Gaia Pigino: Inside the cell

Gaia Pigino studies the molecular mechanisms and principles of self-organization in cilia using 3D cryo-EM.

Gaia Pigino was only 3 yr old when she became fascinated with nature in the beautiful countryside of Siena, Italy, where she grew up. The neighbor’s daughter showed her a hen in the chicken coop, and they caught it in the act of laying an egg. Gaia remembers, “This was for me almost a shock, as my experience about eggs was that they come directly out of paper boxes!” Her father was also an important part of awakening Gaia’s curiosity for the amazing things in nature. He used to bring home the award-winning magazine Airone, the Italian equivalent of National Geographic. Gaia never missed an issue; even before learning to read, she could spend hours looking at the captivating photos of the wildlife. She wanted to understand what she was seeing, and maybe because of that, she was determined to do science.Gaia Pigino. Photo courtesy of Human Technopole.Gaia took her first “scientific” steps with Professor Fabio Bernini and Professor Claudio Leonzio at the University of Siena, where she studied bioindicators of soil contamination and detoxification strategies of soil arthropods as part of her PhD project. But it was later, when she joined the laboratory of Professor Pietro Lupetti and met Professor Joel Rosenbaum, a pioneer of cilia research, that Gaia discovered the world of 3D EM and felt her place was “inside a single cell.” She solidified her interest in the structure of protein complexes of cilia and flagella and boosted her passion for cryo-electron tomography (ET) in the laboratory of Professor Takashi Ishikawa, first at the ETH Zurich and then at the Paul Scherrer Institut in Switzerland. In 2012, Gaia started her own laboratory at the Max Planck Institute of Molecular Cell Biology and Genetics in Dresden, Germany, with the vision of creating a truly interdisciplinary laboratory. Her team combines techniques from different fields such as biophysics, cell biology, and structural biology to answer open questions in the cilia field. Gaia recently moved countries again—this time to take over the position of Associate Head of the Structural Biology Research Centre, at the Human Technopole, Milan, Italy.We reached out to Gaia to learn more about her scientific journey and future research directions.What interested you about cilia?The first thing that attracted me toward cilia and flagella were some EM micrographs, by Professor Romano Dallai in Siena, that showed the beautiful geometrical microtubular structures of sperm flagella. I was intrigued by the apparent perfection of these organelles that clearly showed me that a cell is a coordinated system of complex molecular machines, the mechanism of many of which we do not understand. Soon after, Professor Joel Rosenbaum introduced me to the bidirectional transport of components inside cilia, which, he explained to me, is required for both assembly and function of virtually all cilia and flagella, from the motile cilia in our lungs to the primary cilium in our kidneys. He called it intraflagellar transport (IFT) and compared it to a Paternoster elevator, where the individual cabins were what we now call IFT trains. I was completely fascinated by the IFT system, the structure, the function, the dynamics, and the mechanism of which were still largely unknown. Quickly, I realized that in addition to IFT, cilia represent a virtually infinite source of open biological questions waiting to be solved, from the mechanics and regulation of the beating to the sensory function of primary cilia, and their importance for human health.What are some of the scientific questions currently of interest in your laboratory?In the past few years, we have made substantial contributions to the current understanding of the structure and the mechanism of the IFT (1, 2, 3). Currently, we are investigating how the structure of IFT trains relates to their functions by looking, in cryo-electron tomography, at how anterograde trains transform into retrograde trains and at how different ciliary cargoes are loaded on the trains. Beside this more classical line of research, we are exploring other approaches to study IFT, for instance we have developed a method to reactivate IFT trains in vitro on reconstituted microtubules. We want to use this approach to investigate the behavior of IFT trains, and their motors, in experimentally controllable conditions, e.g., in the presence of only certain tubulin posttranslational modifications. We have also made interesting discoveries about the distribution of tubulin posttranslational modifications on the microtubule doublets of the axoneme and how this spatially defined tubulin code affects the function of different ciliary components. We hope we will be able to share these new “stories” with the structural and cell biology community very soon!What kind of approach do you bring to your work?I believe that the main reason for why science became an integral, and dominant, part of my life is because it provides infinite riddles and continuous challenges. I have always been curious about how things work in nature, but I quickly realized that learning from books didn’t satisfy me. My desire was to be at the frontline, to be among the ones that see things happening in front of their eyes, at the microscope, for the first time. I wanted to be among the ones that make the discoveries that students read about in textbooks. Thus, what I bring to my work is an endless desire of solving biological riddles, curiosity, creativity, determination, and energy, with which I hope to inspire the members of my team. My laboratory uses an interdisciplinary approach; we use whatever method, technique or technology is needed to reach our goal, from the most basic tool to the most sophisticated cryo-electron microscope. And if the method we need does not yet exist, we try to invent it.A young Gaia Pigino (3 yr old) the day she discovered how eggs are made. Photo courtesy of Giancarlo Pigino.Could you tell us a bit about the Structural Biology Research Centre at the Human Technopole (HT)?At the HT Structural Biology Centre, we are working to create a vibrant and interdisciplinary scientific environment that will attract molecular, structural, cell, and computational biologists from all over the world. We are creating fantastic facilities, including one of the most well equipped and advanced electron microscopy facilities in Europe—and likely the world—headed by Paolo Swuec. My team, together with the teams of my colleague Alessandro Vannini and the research group leaders Ana Casañal, Francesca Coscia, and Philipp Erdmann, already cover a vast range of competences and know-how from classical molecular and structural biology approaches, such as crystallography and protein biophysics, to cryo-CLEM, cryo-FIB SEM and cryo-ET, all of which allow us to address questions in cell biology. Our goal is to create a scientific infrastructure and culture that will enable biologists to obtain a continuum of structural and functional information across scales.What did you learn during your PhD and postdoc that helped prepare you for being a group leader? What were you unprepared for?I learned that everyday research is mostly made of failures, but that with the right amount of obsession, persistence, curiosity, and creativity, it is always possible to succeed and discover new things. Being given the freedom to develop your own ideas and your own project very early in your career is a treat; science is not only about having good ideas! One needs to follow up on these ideas with intense work and troubleshooting to make them reality. In addition, I realized that being fearless and attempting what is considered too difficult by others, despite challenges, can turn into a worthy learning experience. Also, how you present your work to the scientific community matters for swinging the odds of success in your favor. Different places might work in very different ways, and conducting good science does not only depend on you, but also on the possibilities given to you by your environment.What was I unprepared for?—I guess several things, but one comes immediately to mind: I underestimated how much being responsible not only for my own life and career, but also the career of students, postdocs, and others in the laboratory, would affect me personally.Structure of the 96-nm axonemal repeat reconstructed by cryo-ET and subtomogram averaging. Image courtesy of Gonzalo Alvarez Viar, Pigino Lab.What has been the biggest accomplishment in your career so far?This is a tricky question for me... I tend to look into the future more than celebrating the past. I fight to succeed in something, but as soon as I conquer it, I find it less of an achievement than the thing I could conquer next. Nevertheless, I am happy about the discoveries and the papers published together with my students and postdocs (1, 2, 3, 4, 5). I am extremely excited about the fact that after many years of work I am now leading an interdisciplinary laboratory, where we combine techniques from different fields. I am also happy that three times my husband and I were able to move from one world class academic institution to the another to start exciting and fitting jobs and could still live together in the same place. We worked hard for this, but we also got lucky.What has been the biggest challenge in your career so far?I studied French in school; I had almost no exposure to spoken English until the end of my PhD. To avoid having to show my English insufficiencies, I did hide beside the board of my poster at the first international conference I attended in 2004! It took me a while to overcome this barrier and feel confident to express my thoughts and ideas in English.What do you think you would be if you were not a scientist?I had been a good fencer during my youth. I was a member of the Italian National Team between ages 14 and 19 and saw quite a bit of the world, which was cool! When my sporting career failed, due to diabetes, I was torn between art and science. I guess that in a parallel universe, I am a wildlife photographer and a potter specialized in wood kiln firing. [Gaia confesses that she misses “the amazing and addictive adrenaline rush of a good fencing match!”]Any tips for a successful research career?Do not compare your performances to the ones of the people at your career stage; compare yourself with people that are already successful one level higher than you currently are at. For example, if you are a PhD student, ask yourself what in your current performance separates you from being a good postdoc—once a postdoc, what is missing to be a good PI.     

Ye Tian: Surveilling stress to live longer

Ye Tian investigates how mitochondrial stress signaling pathways regulate longevity using C. elegans as a model system.

