Twenty years of research on primordial germ cells.

Just twenty years ago I was preparing a research project centred on establishing methods for the isolation and culture of mouse primordial germ cells (PGCs). The project had been suggested to me by Anne McLaren and was to be developed at the Medical Research Council (MRC) "Mammalian Development Unit" in London under the direction of Anne herself. At that time I was a young postdoctoral researcher at the Institute of Histology and Embryology of the University of Rome "La Sapienza" and did not imagine that my decision to be involved in this project would signal a profound switch in my scientific life. From then on my research would mostly concentrate on primordial germ cell biology. I feel like saying that the modern history of mammalian primordial germ cells began twenty years ago at the MRC Mammalian Development Unit under Anne McLaren's impulse. It is not surprising that among the most active researchers in the last twenty years in studying mammalian primordial germ cells, three, namely Chris Wylie, Peter Donovan and myself, began their studies under Anne McLaren's guidance. Over the years, Anne's suggestions and encouragement were always precious for my studies and her presence marked my most important findings on PGC biology. She often invited me to present the results obtained in my laboratory to workshops and congresses. In the present article some of these results particularly influenced by Anne's teaching and suggestions will be briefly reviewed.     

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.     

Reminiscences,collaborations and reflections

This is a personal account by a semi old-timer who completed his official term as a professor of plant biochemistry at Nagoya University in Japan in 1992. My university student life began soon after the World War II (1948). I shared the hardships of many in my age group, in that life was difficult during my college years. I was fortunate to have the opportunity of studying in the USA on a Fulbright scholarship first at Purdue University (1955–1956), and then at the University of California, Berkeley (1956–1957). My graduate study and postdoctoral training in the new world were vitally refreshing and stimulating, which gave me the impetus for becoming a natural scientist associated with academic institutions. Consciously and subconsciously I was impressed by the friendly and liberal atmosphere surrounding young students as well as senior scholars in the United States. But more importantly, I was inspired by the critical and competitive minds prevailing among these people.The appointment as a biochemist at the International Rice Research Institute (IRRI) in the Philippines (1962–1964) was the real start of my professional career. The work was continued upon my return to Nagoya to become a staff member of the Research Institute for Biochemical Regulation (1964–1992). Throughout the years, my major research interest has covered photosynthesis as a whole, involving photosynthetic CO₂-fixation (RuBisCO), carbohydrate metabolism, e.g. starch biosynthesis and breakdown (-amylase), and metabolic regulation, which are interrelated in the basic metabolism of plant cells.I shall briefly describe in this article highlights from my studies and discoveries made and I shall also discuss their possible significance in plant metabolism, with the hope that it does not contradict my sense of humility: They are (a) discovery of ADPG in plants and its role in starch biosynthesis; (b) structure-function relationship of RuBisCO proteins, in particular on heterologous recombination of their subunits of plant-type enzyme molecules derived from the prokaryotic photosynthetic bacteria; (c) molecular evolution of RuBisCO genes; (d) mode of actions (formation, intracellular transport and secretion) of rice seed -amylase and its structural characteristics (distinctive glycosylation), and (e) DNA methylation and regulatory mechanism of photosynthesis gene expression in plastids (amyloplasts). In each step of my research, I shared joy, excitement, disappointment, and agony with my colleagues, an experience that may be common to all researchers. Although it is now becoming well recognized among the scientific community in Japan, I want to point out that interaction of multinational scientific minds in the laboratory produces a vital and creative atmosphere for performance of successful research. I experienced and realized this important fact in my earlier days in the USA and the Philippines. Inasmuch as I believe that this is the most crucial element for any research laboratory to possess, I fondly remember the friendships gained with numerous overseas visitors and collaborators who have contributed immensely to our work.Written at the invitation of Govindjee.     

Fifty-eight years in science--a commentary

After 58 years in science, mostly in pharmacology, one gains perspective. Mine is that there have been important changes over this time, some good and some questionable. In this commentary, I try to reveal how I got to this stage, partially explaining my biases, and possibly helping others learn from my experiences including mistakes. Changing from seeking an M.D. to cellular biology and then to pharmacology early in my career were the best moves I made. The next best move was migration to Canada, away from the McCarthy-McCarran hysteria. Arriving at a time after the end of World War II when science in Canada was expanding was very good luck. I had an excellent opportunity to enjoy both the administration (as Chair of the first independent Department of Pharmacology at the University of Alberta) and the practice of pharmacology (as a practitioner of research on smooth muscle in health and disease). For me, the practice of research has always won over administration when a choice had to be made. Early on, I began to ask questions about educational practices and tried to evaluate them. This led me to initiate changes in laboratories and to seek nondidactic educational approaches such as problem-based learning. I also developed questions about the practice of anonymous peer review. After moving to McMaster in 1975, I was compelled to find a solution for a failed "Pharmacology Program" and eventually developed the first "Smooth Muscle Research Program". Although that was a good solution for the research component, it did not solve the educational needs. This led to the development of "therapeutic problems", which were used to help McMaster medical students educate themselves about applied pharmacology. Now these problems are being used to educate pharmacology honours and graduate students at the University of Alberta. The best part of all these activities is the colleagues and friends that I have interacted with and learned from over the years, and the realization that many of them have collaborated with me again in this volume.     

