The pH profile for acid-induced elongation of coleoptile and epicotyl sections is consistent with the acid-growth theory

The acid-growth theory predicts that a solution with a pH identical to that of the apoplast of auxintreated tissues (4.5–5.0) should induce elongation at a rate comparable to that of auxin. Different pH profiles for elongation have been obtained, however, depending on the type of pretreatment between harvest of the sections and the start of the pH-incubations. To determine the acid sensitivity under in vivo conditions, oat (Avena sativa L.) coleoptile, maize (Zea mays L.) coleoptile and pea (Pisum sativum L.) epicotyl sections were abraded so that exogenous buffers could penetrate the free space, and placed in buffered solutions of pH 3.5–6.5 without any preincubation. The extension, without auxin, was measured over the first 3 h. Experiments conducted in three laboratories produced similar results. For all three species, sections placed in buffer without pretreatment elongated at least threefold faster at pH 5.0 than at 6.0 or 6.5, and the rate elongation at pH 5.0 was comparable to that induced by auxin. Pretreatment of abraded sections with pH-6.5 buffer or distilled water adjusted to pH 6.5 or above gave similar results. We conclude that the pH present in the apoplast of auxin-treated coleoptile and stems is sufficiently low to account for the initial growth response to auxin.Abbreviations FS free space - IAA indole-3-acetic acid This research was supported by a grant from the National Adonautics and space Administration (NASA), NAGW 1394 to R.E.C., NASA grant NAGW-297 to M.L.E., and NASA grant NAG 1849 to D.L.R.     

Science in the age of mechanical reproduction: moral and epistemic relations between diagrams and photographs

Sociologists, philosophers and historians of science are gradually recognizing the importance of visual representation. This is part of a more general movement away from a theory-centric view of science and towards an interest in practical aspects of observation and experimentation. Rather than treating science as a matter of demonstrating the logical connection between theoretical and empirical statements, an increasing number of investigations are examining how scientists compose and use diagrams, graphs, photographs, micrographs, maps, charts, and related visual displays. This paper focuses on diagrams in biology, and tries to demonstrate how diagrams are an integral part of the production of scientific knowledge. In order to disclose some of the distinctive practical and analytical uses of diagrams, the paper contrasts the way diagrams and photographs are used in biological texts. Both diagrams and photographs are shown to be “constructions” that separately and together mediate the investigation of scientific phenoman.     

Simulating effects of environmental factors on biological control of Tetranychus urticae by Typhlodromus pyri in apple orchards

Successful biological control of mites is possible under various conditions, and identifying what are the requirements for robust control poses a challenge because interacting factors are involved. Process-based modeling can help to explore these interactions and identify under which conditions biological control is likely, and when not. Here, we present a process-based model for population interactions between the phytophagous mite, Tetranychus urticae, and its predator, Typhlodromus pyri, on apple trees. Temperature and leaf nitrogen concentration influence T. urticae rates of development and reproduction, while temperature and rate of ingestion of prey and pollen influence T. pyri rates of survival and reproduction. Predator and prey population dynamics are linked through a stage structured functional response model that accounts for spatial heterogeneity in population density throughout the trees. T. urticae biomass-days (BMD’s), which account for sizes of larvae, nymphs and adults, indicate level of mite-induced leaf damage. When BMD’s exceed 290 per leaf, there are economic losses. When BMD’s exceed 350 per leaf, T. urticae population growth is curbed and eventually the population decreases. Simulations were run to determine which conditions would lead to current year economic loss and increased risk of loss in the following year, i.e. where more T. urticae than T. pyri are present at the end of September. Risk was high with one or more of the following initial conditions: a high prey: predator ratio (10:1 or more); a low to intermediate (0.04–0.2 T. urticae per leaf) initial density; T. urticae with a higher initial proportion of adult females than T. pyri; and a delayed first detection of mites, whether in late July, or sometimes in late June, but not in early June. Warm summer weather, higher leaf nitrogen and T. urticae immigration into trees were also risk factors. Causes for these patterns based on biological characteristics of T. urticae and T. pyri are discussed, as are counter measures which can be taken to reduce risk.     