An avid reader, Ye Tian used to save up her child allowance with the sole purpose of buying science fiction books. Reading and solving mathematical problems were her favorite hobbies; indeed, she liked mathematics so much that she was about to enroll herself as an architecture major but finally chose biotechnology. Ye moved from her hometown in the Northwest of China, Baoji—famous for housing the Zhou dynasty’s bronzeware and being close to the Terracotta Army—to Beijing for her college and graduate studies.Ye is proud of being among the earliest researchers working on Caenorhabditis elegans in her country; for her PhD studies, she joined the lab of Hong Zhang, who at that time has just established the first C. elegans lab in China at the National Institute of Biological Sciences in Beijing. Ye identified epg-2 as an adaptor for cargo recognition during autophagy. In 2010, she crossed the Pacific toward the U.S. West Coast for her postdoctoral training in the aging field with Andrew Dillin, first at the Salk Institute in San Diego and then at the University of California, Berkeley. There, she discovered that mild mitochondrial stress during development in worms rewires their chromatin landscape to establish specific gene expression patterns throughout the lifespan and promote longevity.Ye Tian. Photo courtesy of Ye Tian.Ye came back to China at the end of 2016 to start her own lab at the Institute of Genetics and Developmental Biology of the Chinese Academy of Sciences. Her research team studies mitochondrial stress signaling pathways and their interplay with aging. We chatted with her to learn more about her next scientific plans.What interested you about the interplay between mitochondria and aging?I became interested in mitochondrial biology during my postdoc in Andrew Dillin’s lab. Since the origin of eukaryotic cells, mitochondria have been a driving force of evolution. During reproduction, mitochondria are passed from the mother to the offspring through egg cells and they exhibit a unique inheritance pattern. As essential hubs that dictate cellular metabolism, it is clear now that mitochondria and the nucleus maintain a bidirectional communication. Early life “stressed” mitochondria communicate with the nucleus to induce gene expression changes that are beneficial on longevity and persist throughout the lifespan. The fact that mitochondrial function is crucial to aging fascinated me; I wanted to continue exploring that topic further, and that’s why I established my lab around the question of how mitochondrial surveillance mechanisms regulate the aging process.What are you currently working on? What is up next for you?My research team focuses on the interplay between mitochondrial stress signaling pathways and aging. The first work that my lab published was a project that I started during my postdoc. The Dillin lab reported a phenomenon in which perturbations of mitochondria in neurons induced a mitochondrial stress response in the peripheral tissues and hypothesized that a secreted signal molecule, named after mitokine, is required for the cell non-autonomous regulation (1). The identity of this molecular signal remained elusive for almost ten years until we found that a secreted Wnt ligand, EGL-20, functions as the mitokine to coordinate mitochondrial stress signaling across tissues and promote longevity of the organism (2). We are also interested in how the crosstalk between mitochondria and the nucleus influences lifespan. We found that mitochondrial perturbations alter the nuclear epigenome to induce longevity via the histone deacetylation complex NuRD in response to cellular acetyl-CoA levels, the key metabolite at the entry point of the Krebs cycle (3).Lab group picture; current lab members (2021). Photo courtesy of Ye Tian.Our latest work stemmed from a serendipitous observation that neuronal mitochondrial stress is sensed by and transmitted through the mitochondria in the germline. Intergenerational, maternal inheritance of elevated levels of mitochondrial DNA via the mitokine Wnt/EGL-20, which causes the activation of the mitochondrial unfolded protein response (UPR^mt), provides descendants with a greater tolerance to environmental stress. This makes the offspring live longer (4).Among our short-term scientific plans, we’re determining how mitochondria functions during the aging process at both the genetic and biochemical levels and searching for ways to apply our findings from C. elegans to neurodegenerative disease models in mammals.What kind of approach do you bring to your work?The curiosity about how things work drives me; what I enjoy the most is when I see things happening in front of my eyes and when I figure out why they occur that way. That enthusiasm is what I try to spread to my team every day. In the lab, we rely on C. elegans as our model system and on genetics to dissect complex biological processes like aging. We have also adapted modern biochemical and imaging techniques as well as bioinformatics to complement our genetic studies. I’m a geneticist at heart, and I like to initiate a project with a well-designed genetic screen. The best part is that the screen often leads me to answers I was not expecting, and that’s genuinely inspiring!What did you learn during your PhD and postdoc that helped prepare you for being a group leader? What were you unprepared for?Like most scientists, my research career has gone through ups and downs. I had to change my research project in the last year of my graduate school; that was nerve-racking, but I eventually managed to redirect my thesis and get exciting results under time pressure, thanks in large to the support of my parents, mentors, and lab mates. That helped me prepare to become a principal investigator; I gained confidence in problem solving, and since I’ve experienced the stress of dealing with last-minute scope changes firsthand, I connect better with my students.I guess, as many other non-native English speakers, I wasn’t prepared for writing grants and papers fluently in English. This issue wasn’t obvious during my graduate and postdoctoral studies, as my mentors were always there for me and proofread and edited my writing. Now I have to stand up for myself. I spend most of my time writing; I’ve improved my writing skills but it’s still an ongoing process.Reconstruction of the nerve system of C. elegans by confocal microscopy. Green corresponds to YFP-labeled neuronal specific marker Q40, and red labels germline specific mitochondrial outer membrane protein TOMM-20::mkate2. Image courtesy of Ye Tian’s lab.What has been the biggest accomplishment in your career so far?My very first PhD student, Qian Zhang, graduated with two first-author papers and decided to pursue a research career in academia. Being responsible for someone else’s career is challenging but also rewarding.What has been the biggest challenge in your career so far?I use the model organism C. elegans for my research in aging, so from time to time, peers criticize the relevance of my work to human health. I’m used to justifying my scientific approach to funding agencies and peers in other fields, but sometimes it’s exhausting or not pleasant.Who were your key influences early in your career?My PhD mentor, Hong Zhang. He is very passionate about the science he does, and he is courageous to shift his research directions to answer new biological questions.What is the best advice you have been given?I think the best advice I’ve gotten is that “tomorrow is another day.” It reminds me to keep going and be optimistic.What hobbies do you have?I love art and music. When I was in San Diego, I used to play in the Chinese Music Band; I miss my musician friends over there. In my teens, I used to hike mountainside trails along the river with my parents. Now, running has become my new favorite hobby. I enjoy the tranquility and peace of mind while running; it’s soothing.     

Elda Grabocka: Stress-buffering comes in granules

Elda Grabocka investigates the role of stress granules in obesity and cancer.

When one thinks of high school, sharing hallways with students from 80 different countries is not the usual image that springs to mind. This was indeed Elda Grabocka’s experience. She grew up in Pogradec, a remote town in Albania—her parents, both physicians, were assigned to this location by the state. Elda won one of the two spots available for Albanian students in a national competition to attend the United World College of the Adriatic in Trieste, Italy, a high school focused on social change that brings together students from around the globe to promote intercultural understanding. Elda still remembers, with a smile on her face, the first glimpse at the laboratories as the senior students were working on their thesis projects: “That was exactly what I wanted to do!” She barely spoke English at the time and had to catch up to the level of her peers, but her perseverance and passion prevailed, and she obtained the International Baccalaureate Diploma (IBD). For the independent study of the IBD program, she submitted a research project in chemistry, which ended up being an important learning and life lesson: “That helped me understand that I was more suited to biology! In hindsight, it was great to have that experience so early; I certainly had no awareness then how essential failing and then learning from your failures is to science, but having a level of comfort with it from the beginning was probably a bonus.”Elda Grabocka. Photo courtesy of Chris Hamilton Photography.But science was not the only professional option Elda contemplated—her volunteering experience with relief organizations in various refugee camps made her consider a career in public health and humanitarian relief efforts. She finally sought a PhD degree in molecular pharmacology and structural biology in the laboratory of Phil Wedegaertner at Thomas Jefferson University. After studying heterotrimeric G-proteins and how the subcellular localization of their exchange factors regulates function, Elda felt the need to seek greener pastures. She went on to do a postdoc on one of longest-studied oncogenes, RAS—her choice wasn’t motivated by the field, but by the mentor, Dafna Bar-Sagi. Elda’s admiration for Dafna is notable when she speaks about her time at the New York University Langone Medical Center: “It’s remarkable how many novel aspects of RAS biology that have shaped and then re-shaped our thinking about this oncogene have come out of her lab; I felt there was a depth and breadth to her approach to scientific research that if I could learn, I’d be able to see more of the angles, so to speak, ask better questions; she has really expanded my mind in all those aspects.” Elda’s work focused on the interplay between the mutated forms of RAS and the wild-type isoforms, which she and others have shown is context dependent, with the wild-type isoforms acting as both tumor suppressors and tumor promoters (1). While still in Dafna’s laboratory, Elda pursued a more independent scientific interest: the role of stress granules in mutant KRAS cells. In 2016, Elda returned to her alma mater, joining the Department of Cancer Biology at the Sidney Kimmel Cancer Center at Jefferson as an assistant professor, with stress granules in cancer as the focus of her laboratory. We contacted her to learn more about her research journey.What interested you about stress granules and their connection with obesity and cancer?I became interested in stress granules and their potential role in cancer early in my postdoc. I read a review by Stephen Elledge’s group where they described the “stress phenotype” of cancer as an important player in tumorigenesis. I realized that cancer cells exist mostly in a state of stress—for example, mutated genes, like oncogenic RAS, are potent inducers of many types of cellular stresses. I was working on a RAS ubiquitination project, and one of the candidates for a RAS de-ubiquitinating enzyme we were looking at was implicated in stress granule formation. Little was known about stress granules at the time—they are induced by types of stresses associated with tumors (hypoxia, oxidative stress, osmotic pressure, proteotoxic stress, metabolic stress, etc.), so the question I asked was whether stress granules could function as a stress coping/adaptation mechanism in cancer. Indeed, I found that stress granules are prevalent in tissues from patients with pancreatic cancer and mouse models of pancreatic cancer. Remarkably, not all cancer cells are the same in their capacity to form stress granules—all cells will make stress granules under stress, but KRAS mutant cancer cells have a heightened ability to do so because signaling from mutant KRAS enhances the levels of a critical molecule to stress granule formation, 15-deoxy-prostaglandin J2 (2). This enhanced capacity to make stress granules, in turn, renders KRAS mutant cells more resistant to stress and more dependent on stress granules; inhibition of stress granules leads to increased cell death in KRAS mutant versus KRAS wild-type cancer cells.Immunofluorescence staining of pancreatic ductal adenocarcinoma tissue showing cancer cells in red, stress granules in green, and nuclei in blue. Image courtesy of the Grabocka laboratory.The work establishing this dependence was in vitro, so the primary goal when I started my laboratory was to determine their relevance in tumorigenesis, which led me to explore their connection to obesity and cancer for several reasons. First, obesity is a major predisposing factor for several cancers, including pancreatic and colon, which are prevalent KRAS-driven cancers for which treatment options are limited. Second, obesity is a complex pathology which likely impacts the pathobiology, the therapy response, and even the evolution of cancers that arise in this setting. Given that cell stress and inflammation are key features in obesity, this would make the ideal background to study the contribution of stress granules in tumorigenesis. I think this pre-existing stress [obesity] might necessitate the engagement of stress adaptive mechanisms from the early stages of tumorigenesis and may also lead to a high dependence on these processes.What are you currently working on, and what is up next for you?It’s a very exciting time to be working on stress granules! The field has grown significantly over the past 10 yr or so, especially with the renewed interest in phase separation. As organelles that form via phase separation when a cell is under stress, stress granules are perhaps one of the best examples of phase separation in vivo and a great platform to understand its relevance. The recent advances in defining the composition, as well as key molecular drivers and their functional domains in stress granule assembly, have been of great benefit. We are now better positioned to define the stress granule–specific functions in health and disease. Because stress granules are induced by various types of stresses, they could function as a pan-stress adaptation mechanism in cancer. This is a very appealing angle, as if we can solve how stress granules enable stress adaptation, which is a major focus of my laboratory, we could have better anti-cancer therapies.The composition of stress granules, comprising hundreds of proteins and mRNAs involved in several aspects of cell biology, prompted me to ask whether cytoprotection under stress is their main and/or only function. What other cellular processes stress granules regulate, whether these vary with the type of stress, and how such processes are integrated into the stress response of cancer cells are burning questions we are currently working on, as the answers will advance our understanding of the role of stress granules in cancer. The “chronic stress” of cancer is heterogenous in both spatial and temporal terms, as well as in the type of stress and intensity. I am also very curious to see if and how heterogeneity in stress stimuli impact the composition of stress granules and the processes they regulate, and how this may affect tumor evolution. Also, cancer cells are not the only cells in the tumor that make stress granules. As a matter of fact, we reported that KRAS mutant cells can stimulate stress granule formation in a paracrine manner. An ongoing project in the laboratory that I’m very excited about is focused on understanding the contribution of stress granules to the pro-tumorigenic microenvironment.What kind of approach do you bring to your work?My approach is very hypothesis and observation driven; the latter in the sense that it can often be that initial spark that inspires an idea, draws connections, and looks for context and meaning. I also find that sometimes the answer to my next question or the question I don’t know to ask yet is hidden right in front of my eyes, so paying careful attention to the data is key. It is also where objective and critical evaluation of experimental results starts. There’s one line that’s firmly ingrained in my mind from my postdoctoral training, which is “Science is self-correcting.” It’s a note of caution that if you don’t pay attention and see only what you want to see, it will still eventually prove you wrong, and you’d have wasted a lot of time in the process. So I try to minimize that waste as much as possible—unavoidable entirely, having a favorite hypothesis is part of the scientific thinking process, but crucial to remember to follow the data and not just convince yourself.What has been the biggest accomplishment in your career so far?I’m still quite early in my career to start listing accomplishments. I feel privileged to do the work I do; I essentially get funded to pursue ideas that I find interesting. So I have a hard time with this question because it has a hint of pride, and when you start adding pride to privilege, as a junior principal investigator especially, it gets a bit too self-serving. I hope that the work we are doing stands the test of time and leads to or helps lead to a meaningful impact on patients’ lives—that would be a great accomplishment.What has been the biggest challenge in your career so far?The past two years of COVID have certainly been a different reality, and a constantly shifting one at that. From a career perspective, so much of a scientific career happens at the bench: experiments happen at the bench, we train at the bench, animal work is long and requires multiple dedicated essential personnel and facilities, so inevitably, remote work, or shift work, limited occupancy, and the shortages we are now seeing in the supply chain have been a major challenge for everyone. I do think junior laboratories like mine experience that a bit harder. The bandwidth to absorb these challenges is much smaller if you’re just starting out, or if you’ve had a laboratory for a couple of years and are just ramping up. I must say though that it has made for stronger teamwork in the laboratory, and we’ve had to be really focused and efficient—so there’s an upside!Out for a paddle. Photo courtesy of Elda Grabocka.Any tips for a successful research career?Hard to say, because certainly it means different things to different people. The only tip I would give perhaps is to define what that means, what that success looks like for oneself, and be true to that. I expect how each one defines it also changes with time and experience, but I do think it’s very important to identify what success means as early as possible and let that be what you measure your efforts against. It’s easy to get distracted, overwhelmed, or even disheartened otherwise. My own definition is quite simple: success is doing what I love to do, working toward answering a meaningful scientific question, and enabling/supporting my trainees to reach their potential—keeping that in mind has been very important and helpful.     