Plans of mice and men: from bench science to science policy

The transition from bench science to science policy is not always a smooth one, and my journey stretched as far as the unemployment line to the hallowed halls of the U.S. Capitol. While earning my doctorate in microbiology, I found myself more interested in my political activities than my experiments. Thus, my science policy career aspirations were born from merging my love of science with my interest in policy and politics. After receiving my doctorate, I accepted the Henry Luce Scholarship, which allowed me to live in South Korea for 1 year and delve into the field of science policy research. This introduction into science policy occurred at the South Korean think tank called the Science and Technology Policy Institute (STEPI). During that year, I used textbooks, colleagues, and hands-on research projects as my educational introduction into the social science of science and technology decision-making. However, upon returning to the United States during one of the worst job markets in nearly 80 years, securing a position in science policy proved to be very difficult, and I was unemployed for five months. Ultimately, it took more than a year from the end of the Luce Scholarship to obtain my next science policy position with the American Society for Microbiology Congressional Fellowship. This fellowship gave me the opportunity to work as the science and public health advisor to U.S. Senator Harry Reid. While there were significant challenges during my transition from the laboratory to science policy, those challenges made me tougher, more appreciative, and more prepared to move from working at the bench to working in the field of science policy.     

A Reminiscence

Leslie Orgel and Francis Crick with Gobind Khorana in Madison, Wisconsin (December 1965). I first met Leslie at the Endicott House (MIT) in February 1964. Leslie was then spending a period of time at MIT and the occasion was a party for him. During our conversation, Leslie talked about starting some experimental work. He seemed to be particularly interested in polyphosphates and the chemical activation of small molecules (building blocks).Shortly after his move to the Salk Institute in the Fall of 1964 I visited him in January 1965. He already had a lab going. I remember meeting Jim Ferris, in particular, and John Sulston sometime later. That particular time was exciting for my research as well. We had the first results on the Genetic Code using the chemical-biochemical approach that my lab had developed. Francis Crick was also at the Salk Institute during the time of my visit. Both Leslie and Francis were very excited by my results and they began to ask a lot of questions and gave me a whole lot of suggestions about further experiments. In fact, my thinking and planning of things that we were doing were so scrutinized and clarified during these discussions that, it seemed to me, my own group had only to turn out all the experiments that were needed. These interactions with Francis and Leslie continued intensively throughout that year and later. In fact, both Leslie and Francis accepted my invitation to Madison in December 1965 for more discussions.Since those early days of the Salk Institute, I have made numerous visits over the years to Leslie and his research group. It has always been very exciting to learn about the many discoveries bearing on chemical evolution that have unfolded from Leslie's research group. In addition, I have always benefitted from the insightful comments that Leslie invariably provided on my own research. I look forward to our continued interactions and friendship in the future.Leslie, A Happy Birthday!     

Discourse on the development of EEG diagnostics and biofeedback for attention-deficit/hyperactivity disorders

This article presents a review of work that my colleagues and I have been doing during the past 15 years developing a rationale for the diagnosis of attention-deficit/hyperactivity disorder (ADHD) and treatment of ADHD employing EEG biofeedback techniques. The article first briefly reviews the history of research and theory for understanding ADHD and then deals with the development of EEG and event-related potential (ERP) assessment paradigms and treatment protocols for this disorder, including our work and that of others who have replicated our results. Illustrative material from our current research and child case studies is included. Suggestions for future experimental and clinical work in this area are presented and theoretical issues involving the understanding of the neurophysiological and neurological basis of ADHD are discussed.Over the years, many people have been involved both in my laboratory and at Southeastern Biofeedback Institute working with me in developing this area; I wish to acknowledge some of them. They are specifically Dr. Margaret Shouse and Dr. Chris Mann, who have been involved in the initial and recent stages of my research; Ms. Jennifer Samples, who has worked with us in the Institute for many years and has helped us in training many of the children that have benefited from EEG biofeedback. I would especially wish to acknowledge the skill and dedication of Judith O. Lubar, of Southeastern Biofeedback Institute, who has worked with me clinically in terms of developing treatment protocols for ADHD biofeedback and who has trained many of the children who have successfully completed EEG Biofeedback. I would like to acknowledge the generous help of the Lexicor Corporation of Boulder, Colorado who have provided support and instrumentation for recent studies in this area. Mr. Rod Bunn and Mr. Robert Muenchen, who have provided computer support, programming, and statistical assistance in evaluating data in various studies, are gratefully acknowledged. Some of this research was supported by a grant under the ESEA Title IV-C Program for the handicapped. I also gratefully acknowledge Children's Hospital of Knoxville, TN, who have provided essential contract support for our laboratory at the University of Tennessee.     