Successional dynamics of the cultivated kelp microbiome

Kelp are important primary producers that are colonized by diverse microbes that can have both positive and negative effects on their hosts. The kelp microbiome could support the burgeoning kelp cultivation sector by improving host growth, stress tolerance, and resistance to disease. Fundamental questions about the cultivated kelp microbiome still need to be addressed before microbiome-based approaches can be developed. A critical knowledge gap is how cultivated kelp microbiomes change as hosts grow, particularly following outplanting to sites that vary in abiotic conditions and microbial source pools. In this study we assessed if microbes that colonize kelp in the nursery stage persist after outplanting. We characterized microbiome succession over time on two species of kelp, Alaria marginata and Saccharina latissima, outplanted to open ocean cultivation sites in multiple geographic locations. We tested for host-species specificity of the microbiome and the effect of different abiotic conditions and microbial source pools on kelp microbiome stability during the cultivation process. We found the microbiome of kelp in the nursery is distinct from that of outplanted kelp. Few bacteria persisted on kelp following outplanting. Instead, we identified significant microbiome differences correlated with host species and microbial source pools at each cultivation site. Microbiome variation related to sampling month also indicates that seasonality in host and/or abiotic factors may influence temporal succession and microbiome turnover in cultivated kelps. This study provides a baseline understanding of microbiome dynamics during kelp cultivation and highlights research needs for applying microbiome manipulation to kelp cultivation.     

Diversity,equity, and inclusion in the melanoma research community

The inaugural Diversity and Inclusion in Science Session was held during the 2021 Society for Melanoma Research (SMR) congress. The goal of the session was to discuss diversity, equity, and inclusion in the melanoma research community and strategies to promote the advancement of underrepresented melanoma researchers. An international survey was conducted to assess the diversity, equity, and inclusion (DEI) climate among researchers and clinicians within the Society for Melanoma Research (SMR). The findings suggest there are feelings and experiences of inequity, bias, and harassment within the melanoma community that correlate with one's gender, ethnic/racial group, and/or geographic location. Notably, significant reports of inequity in opportunity, discrimination, and sexual harassment demonstrate there is much work remaining to ensure all scientists in our community experience an academic workplace culture built on mutual respect, fair access, inclusion, and equitable opportunity.     

Enrichment of adeno-associated virus serotype 5 full capsids by anion exchange chromatography with dual salt elution gradients

Adeno-associated virus-based gene therapies have demonstrated substantial therapeutic benefit for the treatment of genetic disorders. In manufacturing processes, viral capsids are produced with and without the encapsidated gene of interest. Capsids devoid of the gene of interest, or “empty” capsids, represent a product-related impurity. As a result, a robust and scalable method to enrich full capsids is crucial to provide patients with as much potentially active product as possible. Anion exchange chromatography has emerged as a highly utilized method for full capsid enrichment across many serotypes due to its ease of use, robustness, and scalability. However, achieving sufficient resolution between the full and empty capsids is not trivial. In this work, anion exchange chromatography was used to achieve empty and full capsid resolution for adeno-associated virus serotype 5. A salt gradient screen of multiple salts with varied valency and Hofmeister series properties was performed to determine optimal peak resolution and aggregate reduction. Dual salt effects were evaluated on the same product and process attributes to identify any synergies with the use of mixed ion gradients. The modified process provided as high as ≥75% AAV5 full capsids (≥3-fold enrichment based on the percent full in the feed stream) with near baseline separation of empty capsids and achieved an overall vector genome step yield of >65%.     

Planar cell polarity proteins differentially regulate extracellular matrix organization and assembly during zebrafish gastrulation

Zebrafish gastrulation cell movements occur in the context of dynamic changes in extracellular matrix (ECM) organization and require the concerted action of planar cell polarity (PCP) proteins that regulate cell elongation and mediolateral alignment. Data obtained using Xenopus laevis gastrulae have shown that integrin–fibronectin interactions underlie the formation of polarized cell protrusions necessary for PCP and have implicated PCP proteins themselves as regulators of ECM. By contrast, the relationship between establishment of PCP and ECM assembly/remodeling during zebrafish gastrulation is unclear. We previously showed that zebrafish embryos carrying a null mutation in the four-pass transmembrane PCP protein vang-like 2 (vangl2) exhibit increased matrix metalloproteinase activity and decreased immunolabeling of fibronectin. These data implicated for the first time a core PCP protein in the regulation of pericellular proteolysis of ECM substrates and raised the question of whether other zebrafish PCP proteins also impact ECM organization. In Drosophila melanogaster, the cytoplasmic PCP protein Prickle binds Van Gogh and regulates its function. Here we report that similar to vangl2, loss of zebrafish prickle1a decreases fibronectin protein levels in gastrula embryos. We further show that Prickle1a physically binds Vangl2 and regulates both the subcellular distribution and total protein level of Vangl2. These data suggest that the ability of Prickle1a to impact fibronectin organization is at least partly due to effects on Vangl2. In contrast to loss of either Vangl2 or Prickle1a function, we find that glypican4 (a Wnt co-receptor) and frizzled7 mutant gastrula embryos with disrupted non-canonical Wnt signaling exhibit the opposite phenotype, namely increased fibronectin assembly. Our data show that glypican4 mutants do not have decreased proteolysis of ECM substrates, but instead have increased cell surface cadherin protein expression and increased intercellular adhesion. These data indicate that Wnt/Glypican4/Frizzled signaling regulates ECM assembly through effects on cadherin-mediated cell cohesion. Together, our results demonstrate that zebrafish Vangl2/Prickle1a and non-canonical Wnt/Frizzled signaling have opposing effects on ECM organization underlying PCP and gastrulation cell movements.     