Dr. Manners

Good manners make a difference—in science and elsewhere. This includes our social media etiquette as researchers. Subject Categories: S&S: History & Philosophy of Science, Methods & Resources, S&S: Ethics

Elbows off the table, please. Don’t chew with your mouth open. Don’t blow your nose at the table. Don’t put your feet up on the chair or table. And please, do not yuck my yum. These are basic table manners that have come up at some of our lab meals, and I have often wondered if it was my job to teach my trainees social graces. A good fellow scientist and friend of mine once told me it was absolutely our place as mentors to teach our trainees not only how to do science well, but also how to be well‐mannered humans. While these Emily Post‐approved table manners might seem old‐fashioned (I’m guessing some readers will have to look up Emily Post), I strongly believe they still hold a place in modern society; being in good company never goes out of style.Speaking of modern society: upon encouragement by several of my scientist friends, I joined Twitter in 2016. My motivation was mainly to hear about pre‐prints and publications, conference announcements and relevant news, science or otherwise. I also follow people who just make me laugh (I highly recommend @ConanOBrien or @dog_rates). I (re)tweet job openings, conference announcements, and interesting new data. Occasionally, I post photos from conferences, or random science‐related art. I also appreciate the sense of community that social media brings to the table. However, social media is a venue where I have also seen manners go to die. Rapidly.It is really shocking to read what some people feel perfectly comfortable tweeting. While most of us can agree that foul language and highly offensive opinions are generally considered distasteful, there are other, subtler but nonetheless equally—if not more—cringe‐worthy offenses online when I am fairly certain these people would never utter such words in real life. In the era of pandemic, the existence of people tweeting about not being able to eat at their favorite restaurant or travel to some destination holiday because of lockdown shows an egregious lack of self‐awareness. Sure it sucks to cancel a wedding due to COVID‐19, but do you need to moan to your followers—most of whom are likely total strangers—about it while other people have lost their jobs? If I had a nickel for every first‐world complaint I have seen on Twitter, I’d have retired a long time ago; although to be honest, I would do science for free. However, these examples pale in comparison with another type of tweeter: Reader, I submit to you, “the Humblebragger.”From the MacMillan Buzzword dictionary (via Google): a humblebrag is “a statement in which you pretend to be modest but which you are really using as a way of telling people about your success or achievements.” I would further translate this definition to indicate that humblebraggers are starved for attention. After joining Twitter, I quickly found many people using social media to announce how “humble and honored” they are for receiving grant or prize X, Y, or Z. In general, these are junior faculty who have perhaps not acquired the self‐awareness more senior scientists have. Perhaps the most off‐putting posts I have seen are from people who post photos of their NIH application priority scores right after study section, or their Notice of Awards (NOA). When did we ever, before social media, send little notes to each other—let alone to complete strangers—announcing our priority scores or NOAs? (Spoiler: NEVER)Some of you reading this opinion piece might have humblebragged at one or time or another, and might not understand why it is distasteful. Please let me explain. For every person who gets a fundable score, there are dozens more people who do not, and they are sad (I speak from many years of experience). While said fundable‐score person might be by someone we like—and I absolutely, positively wish them well—there are many more people who will feel lousy because they did not get funding from the same review round. When has anyone ever felt good about other people getting something that they, too, desire? I think as children, none of us liked the kid on the playground who ran around with the best new Toy of the Season. As adults, do we feel differently? Along these lines, I have never been a fan of “best poster/talk/abstract” prizes. Trainees should not be striving for these fleeting recognitions and should focus on doing the best science for Science’s sake; I really believe this competition process sets people up for life in a negative way—there, I’ve said it.Can your friends and colleagues tweet about your honors? Sure, why not, and by all means please let your well‐wishers honor you, and do thank them and graciously congratulate your trainees or colleagues for helping you to get there. But to post things yourself? Please. Don’t be surprised if you have been muted by many of your followers.It is notable that many of our most decorated scientists are not on Twitter, or at least never tweet about their accomplishments. I do not recall ever seeing a single Nobel laureate announce how humbled and honored they are about their prize. Of course, I might be wrong, but I am willing to bet the numbers are much lower than what I have observed for junior faculty. True humility will never be demonstrated by announcing your achievements to your social media followers, and I believe humblebragging reveals insecurity more than anything. I hope that many more of us can follow the lead of our top scientists both in creativity, rigor, and social media politeness.     

A Tale of Mother and Daughter

Loving science and nature and being a scientist can be very different, yet the two are so intertwined in a scientist''s life that you will certainly experience both aspects. This essay presents my perspective on how, as one who loves science and nature, I came to fall in love with centrosome behavior in stem cells and how I came to run a lab as a scientist. When I started, there was a big gap between my love for science and my experience as a scientist. I filled this gap by learning a “laid-back confidence.”Before the beauty of cell biology (or whatever you love), who you are (i.e., your age, gender, or race) is immaterial. Yet history shows that the ease with which you can pursue science is influenced by who you are. This has certainly been my experience. The key is to find a way to fill in the gap between who you are and what you are (i.e., a scientist), a goal in which we must all support each other. It is my hope that this essay will convey something helpful to those who are at early stages of their career and might be encountering obstacles because of who they are.     