Discovering the Mechanisms of Neurodegeneration in Prion Diseases

The purpose of my chapter in this issue of Neuroscience Reviews dedicated to Dr. Lawrence Eng is to summarize my contributions to understanding the mechanisms of neurodegeneration in prion diseases. I explain that I was able to advance the field of prion disease neuropathology largely because of the foundation of neurochemistry and immunohistochemistry that I learned while working 5 years in Dr. Engs laboratory. In my review, I relate how my Neuropathology Research Laboratory began as a collaboration with Dr. Stanley Prusiner 20 years ago that led from immunohistochemical staining of amyloid plaques in rodent and human brains using prion protein-specific antibodies to molecular evidence that the abnormal prion protein, PrP^Sc, is the cause of the clinically relevant neuropathological changes in animal and human prion diseases.Special issue dedicated to Dr. Lawrence F. Eng.     

A more cost-effective method of preoperative computerized imaging

Stimulated by the explosive expansion of the computerized desk top publishing industry during the past few years, microcomputer hardware and software are evolving at a staggering rate. Memory is rapidly increasing, and prices are declining. I have found that with the hardware and software described in this paper, I was able to obtain, in a much more cost-effective manner, as useful preoperative information for my practice as I could obtain with more expensive "turnkey" (only one use) computerized imaging systems. This type of microcomputer, of course, is not limited to just the imaging system, but can be used for a variety of other programs as well, such as word processing, slide labeling and production, spreadsheet functions, billing and filing, and numerous business and other applications. The ease of use with readily available 35-mm slides of my patients has greatly enhanced the appeal of this system. Computerized imaging, when used as an educational tool, can be very helpful in preoperative planning, resident teaching, and for illustration and discussion of a patient's proposed surgery. The electronic imaging disclaimer compiled by the American Society of Plastic and Reconstructive Surgeons has been extremely helpful in clarifying the limits of computerized imaging and reducing any false expectations that my patients might have. All of us are experiencing the dawn of a very exciting evolution.     

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.”     

You are the most valuable reagent in the lab: mental health before and during the pandemic

No one maps out their tenure as a postdoc anticipating a life-altering tragedy. But mental health crises of all kinds affect academic trainees and staff at similar or higher levels than the general public. While the mental health resources available to trainees are often set by healthcare providers, all levels of university leadership can work to remove material and immaterial obstacles that render such resources out of reach. I describe how access to care via telemedicine helped me following a loss in my family.