A structured analysis of risk to important wildlife elements in three Australian Wildlife Conservancy sanctuaries

Where data and information are lacking, structured expert risk assessments can provide a powerful tool to progress natural resource planning. In many situations, practitioners make informal assessments of risk within small groups that typically constitute employees. In this study, we report on three small (in terms of experts) structured expert‐based risk assessment case studies conducted by expert employees of a not‐for‐profit organisation (Australian Wildlife Conservancy) to demonstrate the utility of the approach. The case studies were carried out for three wildlife sanctuaries managed by AWC: Faure Island, Karakamia and Paruna. The likelihood that a set of direct risk factors would cause management failure for sets of important wildlife elements in the three sanctuaries was elicited from the small group of ecological experts. The analysis was couched in terms of a management aim to not lose species from each wildlife element over the management period of 25 years with current management. The experts believed, in particular, that increasing temperature and decreasing water availability associated with climate change were likely to impact significantly upon the vegetation elements and water‐reliant fauna associated with the sanctuaries. Some vegetation elements were also thought likely to be at risk of over‐grazing, unsuitable fire regimes and, in some cases, disease. In addition to predation by exotic predators at one sanctuary, the experts identified additional direct risk factors for various fauna elements associated with expected changes to the vegetation elements, including reduction in food availability, nesting habitat, and generally important life media. From the risk analyses, a preliminary conceptual model was developed to underpin monitoring and to indicate areas for possible management intervention and research. The case studies demonstrate that even in a small workplace team, structured risk assessments can be efficiently accomplished and can provide expedient and transparent information that effectively captures and aggregates the views of the experts.     

Structural insights into the allosteric effects of 4EBP1 on the eukaryotic translation initiation factor eIF4E

The eukaryotic translation initiation factor eIF4E plays key roles in cap-dependent translation and mRNA export. These functions rely on binding the 7-methyl-guanosine moiety (5'cap) on the 5'-end of all mRNAs. eIF4E is regulated by proteins such as eIF4G and eIF4E binding proteins (4EBPs) that bind the dorsal surface of eIF4E, distal to the cap binding site, and modulate cap binding activity. Both proteins increase the affinity of eIF4E for 5'cap. Our understanding of the allosteric effects and structural underpinnings of 4EBP1 or eIF4G binding can be advanced by obtaining structural data on cap-free eIF4E bound to one of these proteins. Here, we report the crystal structure of apo-eIF4E and cap-free eIF4E in complex with a 4EBP1 peptide. We also monitored 4EBP1 binding to cap-free eIF4E in solution using NMR. Together, these studies suggest that 4EBP1 transforms eIF4E into a cap-receptive state. NMR methods were also used to compare the allosteric routes activated by 4EBP1, eIF4G, and the arenavirus Z protein, a negative regulator of cap binding. We observed chemical shift perturbation at the dorsal binding site leading to alterations in the core of the protein, which were ultimately communicated to the unoccupied cap binding site of eIF4E. There were notable similarities between the routes taken by 4EBP1 and eIF4G and differences from the negative regulator Z. Thus, binding of 4EBP1 or eIF4G allosterically drives alterations throughout the protein that increase the affinity of eIF4E for the 5'cap.     

The RhoGAP domain of CYK-4 has an essential role in RhoA activation

Cytokinesis in animal cells is mediated by a cortical actomyosin-based contractile ring. The GTPase RhoA is a critical regulator of this process as it activates both nonmuscle myosin and a nucleator of actin filaments [1]. The site at which active RhoA and its effectors accumulate is controlled by the microtubule-based spindle during anaphase [2]. ECT-2, the guanine nucleotide exchange factor (GEF) that activates RhoA during cytokinesis, is regulated by phosphorylation and subcellular localization [3-5]. ECT2 localization depends on interactions with CYK-4/MgcRacGAP, a Rho GTPase-activating protein (GAP) domain containing protein [5, 6]. Here we show that, contrary to expectations, the Rho GTPase-activating protein (GAP) domain of CYK-4 promotes activation of RhoA during cytokinesis. Furthermore, we show that the primary phenotype caused by mutations in the GAP domain of CYK-4 is not caused by ectopic activation of CED-10/Rac1 and ARX-2/Arp2. However, inhibition of CED-10/Rac1 and ARX-2/Arp2 facilitates ingression of weak cleavage furrows. These results demonstrate that?a GAP domain can contribute to activation of a small GTPase. Furthermore, cleavage furrow ingression is sensitive to the balance of contractile forces and cortical tension.