Spinning plates and juggling balls

A PhD thesis is a project with an established goal and a deadline. As such, the tools, strategies and insight of professional project management can be used effectively to improve both research success and personal well-being.A project is a “temporary endeavour undertaken to create a unique product, service or result” [1]. Although this is a generic definition, it pretty much describes any PhD research project. There are many ways to manage a project effectively and efficiently. Unfortunately, most of us are so busy with our science that we forget about the importance of planning and management to our own success, sanity and health. Instead, we approach our first three years of genuine scientific endeavour wide-eyed and unprepared to juggle the hundreds of tiny balls that make up a PhD. Several techniques from the realm of ‘project management'' might therefore be helpful for PhD students who need to plan and manage the many competing demands that doctoral research can place on them.A PhD comprises both the research itself and the acquisition of skills and knowledge that will facilitate your future career. As such, it is of paramount importance to establish your own objectives early on. For example, alongside dividing your project into work packages—smaller projects that might be discrete or might build on each other—it is also essential to define which so-called transferable skills—additional knowledge and experience that might improve your job prospects—you feel will be of greatest use to you, depending on what you want to do after your PhD. The importance of these skills is becoming more widely recognized and taken far more seriously, and you should find that your supervisor is willing to give you the time to pursue them—your institute or university usually requires that he or she does so. More importantly, you should give yourself the time to invest in these skills, as they are going to be vital to everything you do once your PhD project is over.Doctoral research requires a multitude of skills, most of which you will inevitably lack when you commence your PhD programme. The first step is to identify the gaps in your knowledge to plan what skills on which to focus. This will allow you to acquire them in good time, either through professional activities—shadowing a postdoc, teaching undergraduates, joining journal clubs and blogging—or through both internal and external courses and workshops to improve communication, presentation, writing, networking and other skills. In addition to your planned skills acquisitions, you will also have situations arise, in which you need to acquire new skills quickly. The more you plan training activities and skills acquisition in advance, however, the smoother this aspect will be of your PhD. By way of example, part of my own PhD project relates to statistical analysis of data. An early analysis highlighted several areas in which I had to improve my skills, including hierarchical cluster analysis, principal component analysis and χ² testing against standard distributions. Having identified these gaps in knowledge early on in my doctoral programme, I could plan ahead accordingly when and how to acquire these skills.The full scope of your PhD project is usually unknown at the outset, and even the direction of your research might well change before you are finished. ‘Rolling wave planning'' is a technique that allows you to take these facts into account and plan the short-term future in detail, with a high-level provision for medium- to long-term activities. For those new to developing project schedules, I advocate a simple five-step approach. First, make an ordered list of high-level activities needed to achieve your goal. Second, expand this list by adding lower-level activities for which you have a detailed understanding of the scope, for example work to be performed in the next six months. You now have a work breakdown structure. Third, turn this work breakdown structure into a dependency-driven list by adding associations between the activities, for example by adding links to precursor activities that need to be completed before another activity can be started. Fourth, estimate the duration of each activity and extrapolate the start and end dates beginning with the first scheduled activity. Finally, as you progress through your research, and the scope of future activities becomes clearer, update the project schedule with these low-level activities as they become known. This approach of generating a hybrid-level project schedule, and updating with detailed activities as the scope becomes clearer, is known as ‘rolling wave planning''.…we approach our first three years of genuine scientific endeavour wide-eyed and unprepared to juggle the hundreds of tiny balls that make up a PhDThere is a range of professional software to help develop project schedules, but there are also various freeware tools available. Alternatively, you can use one of the many word processing or spreadsheet applications to make a simple Gantt chart. Along with the technical scope of your doctoral research, it would also be useful to include milestones that your institution enforces; for example literature review submission, formal progress reports and thesis chapter outlines. Including these in your rolling wave planning will allow you to keep in mind the bigger picture and the formal aspects that must be completed for your PhD, in parallel with the progress you are making towards your specific research subject.It is of course a cliché, but it is true that ‘failing to plan is planning to fail''. Of course the fluid nature of research makes it difficult to estimate accurately the time that it will take to complete various experiments, especially as a novice researcher. I therefore believe that although experiments do overrun and PhD projects can change, developing a project schedule is not a futile activity. By having a plan, even if it is made up of ‘guesstimates'', you can forecast roughly how much time you have left for your research and roughly what you can realistically hope to achieve. After all, without a plan, how can you predict when you will complete your research, submit your thesis and ultimately gain your PhD?Doctoral research requires a multitude of skills, most of which you will inevitably lack when you commence your PhD programmeThe serious consideration of scope is necessary in any project, but even more when you are simultaneously project manager, research scientist and key stakeholder. This raises various crucial questions regarding scope management: what is my doctoral research all about (the goal), and what work do I need to do to meet this goal? Once this has been agreed between you and your supervisor(s), it is essential to manage the scope of your project—the breadth and number of experiments you will perform—and how this will achieve your goal(s). Furthermore, be specific—knowing exactly what you want to achieve will keep you motivated until you get there.Project managers often use the concept of the triple constraint to manage work: scope, time and cost are intricately linked in a project and the different level of focus that each is given affects the perceived quality (by others) of project deliverables (Fig 1). Project managers understand that any deviation in one of the triple constraints changes one or both of the others. This is where the project schedule really comes into its own by allowing you to forecast when you will complete the agreed goals of your PhD project. For example, is your doctoral programme for a fixed-term period? If so, then once a project schedule has been agreed that uses all of the time available, any project overruns will cause an overrun to the overall PhD. The two main possibilities for a PhD student to manage this situation and bring the projected completion back into acceptable timescales are either to work longer or to reduce the scope or goals of the project, either by conducting fewer experiments to answer the same question or by modifying the depth of the question being asked. This leads to the issue of whether there is a minimum set of goals that need to be achieved, or whether several agreed activities are ‘nice to haves'', but are not crucial for the overall PhD. I believe that your supervisor(s) are best suited to answer questions about the minimum goals and the scope needed to achieve them.Open in a separate windowFigure 1The project management triangle as applied to a PhD. Three competing constraints influence project management: time, scope and cost. The time constraint reinforces that projects are temporary endeavours, and that in most cases have defined timescales (absolute deadlines). The cost constraint refers to the budgeted amount allocated to the project that, from the perspective of doctoral research students, will predominantly be focused on the amount and duration of the stipend awarded, but might also incorporate various expenses such as bench fees, conference fees and consumables. For those changing career, the cost might also comprise an element of salary sacrifice. The scope constraint refers to what must be done, produced or developed to meet the objective of the project, which in the case of a PhD generally comprises the actual doctoral research to be performed, development (and submission) of the thesis, publication of one or more journal articles, presentation at conferences and potentially teaching. The triple constraint principle highlights that any change to one of the constraints will have an impact on one or both of the other constraints. For example, increased scope typically leads to increased time and cost; tight time constraints usually mean that an overrun in activities (such as experimentation) might have a knock-on effect of requiring the scope to be reduced to submit your thesis on time, or increasing the overall amount of time required to complete your PhD. Similarly, a tight budget could mean you cannot gain access to various resources, resulting in either increased time or a reduction in scope. Recently, a fourth component of the project management triangle has been introduced highlighting that along with the three constraints competing with each other, they also interact to form a fourth dimension of quality.If you need to complete your doctoral programme within a specified time frame, then you need to manage your goals and scope mercilessly—do not allow additional research questions or extra experiments to take away precious time. This does not mean that you cannot deviate, but any deviations need to be managed. Remember, whether you wish to remain in scientific research or not, the PhD is a stepping-stone to your future career and not the end goal in itself. Once you have achieved the goals agreed with your supervisor, it is more beneficial for you to write-up your doctoral thesis and move on [2].Good communication is essential in every area of work, but even more so for a PhD as you are simultaneously learning how to research along with doing the research. Often, access to your supervisor is limited by constraints on his/her or your time, which means that clear communication is vital. Do not assume that your supervisor knows every intricate detail of what you are doing; he or she might have a large group in which each member is looking at complementary aspects of a more general topic. It is, therefore, your responsibility to ensure that all your stakeholders—supervisors, postdoc leads and any others involved—know what you are doing and, more importantly, why you are doing it.This is another area in which the apt use of technology can maximize efficiency. Subject to institutional licensing, collaboration tools such as SkillsForge or Evernote can improve communication between stakeholders. For example, meeting minutes, action points to be followed and research results can be uploaded for sharing. Supervisors can then review the material at a convenient time to ensure that they stay up to date with the progress of each student within their research group.As PhD students usually aspire to become research scientists, it is of paramount importance that you learn the correct application of the scientific method and the context in which your work is being done. Before jumping into practical work—wet-lab experiments or computational modelling—it is important to understand the meaning and relevance of your project in relation to existing knowledge and the underlying science. For example, the hypothesis-driven research life cycle in systems biology [3,4]—my own field—advises that computational models should be developed on the basis of wet-lab data relating to the underlying biological system. Almeida-Souza and Baets state that a PhD in science is an opportunity to learn how to tackle problems scientifically and, as such, requires the development of skills in critical thinking, hypothesis formulation and experimental design [5]. I believe that the requirement for these skills is universal across the sciences, and that molecular biosciences and computational systems biology are no different.The serious consideration of scope is necessary in any project, but even more when you are simultaneously project manager, research scientist and key stakeholderTherefore, before the first wet-lab experiment is performed, or the first line of code is written, it is essential that we understand why the experiment is important and what results we might expect to support our initial hypotheses. Furthermore, regarding computational systems biology, I believe that it is also essential for wet-lab and computational researchers to collaborate to ensure both have a consistent understanding of the data and the purpose of the computational model. After all, for the most part, computational models are developed for their predictive capacities and to allow hypothesis generation for subsequent wet-lab experimentation. Baxter et al have extensively covered this area and advocate not only designing the project up-front, but also the need for quality control [6].You need to manage the scope and goals of your PhD mercilessly and, at the same time, be flexible enough to grasp new opportunities. Conversations at conferences, for instance, can open up opportunities for collaboration and take your research in a direction that you had not considered previously. In my case, I was invited to turn a conference paper relating to my masters degree into a full paper for a special issue of a well-known bioinformatics journal. Although it was not related to my doctoral research, the prospect was too good to turn down. I therefore discussed the idea with my PhD supervisor, and once we were in agreement, I updated the project schedule to incorporate this new activity, trying to mitigate as much as possible the resulting slippages to my doctoral research. In essence, I had performed an impromptu risk–reward analysis and decided that the reward that would be gained from publishing this work outweighed the risk of a slight overrun of my PhD thesis. It must be noted that I was lucky in this instance, as my PhD supervisor also supervised the research project during my master''s degree, so a full paper would be beneficial for both of us.A project risk is “an uncertain event or condition, that if it occurs, has an effect on at least one project objective” [1]. The positive side to risks is that the likelihood of their future occurrence can be mitigated by planning in the present. Once a risk is realized, however, and its effects begin to be felt, it has turned into a project issue. The first step in trying to manage risks is their identification. Risk identification in this context is the process of determining which events, if they occurred, would affect your research. In the context of a molecular biosciences PhD, I believe that general risks relate to access to resources, such as people—postdocs and collaborators, for example—reagents, cell lines and shared equipment. For example, if your work uses fluorescent proteins within single cell analysis, how would you be affected if the fluorescence microscope was booked out by another research lab? Similarly, in computational systems biology, if the design process for your computational model requires access to wet-lab data, what would the effect(s) be if this was not available?Once risks are identified, it is important to develop risk response plans. By using the above example of access to a microscope, what should your response be if you cannot gain access? The initial risk response would be to liaise with the other research lab to understand their requirements and ascertain whether you could gain access at a mutually convenient time. Alternatively, another approach might be to work outside normal office hours, either throughout the evenings or on the weekend, subject to health and safety procedures at your institution and your own health and well-being. I believe that a degree of creativity is often required when developing effective risk response plans.A PhD thesis is a hefty document that might run to many hundreds of pages. They are generally not written as a single large document from start to finish, but as chapters. In the molecular biosciences, a thesis consists of an initial literature review early in the doctoral programme, work-in-progress documents for materials and methods, experimental results throughout the middle section, which is followed by analysis and critical evaluation towards the end of your experimental work. Whether through software tools or through your own manual methods, such as keeping a configuration log and keeping a copy of each version of your working documents, it is essential that you maintain an up-to-date repository of all your notes. I have found through experience that it is beneficial to save not only the final versions, but also each of the working drafts of documents generated throughout your PhD. Ideas previously discounted, and thus removed from more recent versions of documents, might once again take centre stage at a later date.The positive side to risks is that the likelihood of their future occurrence can be mitigated by planning in the presentThis can be aided through the development and use of a project library with a logical folder structure to facilitate easy access to documentation. Noble [7] provides an in-depth discussion of organizing your computational biology project—in particular the value of version control—but the concepts are transferable across disciplines. Furthermore, do not forget to back-up your work, and without seeming too pessimistic, back-up your back-up!Finally, look after the most important resource: you. Exercise, diet, alcohol, caffeine and holidays all affect your well-being. Holidays and time away from the lab or office allow you to take a step back from the detail and reflect on your experiences and progress. Sometimes, time off allows you to process issues subconsciously and develop new approaches to overcome problems that you have been tackling for extended periods of time without success. Finally, holidays also help you recharge your batteries and enthusiasm to return to your project with fresh vigour. If you have sensibly and reasonably planned time off alongside your work, you will be able to enjoy it.Although a PhD requires consistent commitment, you simply cannot—and should not—work at full capacity all of the time. Issues arise periodically throughout any project, and if you have no reserves of energy—either mental or physical—you will be unable to tackle them head on with the step change of performance that is required. Furthermore, doctoral research is a marathon and not a sprint; we all experience the symptoms of burnout from time to time, and sometimes it is better to walk away for a short period to recharge than to carry on, become stale, and ultimately slow down.To conclude, I wish you good luck with your doctoral research, and I hope these techniques help you to manage your PhD project through to successful completion.? Open in a separate windowRichard Alun Williams     

Ten simple rules for finding and selecting R packages

R is an increasingly preferred software environment for data analytics and statistical computing among scientists and practitioners. Packages markedly extend R’s utility and ameliorate inefficient solutions to data science problems. We outline 10 simple rules for finding relevant packages and determining which package is best for your desired use. We begin in Rule 1 with tips on how to consider your purpose, which will guide your search to follow, where, in Rule 2, you’ll learn best practices for finding and collecting options. Rules 3 and 4 will help you navigate packages’ profiles and explore the extent of their online resources, so that you can be confident in the quality of the package you choose and assured that you’ll be able to access support. In Rules 5 and 6, you’ll become familiar with how the R Community evaluates packages and learn how to assess the popularity and utility of packages for yourself. Rules 7 and 8 will teach you how to investigate and track package development processes, so you can further evaluate their merit. We end in Rules 9 and 10 with more hands-on approaches, which involve digging into package code.     

Work life balance?