Over the years, my siblings and close friends have sought mental health resources like therapy, psychoanalysis, or psychiatry, so I loosely understood their benefits. When I was a PhD student I went to therapy briefly, but my counselor and I decided I could do without it. Since I started my postdoc, stress manifested in some new ways but I managed it well with my usual coping strategies and support. That changed one bright December morning in 2019 while I was preparing for our weekly lab meeting. My phone rang indicating a call from my father, whom I had spoken to the night before to celebrate the news of my nephew’s birth. But the voice on the phone was that of a family friend, telling me that my father had died overnight of an undiagnosed heart condition. In the moment I couldn’t even understand what was happening, saying over and over, “but I talked to him last night.” Soon I was sitting at home, dazed, on a string of tearful calls with family and friends.I often read words like “lifted” or “buoyed” to describe the stabilizing support of a network of loved ones. In my case this network was tethering me to reality over the next few weeks, preventing me from spinning off the Earth’s surface in a storm of sorrow and anxiety. The trauma also took a strange physical form and convinced me that I was suffering from a cardiac condition of my own. I had a panic attack during which I went to urgent care convinced my own heart was about to give way. Night after night these physical symptoms prevented me from sleeping.Graced by many loving connections with my siblings, my boyfriend, and close friends, I was actually weathering the process as well as one can. My PI gave me a firm directive to take as much time off as I needed. These were two key elements early in my healing process: a supportive network and an understanding advisor. The third was getting professional help, which I soon realized I needed. Even if I felt OK one day, I didn’t trust that I’d be OK the next. My grief formed too thick and too broad a landscape for me to navigate without help.Deciding to seek mental health resources and realizing that one needs them are often the hardest parts. Connecting with those resources once the decision has been made should be as simple as possible. I called a mental health number, and a triage counselor noted my therapy needs and verified my insurance. She asked what times and locations I preferred and then searched for an open appointment with a therapist who accepted my insurance. She also informed me that my coverage allowed 12 sessions with no copay, which was a pleasant surprise. The therapist who agreed to see me had very few openings, in part because this all happened in December—the holidays are especially busy for therapists. I was aiming for a time after normal working hours, or in the morning before I would head to lab, but none of those times were available. I didn’t like interrupting my workday to trot off to therapy. Taking a long break once a week meant I couldn’t run experiments or mentor my student during that time. But I made the sacrifice because my highest priority was getting the help I needed. There was no shortcut. Prioritizing mental health over lab work is tough for researchers, and I would never have accepted that kind of weekly disruption before my dad’s passing. But as a wonderful mentor of mine used to say, “You are the most valuable reagent in the lab.” She wasn’t describing mental health at the time, but the phrase now provided a guiding principle for my recovery. My first few sessions were on Tuesdays at 2:00 pm.The afternoon break turned out to be less disruptive than I had feared, because I had recently come back to the lab and was working short days. Had she asked, I would have told my PI where I was on Tuesday afternoons, but she wasn’t normally abreast of my daily schedule, so I didn’t seek her approval beforehand. Coordinating experiments with lab members thankfully wasn’t an issue because my work was largely independent; I simply let lab members know that I’d would be out of the lab for a bit on those days.The weeks went by, and the benefits of therapy accrued, helping me in large and small ways as I grieved. In mid-March of 2020, my therapist followed public health guidelines and asked all her clients to transition to remote sessions. While this was easy and sensible, it was still a little disappointing. Therapists are professional empaths, among many other things, and doing away with the physical presence and exchange with her was a blow. Yet therapy via video felt less odd simply because most of my social interactions were now virtual. Thankfully I didn’t have to move out of state for the lockdown (as did many students living in campus housing), which meant I could stay with the same therapist without any insurance complications.A few weeks into lockdown, I asked my therapist whether we had reached the limit of my 12 sessions without a copay. She replied with the good news that my insurance provider had waived all copays for mental health costs due to the pandemic. By that time therapy had generated a platform and an outlet to explore areas of my grief beyond the trauma of my father’s passing. Without needing to weigh the costs and benefits of this resource, I saw my therapist for another 4 months. I slowly took stock of my upbringing in an unconventional family and the loss of my mother when I was 25 and waded through a series of difficult decisions regarding my father’s estate. My father’s death changed me at a depth that is untouched by any amount of therapy or treatment. I’m not “healed”: I feel aged, more brittle, and a little ground down compared with who I had been. But therapy guided me through the worst of my grief, past the acute trauma to help me grasp what I was going through.Since the pandemic began, the number of people reporting increased stress or mental health issues has steadily increased (information on the impact of COVID-19 measures on mental health: https://www.apa.org/workforce/publications/depression-anxiety-coronavirus.pdf) (also see Mental health resources for trainees). I am fortunate to have affordable health insurance and the support from my lab and my department. The ease of finding my institution’s phone number for mental health resources was itself an important benefit. I share these pieces of my story with humility and understanding that not everyone enjoys the privileges that I do and the knowledge that everyone weathers life’s tragedies in their own way. It is not lost on me that some benefits stemmed from a policy change made by a private insurance provider. The provider made the right decision to waive copays, freeing me from having to choose between cost and my mental health needs. Yet had I been a student who had to move out of state due to COVID-19, access to mental health resources might have been disrupted or cut off. The need for reduced out-of-pocket costs for healthcare is known and needs no repetition, but the benefits of telehealth should be a low-cost component of health plans offered to students and staff (information on telehealth recommendations: https://www.apaservices.org/advocacy/news/congress-patient-telehealth?_ga=2.231013471.1538013741.1619359426-1228006513.1619359425 and http://www.apaservices.org/practice/advocacy/state/leadership/telebehavioral-health-policies.pdf?_ga=2.3385904.1067518037.1620039082-1228006513.1619359425.I’m not a cloud of emotions attached to a pair of good pipetting hands, I’m a human who is choosing to spend my time doing research. This observation is easy to repeat, by trainees as much as by faculty and administrators, but much harder to act upon in the midst of conflicting priorities. Consider my story a success: Because I could access the resources I needed, I was able to prioritize my mental health in the midst of my ambitious research program even during the lockdown.MEET THE AUTHORI have been a postdoc in Stefani Spranger’s lab at MIT for 4 years. Supported by an Irvington Fellowship from the Cancer Research Institute, my work examines the behaviors of dendritic cells in tumors that contribute to productive or unproductive anti-tumor immune responses. My doctoral work examined modes of multicellular invasion controlled by the actin cytoskeleton with Margaret Gardel at the University of Chicago. Earlier I was a lab technician with Thea Tlsty at the University of California, San Francisco, which followed a bachelor’s degree in biology at the University of California, Santa Cruz. I serve on the Committee for Students and Postdocs at the American Society for Cell Biology, where I chair the Outreach Subcommittee.     

Lessons learned from Art Pardee in cell cycle, science, and life

This essay is written to honor Dr Art Pardee's 85th birthday (July 13, 2006). In this essay, I have summarized the lessons I learned from Art and the cell-cycle research I performed in Art's laboratory during my postdoctoral training period. I have also summarized some research from my own laboratory that has been inspired by the lessons I learned from Art, including the interactions between cell cycle and cell death regulators and discovery of novel polyphenol- and copper-based proteasome inhibitors. Finally, I have discussed the potential use of these proteasome inhibitors in cancer prevention and treatment.     

The "sense of being stared at" does not depend on known sensory clues.

The "sense of being stared at" can be investigated by means of simple experiments in which subjects and lookers work in pairs, with the looker sitting behind the subject. In a random sequence of trials, the looker either looks at the back of the subject, or looks away and thinks of something else. More than 15,000 trials have already been conducted, involving more than 700 subjects, with an extremely significant excess of correct over incorrect guesses (Sheldrake [1999]). This effect was still apparent in experiments in which subjects were blindfolded and given no feedback, showing it did not depend on visual clues, nor on the subjects knowing if their guesses were right or wrong (Sheldrake [2000]). In this paper I describe experiments I conducted in schools in England in which the subjects were not only blindfolded and given no feedback, but looked at through closed windows. There was again a very significant excess of correct over incorrect guesses (p < 0.004). At my request, schoolteachers in Canada, Germany and the United States carried out similar experiments and found an even more significant positive effect than in my own experiments (p < 0.0002). The fact that positive results were still obtained when visual clues had been effectively eliminated by blindfolds, and auditory and olfactory clues by closed windows, implies that the sense of being stared at does not depend on the known senses. I conclude that peoples' ability to know when they are being looked at depends on an influence at present unknown to science.     

Adventures in Neural Plasticity, Aging, and Neurodegenerative Disorders Aboard the CWC Beagle

This article recounts some of the scientific endeavors of Carl W. Cotman (CWC) during his journeys through the cellular circuitry of the mammalian brain. I have selected for consideration his findings that have been an important impetus for my own research; in several cases our different experiments have provided complementary data to support an hypothesis. Three examples are (i) Carl's studies of the roles of glutamate in synaptic transmission and plasticity in the adult brain and my studies of how glutamate regulates neurite outgrowth and cell survival in brain development; (ii) his and our studies of the mechanisms whereby amyloid -peptide damages and kills neurons; and (iii) Carl's evidence that physical activity regulates neurotrophin levels in the brain and our evidence that dietary restriction has similar effects and is neuroprotective. In case you have not yet realized how I chose a title for this article it is because Carl has a (very distant) connection with Charles Darwin—Darwin sailed on a vessel called the Beagle and Carl has studied beagle dogs, establishing them as a model for understanding the neurobiology of human brain aging.     

Decoding the genetics of rare disease: an interview with Monkol Lek

Monkol Lek, Assistant Professor at Yale University School of Medicine, and Associate Editor at Disease Models & Mechanisms, dedicates his research to finding a genetic diagnosis and improving treatments for rare disease patients. As he originally studied computer engineering at the University of New South Wales in Sydney, Australia, he now utilises computational methods to optimise large-scale genetic studies, provide globally accessible resources for genetic research communities and, importantly, resolve diagnostic odysseys for rare disease patients. Monkol completed his PhD in Prof. Kathryn North''s lab at the University of Sydney, studying the genetics of muscle strength and performance, and then continued his investigation of muscle disease in Prof. Daniel MacArthur''s lab at Massachusetts General Hospital and the Broad Institute. During his postdoc, he led several large-scale studies aimed at distinguishing pathogenic from benign variants, including the Exome Aggregation Consortium (ExAC) project ( Lek et al., 2016). Monkol established his own lab at Yale University School of Medicine, which continues to improve the diagnosis and treatment of rare muscle disease, and also focuses on underserved populations, whose genetic mutations are not as well characterised as those of European ancestry. In this interview, Monkol discusses how his own diagnosis with limb girdle muscular dystrophy has shaped his career and what he envisions for the future of genetic research in rare disease.