There is no perfect recipe to balance work and life in academic research. Everyone has to find their own optimal balance to derive fulfilment from life and work. Subject Categories: S&S: Careers & Training

A few years ago, a colleague came into my office, looking a little irate, and said, “I just interviewed a prospective student, and the first question was, ‘how is work‐life balance here?’”. Said colleague then explained how this question was one of his triggers. Actually, this sentiment isn''t unusual among many PIs. And, yet, asking about one''s expected workload is a fair question. While some applicants are actually coached to ask it at interviews, I think that many younger scientists have genuine concerns about whether or not they will have enough time away from the bench in order to have a life outside of work.In a nutshell, I believe there is no one‐size‐fits‐all definition of work–life balance (WLB). I also think WLB takes different forms depending on one''s career stage. As a new graduate student, I didn''t exactly burn the midnight oil; it took me a couple of years to get my bench groove on, but once I did, I worked a lot and hard. I also worked on weekends and holidays, because I wanted answers to the questions I had, whether it was the outcome of a bacterial transformation or the result from a big animal experiment. As a post‐doc, I worked similarly hard although I may have actually spent fewer hours at the bench because I just got more efficient and because I read a lot at home and on the six train. But I also knew that I had to do as much as I could to get a job in NYC where my husband was already a faculty member. The pressure was high, and the stress was intense. If you ask people who knew me at the time, they can confirm I was also about 30 pounds lighter than I am now (for what it''s worth, I was far from emaciated!).As an assistant professor, I still worked a lot at the bench in addition to training students and writing grant applications (it took me three‐plus years and many tears to get my first grant). As science started to progress, work got even busier, but in a good way. By no means did I necessarily work harder than those around me—in fact, I know I could have worked even more. And I’m not going to lie, there can be a lot of guilt associated with not working as much as your neighbor.My example is only one of millions, and there is no general manual on how to handle WLB. Everyone has their own optimal balance they have to figure out. People with children or other dependents are particularly challenged; as someone without kids, I cannot even fathom how tough it must be. Even with some institutions providing child care or for those lucky enough to have family take care of children, juggling home life with “lab life” can create exceptional levels of stress. What I have observed over the years is that trainees and colleagues with children become ridiculously efficient; they are truly remarkable. One of my most accomplished trainees had two children, while she was a post‐doc and she is a force to be reckoned with—although no longer in my laboratory, she still is a tour de force at work, no less with child number three just delivered! I think recruiters should view candidates with families as well—if not better—equipped to multi‐task and get the job done.There are so many paths one can take in life, and there is no single, “correct” choice. If I had to define WLB, I would say it is whatever one needs to do in order to get the work done to one''s satisfaction. For some people, putting in long days and nights might be what is needed. Does someone who puts in more hours necessarily do better than one who doesn''t, or does a childless scientist produce more results than one with kids? Absolutely not. People also have different goals in life: Some are literally “wedded” to their work, while others put much more emphasis on spending time with their families and see their children grow up. Importantly, these goals are not set in stone and can fluctuate throughout one''s life. Someone recently said to me that there can be periods of intense vertical growth where “balance” is not called for, and other times in life where it is important and needed. I believe this sentiment eloquently sums up most of our lives.Now that I''m a graying, privileged professor, I have started to prioritize other areas of life, in particular, my health. I go running regularly (well, maybe jog very slowly), which takes a lot of time but it is important for me to stay healthy. Pre‐pandemic, I made plans to visit more people in person as life is too short not to see family and friends. In many ways, having acquired the skills to work more efficiently after many years in the laboratory and office, along with giving myself more time for my health, has freed up my mind to think of science differently, perhaps more creatively. It seems no matter how much I think I’m tipping the balance toward life, work still creeps in, and that’s perfectly OK. At the end of the day, my work is my life, gladly, so I no longer worry about how much I work, nor do I worry about how much time I spend away from it. If you, too, accomplish your goals and derive fulfillment from your work and your life, neither should you.     

Alcodemia: are we training our students to be great thinkers or great drinkers?

Academia has fostered an unhealthy relationship with alcohol that has an undeniable impact on the health and behaviour of students and staff. Subject Categories: S&S: History & Philosophy of Science, Chemical Biology, S&S: Ethics

University life has a lot to offer. And, for better or worse, much of it goes hand in hand with a bottle. Believe it or not, I was a bit of teetotaler in my undergraduate days but quickly made up for it in graduate school, where each celebration included inebriation. Indeed, my initial tour of the laboratory I eventually worked in included a refreshing visit to the grad club. Orientation week ended with a marathon beer blitz at a nightclub. The semester’s first invited seminar speaker was welcomed with the sounds of loose change, ice buckets and the clickity‐clack of organic microbrews being opened. Our inaugural genome evolution journal club was such a success that we vowed to spill even more red wine onto our notebooks the following week. In hindsight, I should have realized at this early stage in my studies that I was fostering an unhealthy and unsustainable relationship between biology and booze. Unfortunately, my post‐graduate education in alcohol didn’t stop there.Like many keen students, I arrived at my first scientific conference with a belly full of nerves and a fistful of drink tickets, which I quickly put to good use at the poster session. The successful completion of my PhD proposal assessment was met with pats on the back as I was swiftly marched off to a local pub with no chance of escape. My first peer‐reviewed paper literally arrived with a pop as Champagne was generously poured into plastic cups for the entire laboratory group. My failures, too, were greeted with a liberal dose of ethanol. “Sorry you came up short on that scholarship application, Smitty. It’s nothing a little weapons‐grade Chianti won’t cure.” “That experiment failed again! Come on, let me buy you a lunchtime martini to make up for it.” Soon I learnt that every academic event, achievement or ailment, no matter how big or small, could be appropriately paired with beer, wine or spirit. Missing from the menu were two crucial ingredients for any burgeoning researcher: moderation and mindfulness.But it was the older vintages that really inspired me – the legendary drinking escapades of my scientific mentors, advisors and idols. The tale of professor so‐and‐so who at that epic meeting in 1993 polished off an entire magnum of rosé at dinner and then went on to deliver among the greatest keynote lectures on record at 9 am the following morning. That celebrated chaired researcher who kept the single malt next to the pipette tips for quick and easy access. The grizzled evolutionary ecologist who never went into the field without half a dozen cans of high‐end smoked oysters and two hip flaks, which didn’t contain water. And so, when I was told by someone in the know of how the most famous geneticist on campus wrote that monumental Nature paper (the one I’d read ten times!) while locked in his office for twelve hours with a six‐pack, I bought into the romance hook, line and sinker. The result: I’ve been nursing a recurring headache for nearly two decades and I’m still waiting on that Nature paper. Most importantly, I now realize the various dangers of romanticizing the bottle, especially for individuals in mentorship positions.Like my idols before me, I’ve accrued a cask full of well‐oaked academic drinking stories, except that they haven’t aged well. There is that heroic evening of intense scotch‐fueled scientific discussion, which led to me forfeiting two front teeth to the concrete sidewalk (my mother still thinks it was a squash accident). Or that time I commemorated the end of a great conference in Barcelona by throwing up on the front window of a café while the most prominent minds in my field sipped aperitifs inside (thank god this was before Twitter). Even more romantic: me buying a bottle of Cotes de Nuits Burgundy at Calgary airport on route to a job interview, discreetly opening the bottle in‐flight because economy class wine sucks, and then being met by airport security upon landing. Let’s just say I didn’t get the job. To some, these anecdotes might seem light‐hearted or silly, but they are actually rather sad and underscore the seriousness of substance abuse. Many readers will have their own complicated experiences with alcohol in academia and, I believe, will agree that it is high time we asked ourselves: are we training our graduate students to be great thinkers or great drinkers? Moreover, this question does not address the equally if not more serious issue of excessive drinking among undergraduate students.As I sit at my desk writing this, I think to myself: is it normal that within a two‐minute walk of my university office there are three different places on campus that I can have a beer before lunch, not including the minifridge behind my desk? Is it normal that in my department the first thing we do after a student defends their thesis is go to the grad club where they can have any alcoholic drink of their choosing for free from the goblet of knowledge, which is kept on a pedestal behind the bar? Is it normal that before the COVID pandemic when I was visiting a prominent university for an invited talk, one of the professors I met with offered me a glass of expensive Japanese gin at 11 am in the morning? (And, yes, I accepted the drink.)Of course, if you don’t want to drink you can just say no. But we are learning more and more how institutional cultures – “the deeply embedded patterns of organisational behaviour and the shared values, assumptions, beliefs or ideologies that members have about their organisation or its work” (Peterson & Spencer, 1991) – can have powerful effects on behaviour. Excessive alcohol consumption is undeniably an aspect of collegial culture, one that is having major impacts on the health and behaviour of students and staff, and one that I’ve been an active participant in for far too long. I’ll be turning forty in a few months and I have to face the fact that I’ve already drunk enough alcohol for two lifetimes, and not one drop of it has made me a better scientist, teacher or mentor. The question remains: how much more juice can I squeeze into this forty‐year‐old pickled lemon? Well, cheers to that.     

The story of science: successes,failures, luck,privilege, and bias

I am incredibly honored to receive the 2021 WICB Junior Award for Excellence in Research in WICB’s golden jubilee year. In this essay, I traverse my scientific journey starting with my PhD, highlighting the highs and the lows and how these intersect with luck, privilege, and bias.