You have a very unique career path – could you tell us a little bit about that? My first degree was in computer engineering. When I first went to university, I studied the hardware and software of computers. I really liked the software aspect of the degree, and so I worked for IBM as a software developer when I finished university. However, during the last few years of university, I noticed that my muscles were getting weaker. My university was on a big hill, with classes at the bottom and top of the hill, and I had to stand up for about 3 h a day while commuting on public transport. It started becoming obvious that I had something wrong with my muscles because I felt totally exhausted at the end of the day. It was frustrating, because I felt that my performance at university was impacted by something that had nothing to do with my ability to think. So, I went from doctor to doctor to try to find out what was wrong with me. As a lot of doctors are not trained in rare diseases, they didn''t consider a rare disease diagnosis. Then one doctor did a blood test for creatine kinase (CK), which is leaked into the bloodstream when muscle is damaged. In healthy people, high levels of CK are detected in the bloodstream after they''ve done intensive exercise, like a marathon. If someone hasn''t done something like that, but they have high levels of circulating CK, it could be an indication that there''s something wrong with their muscles. As I had high levels of CK in my bloodstream, I then went to a neurologist, which was when I got a clinical diagnosis. At that point, they didn’t know the root cause of the problem, but they knew that I have a muscle disease based on several tests, including a nerve conduction test.I received this clinical diagnosis during my time in IBM, and that''s when I became dissatisfied with my job, because I felt that I was using all my talents to make a very big, international company richer. I was also becoming frustrated when visiting the neurologist every 6 months, as all they would tell me was that my muscles were getting weaker, which I already knew. I began to think that not much was happening in the neuromuscular disease field if that''s the best they could offer me. I wanted to know what the root cause of my disease was and if there were any treatment options. I came to the conclusion that no one would care about my disease more than I would, because I''m the one that has lived with it every day of my life.That''s when I decided to leave IBM and pursue a career in researching muscle disease. It didn''t go down well with my parents and friends, because I was leaving a well-paid job to go back to university to get paid nothing for an unknown number of years. If I had known my chances of success – completing a meaningful PhD, doing a meaningful postdoc and landing a faculty position – I wouldn''t have gone on this journey. I have been very fortunate, but I wasn''t always in the right place at the right time.When I finished my undergraduate degree in bioinformatics and physiology at the University of New South Wales, I started a PhD in Melbourne, but it didn''t work out, because not all supervisors are perfect. My wife and I then returned to Sydney, where my wife bumped into one of the professors from our undergraduate degree. She explained that we''d had a bad experience in Melbourne with our PhDs, but our passion was still to do muscle research. The professor''s daughter was researching muscle disease in Kathryn North''s lab at the University of Sydney, and she invited us to visit the lab. I was offered an opportunity to do my PhD in Kathryn''s lab, but I was initially reluctant as it was a diagnostic lab, and I was more interested in developing therapies for people with muscle disease. However, I thought I could still learn a lot about muscle physiology and, in the long term, I''m glad that I received training and mentorship from Kathy''s lab. Also, if I hadn''t done my PhD there, I wouldn''t have met Daniel MacArthur, my future boss. He was a very talented student in Kathy''s lab, who taught me a lot about scientific communication among other things, and I taught him some coding skills. He left to work on the 1000 Genomes Project in Cambridge, UK, but I kept in contact with him to get his advice on my project.When I was finishing my PhD, Daniel asked if I wanted to join the lab he was setting up in Massachusetts General Hospital and the Broad Institute. His lab was going to study common loss-of-function mutations in human populations using large datasets from the 1000 Genomes Project, but he offered me a project investigating neuromuscular diseases. As soon as I submitted my PhD thesis, I started working in his lab. This was perfect timing, because it was 2012, when exome sequencing had recently been published in the context of rare diseases (Ng et al., 2010) and, more importantly, it was becoming affordable, in terms of research. I waited over 10 years for a genetic diagnosis, so my goal was that no one should have to wait that long in the future.Through collaboration with our former PhD lab, Daniel and I used samples from undiagnosed patients to find answers for Australian families. The first family had two affected girls with undiagnosed nemaline myopathy, who had been on a diagnostic odyssey for about 9 years. It was amazing how quickly we progressed from receiving the samples to identifying the novel gene, LMOD3, associated with their disease (Yuen et al., 2014). This was part of my main project during my postdoc – working on gene discovery in neuromuscular diseases and finding answers for patients that have been waiting years and years to get a genetic diagnosis (Ghaoui et al., 2015; O''Grady et al., 2016).The project that most people know me for is the ExAC project, which was initially my ‘side’ project during my postdoc. The idea was to create a big database of all rare variants that we see in the general population, so we can better interpret the rare variants that we see in rare disease patients. When we were creating it, we thought that it may be useful to other researchers around the world. Therefore, we tried to ensure, through data-use agreements and consent processes, that we could share as many of our findings as possible. I''m happy to say my side project was quite successful. After that, I led other projects, including an analysis group in the Centre for Mendelian Genomics, to expand that framework and idea across all rare diseases, not just neuromuscular diseases (Baxter et al., 2022).I was having a lot of fun at the Broad Institute, and I was co-author on a lot of high-impact papers. However, the reason I left the Broad Institute was that I wanted to be involved in the full journey for the patients. Sometimes scientists don''t understand that getting a genetic diagnosis is not the end of the journey for a patient. After the diagnosis they want to know what treatment options are available. Yale gave me the opportunity to continue doing the gene discovery and analytical work that I was doing at the Broad Institute, plus the capability of doing experiments with mouse models to investigate gene replacement therapies and other therapeutic approaches.