V. AnanthanarayananMy pursuit for a PhD started with a hiccup—I had applied to several places in the United States, but barely got any offers due to the economic upheaval that happened that year (2008). I had to forgo any dreams of a PhD in the United States and remained in Bangalore, India to complete a project I had started with William (Bill) Thies at Microsoft Research India on a programming language for expressing biology protocols. Applying to U.S. schools was an expensive task, one which I was unwilling to put my family through again. So, a year later, when I recommenced my search for a PhD position, I set my sights on Europe. I had heard about the PhD program at the Max Planck Institute of Molecular Cell Biology and Genetics (MPI-CBG ) at Dresden from a friend who had just joined the institute for her PhD. Fortunately, I received an interview call from MPI-CBG. At the end of a crucial interview week at Dresden, I “matched” with Iva Tolic´’s (now Institut Ruđer Boškovic´, Croatia) lab for my PhD. At the start of my PhD, I knew next to nothing about the cytoskeleton, motor proteins, or microscopy, but I found Iva and my lab members to be some of the warmest and most welcoming people. I made friends for life and graduated with a PhD in Biophysics, with a thesis focused on understanding the regulation of the motor protein cytoplasmic dynein. I was lucky to have been able to get a position at MPI-CBG and join Iva’s lab—of the other three places in Europe I had applied to for a PhD, only one other institute invited me for an interview, which also proved to be unsuccessful.On completing my PhD in 2014, I didn’t quite know what I wanted to do. Due to personal reasons, I had to return to India and was open to options in both industry and academia. But with my training in motor protein and cytoskeleton research, I had some ideas for exploring scientific questions related to dynein activation. However, most labs I approached for a postdoctoral position were not open to a project that was outside the realm of their research focus. Nonetheless, Iva, Nenad Pavin (University of Zagreb), and Jonathon (Joe) Howard (Yale University), who were members of my thesis advisory committee, gave me the courage to continue in academia. In my naïveté, I went ahead and applied for the INSPIRE Faculty Fellowship, which is targeted at fresh PhDs and junior postdoctoral fellows to establish their own independent group at an Indian institute. To my surprise, I ended up getting the fellowship. The next issue was finding a host institute that was preferably in Bangalore, where my partner was based. I applied at a few different places, but only after I attended IndiaBioscience’s Young Investigator Meeting in 2014 did I get the chance to meet representatives of potential host institutes, including the Indian Institute of Science (IISc). After a couple of research seminars at IISc, my application was assessed and I was offered the position of INSPIRE Faculty Fellow at the newly formed Centre for BioSystems Science and Engineering, IISc.While I did not have any additional start-up funding, I was given the infrastructure and the independence to pursue my research program. It was slow and frustrating at the start, not unlike most starting labs. I always wondered if it might have been easier if I had had a regular postdoctoral stint. During this time, I also started recognizing how hard it was to be a woman in Indian academia. As a woman principal investigator, one’s authority, expertise, and ability are constantly called into question. Justifying your presence in academia on a daily basis is an exhausting task. I had a great mentor in Sandhya Visweswariah (IISc) who helped me navigate the system. I also had an extremely supportive partner, who kept me going through some of the worst times. Eventually, my lab and I landed on our feet (more about this in “My INSPIRE’d Journey”). Our research has been recognized with grants and awards, but one of the most rewarding parts of the job is seeing other lab members discovering the joy of science (I wrote about my approach to mentorship recently [https://www.nature.com/articles/s41580-020-0256-6]).Three years into the faculty fellowship, I was able to transition to an Assistant Professor position in the same institute. However, this did not change my experience as a young woman in Indian science, and the implicit and explicit biases continued. In 2020, I accepted a fantastic opportunity to further my lab’s science as an EMBL Australia Group Leader at the Single Molecule Science Node at UNSW Sydney and made the move during a pandemic. My lab’s research focus is in understanding how stochastic and rare events pertaining to cytoskeleton and motor proteins give rise to complexity in intracellular organization. With this theme as the essence of our research, we ask specific questions about motor protein regulation to effect differential cellular trafficking, mitochondria-microtubule interactions, and their role in mitochondrial dynamics, and we aim to determine barcodes of global organelle positioning in health and disease.I have the privilege of being able-bodied, born in an upper middle-class family to college-educated parents who were extremely supportive of my choices. I have also inordinately benefitted from the fact that I was born to an Indian ‘upper caste’ family. I therefore had an undue head start in life. These were circumstances beyond my control and yet played a huge role in how my story turned out. I was embarrassingly ignorant of the rampant misogyny in academia until I had to contend with explicit and implicit gender-based biases myself when I started my independent research group in India. Women make up ∼40% of science PhDs awarded in India but represent only ∼13% of Indian academia (biaswatchindia.com), highlighting the stark gender biases at play in creating a leaky pipeline. While I tried my best to voice my discontent and affect changes to create an equitable environment within my department and institute, it was slow work. In 2020, when the pandemic hit and all conferences and meetings went virtual, conference posters advertised on social media made it immediately apparent just how much women were underrepresented in Indian STEM conferences. So, I teamed up with Shruti Muralidhar (now a scientist at Deep Genomics, Canada) to found BiasWatchIndia, an initiative to document women representation and combat gender-biased panels in Indian STEM conferences.BiasWatchIndia has been in existence for a little over a year now—we have achieved several milestones, but there’s still so much to do. “Manels” (conferences that feature only men) are still as rampant as they were when we first started—40% of all Indian STEM conferences are manels. And while we have just about started to tackle the underrepresentation of women in Indian STEM, we are conscious of the intersectionality of bias with gender, caste, ableism, and socioeconomic background and aim to understand how best we can advocate for all minorities.People who are in power in academia and who oppose equity, diversity, and inclusion initiatives and instead preach merit and equality as the gold standard need to introspect, because when options and opportunities are offered without consideration to the millennia of oppression based on gender, race, and background, it is not promoting equality but upholding values that will continue to oppress underrepresented groups. Still, I am optimistic and hope to see real changes that will result in equity in academia in my lifetime.     

Hongyuan Yang: Tracking lipids,one droplet at a time

Hongyuan Yang investigates lipid trafficking and lipid droplet biogenesis.

Hongyuan Yang grew up in a small city east of Beijing, China. From his childhood, Hongyuan recalls that “food was not abundant, so I was hungry at times, but education was free and good.” Driven by his curiosity for science, after completing his undergraduate studies at Peking University Health Science Center, China, he enrolled at Columbia University, NY, for his doctoral training. Under the guidance of his advisor, Dr. Stephen Sturley, Hongyuan studied lipids in budding yeast. The laboratory’s research department fostered a strong interest in lipids and atherosclerosis, and after earning his PhD, Hongyuan obtained a faculty position at the National University of Singapore (NUS) in 1999. In 2007, he moved to the University of New South Wales (UNSW) in Sydney, Australia, to continue his scientific journey exploring lipids. We contacted Hongyuan to learn more about his career and interests.Hongyuan Robert Yang. Photo courtesy of UNSW.What interested you about lipids?My five-year doctoral study focused entirely on the enzymes Sterol O-Acyltransferases (SOAT, also known as ACAT, Acyl-CoA Cholesterol Acyltransferases), which catalyze the formation of sterol esters from sterols/cholesterol and fatty acyl CoAs (1). SOATs, integral membrane proteins of the ER, are potential therapeutic targets for heart disease and Alzheimer’s disease. Since then, I have been fascinated by two things related to SOAT: first, what happens upstream of SOAT, i.e., how exogenous cholesterol reaches SOAT/ER; and second, what happens downstream of SOAT, i.e., how its product—cholesterol esters—is stored in cells in the form of lipid droplets (LDs).These are fundamental questions in cell biology. While reading on how cholesterol arrives at the ER for esterification by SOAT/ACAT in the late 1990s, I realized that the trafficking of most lipids was poorly characterized with little molecular insight. Significant progress has been made in the last 20 years, but the lack of tools that track the movement of lipids has hampered our understanding of the selectivity, efficacy, and kinetics of lipid trafficking. Few cell biologists cared about LDs ∼20 years ago, even though LDs are prominent cellular structures in many disease conditions. Each LD comprises a hydrophobic core of storage lipids (triglycerides and sterol esters) wrapped by a monolayer of phospholipids. Largely considered inert lipid granules, LDs originate from the ER and are relatively simple cellular structures as compared with other organelles (see image). Now, we know that LDs are not that simple: their biogenesis is tightly regulated, they actively interact with other organelles, and they regulate many aspects of cellular function as well as disease progression. Astonishingly, we still have little understanding of how LDs originate from the ER. I am very much intrigued by the complexity of these two seemingly simple cellular processes, i.e., lipid trafficking and LD biogenesis.What are some of the scientific questions currently of interest in your laboratory?We are currently focusing on how LDs originate from the ER. The first significant paper from my own laboratory was the discovery of seipin as a key regulator of LD formation (2). Results from many groups have demonstrated that seipin can organize the formation of LDs; however, the exact molecular function of seipin remains mysterious. Our data suggest that seipin may directly impact the level and/or distribution of lipids such as phosphatidic acid near sites of LD biogenesis, and the effect of seipin deficiency on LD formation is secondary to changes in local lipids. We are now working hard to test this hypothesis. Moreover, data from my laboratory and others indicated that nonbilayer lipids may have a greater impact on the biogenesis of LDs than that of other ER-derived structures, such as COPII vesicles. This may result from the monolayer nature of the LD surface. We hope to dissect the dynamic changes of lipids at ER domains where LDs are born. More broadly, the ER is a fascinating organelle to me. The simple division of ER into sheets and tubules does not reflect the dynamic nature of this organelle. Dissecting the composition and organization of lipids and proteins of the ER would help answer key questions relating to LD biogenesis, and it is therefore one of our future directions.Another major focus is to understand how cholesterol and phosphatidylserine are moved between organelles. We have been working on how low-density lipoprotein (LDL)–derived cholesterol (LDL-C) reaches the ER for two decades. The release of LDL-C from lysosomes requires the Niemann Pick C1&2 proteins, whose malfunction causes lysosomal cholesterol accumulation and a lethal genetic disorder affecting young children. The Ara Parseghian Medical Research Foundation has led the way in supporting research into cholesterol trafficking, and I take this opportunity to thank their generous support. Once released from lysosomes, LDL-C is believed to reach the plasma membrane first and then the ER. We identified ORP2 as a possible carrier of LDL-C to the plasma membrane using a PI(4,5)P₂ gradient (3). There must be other carriers and/or pathways because ORP2 deficiency only causes a minor accumulation of cholesterol in lysosomes. Another interesting question is what prevents LDL-C from reaching the ER directly from lysosomes, given the close contact between lysosomes and the ER. We reported that ORP5 may bring LDL-C directly to the ER (4). However, it was later found that ORP5 binds and transfers phosphatidylserine, not cholesterol. Thus, our observed link between ORP5 and cholesterol is through some indirect yet unknown mechanism. We have been perplexed by these observations for many years, but a recent study demonstrated that phosphatidylserine is required for the trafficking of LDL-C, establishing a close link between cholesterol and phosphatidylserine (5). We are now trying to understand how the trafficking and distribution of cholesterol, phosphatidylserine, and PI(4,5)P₂ are interconnected. For a long time, I felt that it was impossible to figure out the molecular details governing the cellular trafficking of lipids due to redundant pathways and a lack of tools to track lipids. Recent progress in this field has given me hope.Lipid droplets in a HeLa cell are shown in red (BODIPY), with their surface in green. DAPI (blue) labels DNA. Image courtesy of Hongyuan Yang.What kind of approach do you bring to your work?Besides honesty and open-mindedness, we emphasize rigor and comprehensiveness. We often make our initial discoveries in cell-based screens. This approach has many advantages, but it also gives rise to artifacts and cell-line specific observations. We aim to complement our initial findings with biochemical and structural analyses in vitro as well as animal studies in vivo. To further establish the reproducibility of our data, I often ask my close friends and collaborators to independently repeat the key findings of a study before submission. It generally takes a long time for us to complete a study, but I believe the effort will pay off in the long run.What did you learn during your training that helped prepare you for being a group leader? What were you unprepared for?During my PhD at Columbia, I was most impressed with the general attitude of my mentors toward research. No matter how much they have achieved, they take every new experiment and every poster presentation seriously.As I did not have postdoctoral training, I was somewhat unprepared at the beginning of my independent career. One difficult challenge was knowing when to finish a paper and project. We often kept working and working. I have now gotten a lot better.You’ve done research on three continents throughout your career. Can you tell us about some of these transitions?During the last year of my doctoral studies at Columbia, I was offered a lecturer position by the Department of Biochemistry at NUS. It was a very hard decision to leave the United States, but I was excited by the prospect of starting my own laboratory at a top institution. Life at NUS was very good overall, despite some struggles. I had to make ∼700 slides for teaching during the first year and my start-up fund was 10,000 Singapore dollars (~6,000 USD). But the graduate students were fully supported by the university, and most of them are hard working and talented. The crucial screen that led to the discovery of seipin as a key regulator of LD formation was performed at NUS (2). I enjoyed my time at NUS, where I was promoted and tenured. However, my family and I could not get used to the heat and humidity. We looked for a place with better climate, and it happened that my current employer, UNSW, had an opening in 2006. Moving continents with two kids was very disruptive, and I had zero publications in 2007. Our work on seipin was delayed and almost got scooped. I was also very worried about funding in Australia since I hardly knew anyone and the funding system. It turned out that the Australian community was very supportive of our research from day one. I have also been very fortunate to receive generous support from the Ara Parseghian Medical Research Foundation, based in the United States, after my move to Sydney.Hongyuan’s “metabolism team” after a basketball game. Photo courtesy of Hongyuan Yang.What has been the biggest accomplishment in your career so far?While I am mostly recognized for discovering seipin’s role in lipid droplet formation, I am prouder of the work we have done on lipid trafficking and the oxysterol binding proteins. We struggled mightily for the first 15 years. At one point in 2015, I seriously considered abandoning this line of research. But we persisted and discovered their roles in regulating plasma membrane PI(4,5)P₂ and cholesterol, as well as in lipid droplet formation (3, 6).What has been the biggest challenge in your career so far?The biggest challenge has to do with the subject of my research topic: the fundamental cell biology of lipids. The sorting, distribution, and storage of cellular lipids are clearly very important topics in biology, but they are sometimes too fundamental to explain to funding agencies and new students. These days, lipid research is not as “sexy” as other topics. But there are so many unanswered questions in lipidology. I strongly believe that lipid research is going to be the next “big thing” as new techniques such as cryoEM now allow us to appreciate lipids and membrane proteins with unprecedented clarity.Who were your key influences early in your career?Besides mentors and teachers at Columbia, I really enjoyed reading and studying the works by Drs. Mike Brown and Joseph Goldstein, Ta-Yuan Chang, and Scott Emr. While they were not my teachers, their work inspired and impacted many young scientists, including me.What is the best advice you have been given?I have been given many pieces of great advice during my career. The best one in my view is “Less is more.” I was once told, “You would be better off with a lab of six than twelve.” Initially, I did not get it because I thought that a bigger group would allow me to explore more directions and be more productive. The reality is that, as a little-known junior researcher, few experienced people would join my laboratory. Funding is also a major limiting factor. Supervising a large number of students is fulfilling, but it also takes away some of my own time to think critically about the projects. I have largely kept my group under six, and this allows me to better supervise and guide the trainees. People say, “Once your team has more than 15 members, you become a manager instead of a scientist.” My own experience corroborates that statement because I struggled quite a bit when my group reached 12 at one point.What hobbies do you have?I am heavily into sports, especially basketball and tennis. I follow the NBA closely, and Jeremy Lin is my hero. I still play basketball at least twice a week. I am the captain of a basketball team comprised of scientists working on metabolism (see image). We play real, refereed basketball games against local teams during conferences. As I am getting older, I have also picked up tennis. I watch coaching videos on YouTube but still need a lot of work on my forehand. Through sports, I learned teamwork and the spirit of fighting to the last second. If I were not a scientist, I would probably run a sports-related business.What has been your biggest accomplishment outside of the laboratory?I got married and had children relatively early. Both of my kids are now in college and they appear to be decent human beings. I have been extremely lucky because my wife did most of the heavy lifting in looking after the kids. It was still a struggle for me to balance work and parental duties during the early days of my independent career. I am very proud and happy with where we are as a family right now.Any tips for a successful research career?Everyone is unique. Knowing your strengths and especially your weaknesses can be crucial to your success. My undergraduate training was in medicine and health management, and my PhD work focused on genetics and cell biology, so my understanding of physical chemistry is rather inadequate. I am also very bad at developing new methods. To alleviate these deficiencies, I constantly monitor new methods in my field and I purposefully look for collaborators with strong chemistry backgrounds. I have benefited immensely from such efforts.     

After 2 years of a pandemic,students are struggling to find strong reference letters

Writing and receiving reference letters in the time of COVID. Subject Categories: Careers

“People influence people. Nothing influences people more than a recommendation from a trusted friend. A trusted referral influences people more than the best broadcast message.” —Mark Zuckerberg.

I regularly teach undergraduate courses in genetics and genomics. Sure enough, at the end of each semester, after the final marks have been submitted, my inbox is bombarded with reference letter requests. “Dear Dr. Smith, I was a student in your Advanced Genetics course this past term and would be forever grateful if you would write me a reference for medical school…” I understand how hard it can be to find references, but I have a general rule that I will only write letters of support for individuals that I have interacted with face‐to‐face on at least a few occasions. This could include, for example, research volunteers in my laboratory, honors thesis students that I have supervised, and students who have gone out of their way to attend office hours and/or been regularly engaged in class discussions. I am selective about who I will write references for, not because I am unkind or lazy, but because I know from experience that a strong letter should include concrete examples of my professional interactions with the individual and should speak to their character and their academic abilities. In today''s highly competitive educational system, a letter that merely states that a student did well on the midterm and final exams will not suffice to get into medical or graduate school.However, over the past 2 years many, if not most, students have been attending university remotely with little opportunity to foster meaningful relationships with their instructors, peers, and mentors, especially for those in programs with large enrollments. Indeed, during the peak of Covid‐19, I stopped taking on undergraduate volunteers and greatly reduced the number of honors students in my laboratory. Similarly, my undergraduate lectures have been predominantly delivered online via Zoom, meaning I did not see or speak with most of the students in my courses. It did not help that nearly all of them kept their cameras and microphones turned off and rarely attended online office hours. Consequently, students are desperately struggling to identify individuals who can write them strong letters of reference. In fact, this past spring, I have had more requests for reference letters than ever before, and the same is true for many of my colleagues. Some of the emails I have received have been heartfelt and underscore how taxing the pandemic has been on young adults. With permission, I have included an excerpt from a message I received in early May:Hi Dr. Smith. You may not remember me, but I was in Genome Evolution this year. I enjoyed the class despite being absent for most of your live Zoom lectures because of the poor internet connection where I live. Believe it or not, my mark from your course was the highest of all my classes this term! Last summer, I moved back home to rural Northern Ontario to be closer to my family. My mom is a frontline worker and so I''ve been helping care for my elderly grandmother who has dementia as well as working part‐time as a tutor at the local high school to help pay tuition. All of this means that I''ve not paid as much attention to my studies as I should have. I''m hoping to go to graduate school this coming fall, but I have yet to find a professor who will write a reference for me. Would you please, please consider writing me a letter?I am sympathetic to the challenges students faced and continue to face during Covid‐19 and, therefore, I have gone out of my way to provide as many as I can with letters of support. But, it is no easy feat writing a good reference for someone you only know via an empty Zoom box and a few online assignments. My strategy has been to focus on their scholarly achievements in my courses, providing clear, tangible examples from examinations and essays, and to highlight the notable aspects of their CVs. I also make a point to stress how hard online learning can be for students (and instructors), reiterating some of the themes touched upon above. This may sound unethical to some readers but, in certain circumstances, I have allowed students to draft their own reference letters, which I can then vet, edit, and rewrite as I see fit.But it is not just undergraduates. After months and months of lockdowns and social distancing, many graduate students, postdocs, and professors are also struggling to find suitable references. In April, I submitted my application for promotion to Full Professor, which included the names of 20 potential reviewers. Normally, I would have selected at least some of these names from individuals I met at recent conferences and invited to university seminars, except I have not been to a conference in over 30 months. Moreover, all my recent invited talks have been on Zoom and did not include any one‐on‐one meetings with faculty or students. Thus, I had to include the names of scientists that I met over 3 years ago, hoping that my research made a lasting impression on them. I have heard similar anecdotes from many of my peers both at home and at other universities. Given all of this, I would encourage academics to be more forthcoming than they may have traditionally been when students or colleagues approach them for letters of support. Moreover, I think we could all be a little more forgiving and understanding when assessing our students and peers, be it for admissions into graduate school, promotion, or grant evaluations.Although it seems like life on university campuses is returning to a certain degree of normality, many scholars are still learning and working remotely, and who knows what the future may hold with regard to lockdowns. With this uncertainty, we need to do all we can to engage with and have constructive and enduring relationships with our university communities. For undergraduate and graduate students, this could mean regularly attending online office hours, even if it is only to introduce yourself, as well as actively participating in class discussions, whether they are in‐person, over Zoom, or on digital message boards. Also, do not disregard the potential and possibilities of remote volunteer research positions, especially those related to bioinformatics. Nearly, every laboratory in my department has some aspect of their research that can be carried out from a laptop computer with an Internet connection. Although not necessarily as enticing as working at the bench or in the field, computer‐based projects can be rewarding and an excellent path to a reference letter.If you are actively soliciting references, try and make it as easy as possible on your potential letter writers. Clearly and succinctly outline why you want this person to be a reference, what the letter writing/application process entails, and the deadline. Think months ahead, giving your references ample time to complete the letter, and do not be shy about sending gentle reminders. It is great to attach a CV, but also briefly highlight your most significant achievements in bullet points in your email (e.g., Dean''s Honours List 2021–22). This will save time for your references as they will not have to sift through many pages of a CV. No matter the eventual result of the application or award, be sure to follow up with your letter writers. There is nothing worse than spending time crafting a quality support letter and never learning the ultimate outcome of that effort. And, do not be embarrassed if you are unsuccessful and need to reach out again for another round of references—as Winston Churchill said, “Success is stumbling from failure to failure with no loss of enthusiasm.”     