“I waited over 10 years for a genetic diagnosis, so my goal was that no one should have to wait that long in the future.”

How has being both a researcher and a patient affected your career? When I was first diagnosed, there was a neurologist who discouraged me from researching my own disease and this became the basis of my TEDx talk, because I thought it was very condescending. I thought, “Just because I have this disease, it doesn''t mean that I have a low IQ”. However, this experience motivated me more. I discussed it with Kathy before starting my PhD, and her encouragement and enthusiasm was refreshing. At the time, in the early 2000s, people hadn''t accepted the idea of patients researching their own disease. Things have changed since then, mainly because there are more examples of it now (Branca, 2019), but at the time, it was really hard for me to progress in science. I always thought that people were looking at me with sympathy, and I felt like I had to achieve twice as much to get the same respect as someone else who wasn''t as talented or didn''t work as hard as me. It was frustrating, but in everyday life people still correlate physical disability with intellectual disability. For example, if my wife is pushing me in the wheelchair in public, no one ever directs a question to me because they assume that the physical disability comes with mental disabilities. There are well-known examples of scientists with physical disabilities, like Stephen Hawking, but it is still challenging in academia when you have a physical disability and people make certain assumptions about you.On the other hand, just before starting at Yale, my collaborators at the University of Massachusetts took a skin biopsy from me. With this skin biopsy, they created induced pluripotent stem cells, and, using CRISPR, they corrected my disease-associated gene variant in the cultured cells. They then published this in a Nature article, in which fig. 1 is the experiment in which they corrected my mutation (Iyer et al., 2019). Are there specific skills or knowledge you learned while working in computer engineering that have helped shape and develop your research today? When I started my PhD, there was an increase in how much genetics research, and biological research in general, relied upon big data. It can be very challenging to work with big data if you''re a biologist without a background in computer science. You can go online to teach yourself to an extent, but it gives you an advantage to learn the theory behind a lot of algorithms and other aspects of software engineering, in a formal setting. It makes the difference between building tools that take a week to analyse a set of data and building tools that take a few minutes to analyse the same data. If you can analyse the data more quickly, you can explore different possibilities and ideas much more quickly. You can''t learn everything online, and having a firm foundation of knowledge can enable you to work with big data in an efficient way.The other thing that you learn from computer science is a certain mindset when approaching problem solving. This is because you have to debug code frequently and, due to this fast pace, you learn quickly. This helped me to troubleshoot problems in biological research quickly.

“Getting a genetic diagnosis is not the end of the journey for a patient. After the diagnosis they want to know what treatment options are available.”