An unconventional route to becoming a cell biologist

I am honored to be the E. B. Wilson Award recipient for 2015. As we know, it was E. B. Wilson who popularized the concept of a “stem cell” in his book The Cell in Development and Inheritance (1896, London: Macmillan & Co.). Given that stem cell research is my field and that E. B. Wilson is so revered within the cell biology community, I am a bit humbled by how long it took me to truly grasp his vision and imaginative thinking. I appreciate it deeply now, and on this meaningful occasion, I will sketch my rather circuitous road to cell biology.I grew up in a suburb of Chicago. My father was a geochemist, and for everyone whose parents worked at Argonne National Laboratories, Downers Grove was the place to live. My father’s sister was a radiobiologist and my uncle was a nuclear chemist, both at Argonne; they lived in the house next door. Across the street from their house was the Schmidtke’s Popcorn Farm—a great door to knock on at Halloween. The cornfields were also super for playing hide-and-seek, particularly when you happened to be shorter than those Illinois cornstalks.Open in a separate windowElaine FuchsI remember when the first road in the area was paved. It made biking and roller-skating an absolute delight. Fields of butterflies were everywhere, and with development came swamps and ponds filled with pollywogs and local creeks with crayfish. It was natural to become a biologist. When coupled with a family of scientists and a mother active in the Girl Scouts, all the resources were there to make it a perfect path to becoming a scientist.I could hardly wait until I was in junior high school, when I could enter science fairs. You would think that my science-minded family might help me choose and develop a research project. True to their mentoring ethos, they left these decisions to me. My first project was on crayfish behavior. I recorded the response of the crayfish I had caught to “various external stimuli.” At the end of this assault, I dissected the crayfish and, using “comparative anatomy,” attempted to identify all the parts. The second project was no gentler. I focused on tadpole metamorphosis and the effects of thyroid hormone in accelerating development at low concentrations and death at elevated concentrations. Somehow, I ended up going all the way to the state fair, where it became clear that I had serious competition. That experience, however, whetted my appetite to gain more lab experience and to learn to read the literature more carefully.My experience with high school biology prompted me to gravitate toward chemistry, physics, and math. When it came to college, my father told me that if there was a $2000/year (translated in 2015 to be $30,000/year) reason why I should go anywhere besides the University of Chicago (where Argonne scientists received a 50% tuition cut for their children) or the University of Illinois (then $200/year tuition), we could “discuss” it further. Having a sister, father, aunt, and uncle who went to the University of Chicago, I chose the University of Illinois and saved my Dad a bundle of money. At Illinois, I thought I might revisit biology, but my choices for a major were “biology for teachers” or “honors biology.” The first did not interest me; the second seemed intimidating.I enrolled as a chemistry major. Four years went by, during which time I never took a biology class. I enjoyed quantum mechanics, physics, and differential equations, and problem solving became one of my strengths. In the midst of the Vietnam War era, however, Illinois was a hotbed of activity. I was inspired to apply to the Peace Corps, with a backup plan to pursue science that would be more biomedically relevant than quantum mechanics. I was accepted to go to Uganda with the Peace Corps, but with Idi Amin in office, my path to science was clear. Fortunately, the schools I applied to accepted me, even though, in lieu of GRE scores, I had submitted a three-page essay on why I did not think another exam was going to prove anything. I chose Princeton’s biochemistry program. This turned out to be a great, if naïve choice, as only after accepting their offer did I take a biochemistry class to find out what I was getting into. I chose to carry out my PhD with a terrific teacher of intermediary metabolism, Charles Gilvarg, who worked on bacterial cell walls. My thesis project was to tackle how spores break down one cell wall and build another as they transition from quiescence to vegetative growth.By my fourth year of graduate school, I was trained as a chemist and biochemist and was becoming increasingly hooked on biomedical science. I listened to a seminar given by Howard Green, who had developed a method to culture cells from healthy human skin under conditions in which they could be maintained and propagated for hundreds of generations without losing their ability to make tissue. At the time, Howard referred to them as epidermal keratinocytes, but in retrospect, these were the first stem cells ever to be successfully cultured. I was profoundly taken by the system, and Howard’s strength in cell biology inspired me. It was the perfect match for pursuing my postdoctoral research. The time happened to be at the cusp of DNA recombinant technology.At MIT, I learned how to culture these cells. I wanted to determine their program of gene expression and how this changed when epidermal progenitors embark on their terminal differentiation program. While the problem in essence was not so different from that of my graduate work at Princeton, I had miraculously managed to receive my PhD without ever having isolated protein, RNA, or DNA. Working in a quintessential cell biology lab and tackling a molecular biology question necessitated venturing outside the confines of the Green lab and beyond the boundaries of my expertise. Fortunately, this was easy at MIT. Richard Hynes, Bob Horvitz, Bob Weinberg, and Graham Walker were all assistant professors, and their labs were very helpful, as were those of David Baltimore and Phil Sharp, a mere walk across the street. On the floor of my building, Steve Farmer, Avri Ben Ze’ev, Gideon Dreyfuss, and Ihor Lemischka were in Sheldon Penman’s lab just down the hall, and they were equally interested in mRNA biology, providing daily fuel for discussions. Uttam Rhajbandary’s and Gobind Khorana’s labs were also on the same floor, making it easy to learn how to make oligo(dT)-Sepharose to purify my mRNAs. Vernon Ingram’s lab was also on the same floor, so learning to make rabbit reticulocyte lysates to translate my mRNAs was also possible. Howard bought a cryostat, so I could section human skin and separate the layers. And as he was already working with clinicians at Harvard to apply his ability to create sheets of epidermal cells for the treatment of burn patients, I had access to the leftover scraps of human tissue that were also being used in these operations.The three years of my postdoc were accompanied by three Fuchs and Green papers. The first showed that epidermal keratinocytes spend most of their time expressing a group of keratin proteins with distinct sequences. The second showed that these keratins were each encoded by distinct mRNAs. The third showed that, as epidermal keratinocytes commit to terminally differentiate, they switch off expression of basal keratins (K5 and K14) and switch on the expression of suprabasal keratins (K1 and K10). That paper also revealed that different stratified tissues express the same basal keratins but distinct sets of suprabasal keratins. I am still very proud of these accomplishments, and my MIT experience made me thirst to discover more about the epidermis and its stem cells.My first and only real job interview came during my second year of postdoc, at a time when I was not looking for a job. I viewed the opportunity, initiated by my graduate advisor, as a free trip home to visit my parents and my trial run to prepare me for future searching. I was thrilled when this interview materialized into an offer to join the faculty, for which the University of Chicago extended my start time to allow me to complete my three years with Howard.Times have clearly changed, and it is painful to see talented young scientists struggle so much more today. That said, I have never looked ahead very far, and having a lack of expectations or worry is likely to be as helpful today as it was then. I am sure it is easier said than done, but this has also been the same for my science. I have always enjoyed the experiments and the joy of discovery. There was no means to an end other than to contemplate what the data meant in a broader scope.I arrived at the University of Chicago with a well-charted route. My aim was to make a cDNA library and clone and characterize the sequences and genes for the differentially expressed keratins I had identified when I was at MIT. It was three months into my being at Chicago when my chair lined up some interviews for me to hire a technician. I was so immersed in my science that I did not want to take time to hire anyone. I hired the first technician I interviewed. Fortunately, it worked out. However, I turned graduate students away the first year, preferring to carry out the experiments with my technician and get results. After publishing two more papers—one on the existence of two types of keratins that were differentially expressed as pairs and the other on signals that impacted the differential expression of these keratin pairs, I decided to accept a student, who analyzed the human keratin genes. My first postdoc was a fellow grad student with me at Princeton; she studied signaling and keratin gene expression. My second postdoc was initiated by my father, who chatted with him at the elevator when I was moving into my apartment. He set up DNA sequencing and secondary-structure prediction methods, and the lab stayed small, focused, and productive.I was fascinated by keratins, how they assembled into a network of intermediate filaments (Ifs). When thalassemias and sickle cell anemia turned out to be due to defects in globin genes, I began to wonder whether there might be human skin disorders with defective keratin genes. I had no formal training in genetics, and there were no hints of what diseases to focus on. Thus, rather than using positional cloning to identify a gene mutation associated with a particular disease, we took a reverse approach: we first identified the key residues for keratin filament assembly. After discovering that mutations at these sites acted dominant negatively, we engineered transgenic mice harboring our mutant keratin genes and then diagnosed the mouse pathology. Our diagnoses, first for our K14 mutations and then for our K10 mutations, turned out to be correct: on sequencing the keratins from humans with epidermolysis bullosa simplex (EBS), we found K14 or K5 mutations; similarly, we found K1 or K10 mutations in affected, but not in unaffected, members of families with epidermolytic hyperkeratosis (EH). Both are autosomal-dominant disorders in which patients have skin blistering or degeneration upon mechanical stress. Without a proper keratin network, the basal (EBS) or suprabasal (EH) cells could not withstand pressure. Ironically, family sizes of all but the mildest forms of these disorders were small, meaning that the disorders were not amenable to positional cloning. But the beauty of this approach is that once we had made the connection to the diseases, we understood their underlying biology. In addition, the IF genes are a superfamily of more than 100 genes differentially expressed in nearly all tissues of the body. Once we had established EBS as the first IF gene disorder, the pathology and biology set a paradigm for a number of diseases of other tissues that turned out to be due to defects in other IF genes.Fortunately, I had students, Bob Vassar (professor, Northwestern University) and Tony Letai (associate professor, Harvard Medical School), and a postdoc, Pierre Coulombe (chair, Biochemistry and Molecular Biology, Johns Hopkins University), who jumped into this fearless venture with me. We had to go off campus to learn transgenic technology. I had never worked with mice before. When Bob returned to campus with transgenic expertise, we hired and trained Linda Degenstein, whose love for animal science was unparalleled. Pierre’s prior training in electron microscopy was instrumental in multiple ways. Additionally, I was not a dermatologist and had no access to human patients. Fortunately Amy Paller, MD, at Northwestern volunteered to work with us.The success of this project attests to an important recipe: 1) Pursue a question you are passionate about. 2) In carrying out rigorous, well-controlled experiments, each new finding should build upon the previous ones. 3) If you have learned to be comfortable with being uncomfortable, then you will not be afraid to chart new territory when the questions you are excited to answer take an unanticipated turn. 4) Science does not operate in a vacuum. Interact well with your lab mates and take an interest in their science as well as your own. And wherever you embark upon a pathway in which the lab’s expertise is limited, do not hesitate to reach out broadly to other labs and universities.I have followed this recipe now for more than three decades, and it seems to work pretty well. A lab works only when its students and postdocs are interactive, naturally inquisitive, and freely share their ideas and findings. I have been blessed to have a number of such individuals in my lab over the years. When push comes to shove, I am always inclined first to shave from the “brilliant” category and settle for smart, nice people who are passionate and interactive about science and original and unconventional in their thinking.So what questions have I been most passionate about? I have always been fascinated with how tissues form during development, how they are maintained in the adult, and how tissue biology goes awry in human disorders, particularly cancers. I first began to think about this problem during my days at Princeton. I also developed a dogma back then that I still hold: to understand malignancies, one must understand what is normal before one can appreciate what is abnormal. I think this is why I have spent so much of my life focusing on normal tissue morphogenesis, despite my passion for being at the interface with medicine. And because skin has so many amazingly interesting complexities, and because it is a great system to transition seamlessly between a culture dish and an animal, I have never found a reason to choose any other tissue over the one I chose many years back.I will not dwell on the various facets of skin biology we have tackled over the years. Our initial work on keratins was to obtain markers for progenitors and their differentiating lineages. This interest broadened to understanding how proliferative progenitors form cytoskeletal networks and how the cytoskeleton makes dynamic rearrangements during tissue morphogenesis.From the beginning, the lab has also been fascinated by how tissue remodeling occurs in response to environmental signals. Indeed, signals from the microenvironment trigger changes in chromatin dynamics and gene expression within tissue stem cells. Ultimately, this leads to changes in proteins and factors that impact on cell polarity, spindle orientation, asymmetric versus symmetric fate specifications, and ultimately, the balance between proliferation and differentiation.The overarching theme of my lab over these decades is clear, namely, to understand the signals that unspecified progenitors receive that instruct them to generate a stratified epidermis, make hair follicles, or make sweat and sebaceous glands. And if we can understand how this happens, then how are stem cells born, and how do they replace dying cells or regenerate tissue after injury? And, finally, how does this process change during malignant progression or in other aberrant skin conditions?In tackling tissue morphogenesis, I have had to forgo knowing the details of each tree and instead focus on the forest. There are many times when I stand back and can only admire those who are able to dissect beautiful cellular mechanisms with remarkable precision. But I crossed that bridge some years ago in tackling a problem that mandates an appreciation of nearly all the topics covered in Bruce Alberts’ textbook Molecular Biology of the Cell. I am now settled comfortably with the uncomfortable, and the problem of tissue morphogenesis in normal biology and disease continues to keep me more excited about each year’s research than I was the previous year. Perhaps the difference between my days as a student, postdoc, and assistant professor and now is that my joy and excitement is as strong for those I mentor and have mentored as it is for myself.