What do you think are the key challenges for rare disease research and diagnosis moving forward? I now have a greater appreciation of the challenges because I see it from two points of view: one as a researcher in a group and one as a PI, who leads the research. The diagnosis rate for rare disease is about 50%, so there are still 50% of patients with a disease that has an unknown genetic cause. The gold standard requirement for associating a new disease gene with a novel phenotype is that it presents in multiple unrelated families (MacArthur et al., 2014). However, when you work with rare diseases, there is the issue of small sample numbers. One challenge for basic scientists is creating good collaborations with physician scientists across the world to enable you to create a large enough dataset.The other challenge is the cost of research for these diseases with unknown genetic cause. The 50% of cases for which we know the genetic cause are no longer considered an area of research, as clinical genetic services can now diagnose these patients. To diagnose the remaining patients, you have to use more expensive technologies, such as long-read sequencing.The last thing is the interpretation of rare variants. Although the ExAC project helped with this, there is still a challenge. For example, if a patient has a rare genetic variant, this doesn''t necessarily mean it is the cause of their rare disease. This is because even healthy people have rare variants. So, we have a massive interpretation challenge in rare disease genetics, which can be overcome by creating a laboratory model system with that genetic variant to investigate it further. However, if you had 1000 variants to consider, it''s not going to scale as an animal model. So, an important question is how can we interpret these variants in a scalable manner? This is one of the main driving forces behind the new Subject Focus, ‘Genetic variance in human disease: decoding diversity to advance modern medicine’, that we are launching in DMM. You have led and coordinated several studies involving very large cohorts. From your experience what are the key components of a successful study? I think the key to a successful large cohort study with unsolved rare disease patients, is the amount of structured phenotype data you can collect. This requires a good collaborator, who has the time to prepare that data in a meaningful way, which makes it easier to find other families with the same rare disease. The other thing is to have the ability to recontact patients and collect different samples from them, because we''re moving to a more multi-omics world. Therefore, we need the ability to go beyond just collecting DNA samples. Also, we''re in a world where we''re starting to link data to electronic health records, which allows the collection of deeper and richer phenotype data that enable associations to be made between families.In addition, you can''t work in isolation. In order for us to make a meaningful impact, we need to work with groups that have specialties outside of our own. For instance, we collaborate with groups that specialise in the interpretation of non-coding variants. This is important as variants in these regions could hold the answers for some of those unsolved cases.Another key aspect to a successful study is collaboration with statistical geneticists because some of the more complicated questions are best asked by them. Some of these questions go beyond monogenic diseases. We are seeing convergence between genome-wide association studies, looking for many variants, each with very small contributions to a disease, and studies of Mendelian disease that are looking for one gene that causes disease. The field has to start looking at diseases in the middle of this spectrum, which requires statistical geneticists. This is because you need to make sure that your conclusions are correct. For instance, if you''re asking whether a rare disease is caused by a combination of two genes, then you must have a robust statistical model to show that these variants aren''t presenting together by chance. You have to prove that those two variants are acting in concert, instead of independently, to cause this disease. My colleagues at Yale published a great paper that demonstrated this concept (Timberlake et al., 2016).Lastly, it is important to forge meaningful collaborations beyond academia. A lot of my colleagues are being funded by industry collaboration, and a lot of these companies have access to more samples than we do in academia. You can also collaborate with large biobanks, such as the UK Biobank, which has a rich set of phenotype data and also the ability to recontact patients (Glynn and Greenland, 2020). The FinnGen project is a recent public–private collaboration that combines genetic data with electronic health records from Finnish biobank participants to improve disease diagnosis and treatment (Kurki et al., 2022 preprint). So, working with biobanks and industry is another way of increasing sample numbers, which is the biggest challenge in rare disease research.

“We don''t want to create disparity in terms of health, especially in the context of genetics, which will continue to become more prominent in modern medicine.”

You dedicate a lot of your research towards patients in underserved populations, such as East Asian populations, whose genetic mutations are not as well characterised as those of European ancestry. Can you explain the importance of this? One of the reasons that it took over 10 years for me to get a genetic diagnosis was because the gene that causes my disease was first reported as not commonly associated with disease in populations of European ancestry. The problem with biomedical research is that when people read that, they think it applies to everyone, even patients who have non-European ancestry. Although the gene that causes my disease aligned with my muscle disease phenotype, it wasn''t sequenced because of this assumption. They only decided to sequence this gene once they did linkage analysis of my family, and this was the only gene associated with neuromuscular disease in the linkage region they identified. This is the reason why we need to have good data on all populations. The ExAC and gnomAD studies that I worked on acknowledged that we need good allele frequency data for populations of East Asian, South Asian, Latino and African ancestry, because we don''t want to create disparity in terms of health, especially in the context of genetics, which will continue to become more prominent in modern medicine.If you want to deliver the best healthcare, you have to realise that some variants and diseases are more common in certain populations, such as Tay-Sachs disease, which is common amongst the Jewish community, and sickle cell anaemia, which is more prevalent in populations of African ancestry. By understanding these differences, we can actually find a genetic diagnosis a lot quicker. If it''s not a de novo variant, and is instead a variant inherited in the population, and if you''ve made the discovery in East Asians, there is a better chance of identifying more incidences of this variant in the population in which it was first discovered.I think it''s also good for validation of data, because if you had discovered a potential disease-causing variant and you find that this variant has a frequency of 1% or higher in a non-European population, then it''s impossible for it to be the cause of a rare disease, regardless of its frequency in a European population (Lek et al., 2016).