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Search indexed bioRxiv preprints in genomics, neuroscience, cell biology and bioinformatics. Read source abstracts and check manuscript versions; preprints are not peer reviewed.

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bioRxiv2016

Behavioural effects of temperature on ectothermic animals: unifying thermal physiology and behavioural plasticity

Temperature imposes significant constraints on ectothermic animals, and these organisms have evolved numerous adaptations to respond to these constraints. While the impacts of temperature on the physiology of ectotherms have been extensively studied, there are currently no frameworks available that outline the multiple and often simultaneous pathways by which temperature can affect behaviour. Drawing from the literature on insects, we propose a unified framework that should apply to all ectothermic animals, generalizing temperature's behavioural effects into (1) Kinetic effects, resulting from temperature's bottom-up constraining influence on metabolism and neurophysiology over a range of timescales (from short-to long-term), and (2) Integrated effects, where the top-down integration of thermal information intentionally initiates or modifies a behaviour (behavioural thermoregulation, thermal orientation, thermosensory behavioural adjustments). We discuss the difficulty in distinguishing adaptive behavioural changes due to temperature from behavioural changes that are the products of constraints, and propose two complementary approaches to help make this distinction and class behaviours according to our framework: (i) behavioural kinetic null modeling and (ii) behavioural ecology experiments using temperature-insensitive mutants. Our framework should help to guide future research on the complex relationship between temperature and behaviour in ectothermic animals.

Ecology↗

bioRxiv2016

Gene expression analysis provides insight into the physiology of the important staple food crop cassava

O_LICassava (Manihot esculenta) feeds approximately 800 million people worldwide. Although this crop displays high productivity under drought and poor soil conditions, it is susceptible to disease, postharvest deterioration and the roots contain low nutritional content.\nC_LIO_LIHere, we provide molecular identities for eleven cassava tissue types through RNA-sequencing and develop an open access, web-based interface for further interrogation of the data.\nC_LIO_LIThrough this dataset, we report novel insight into the physiology of cassava and identify promoters able to drive specified tissue expression profiles. Specifically, we focus on identification of the transcriptional signatures that define the massive, underground storage roots used as a food source and the favored target tissue for transgene integration and genome editing, friable embryogenic callus (FEC).\nC_LIO_LIThe information gained from this study is of value for both conventional and biotechnological improvement programs.\nC_LI

Plant Biology↗

bioRxiv2016

Stomatal and physiological response of contrasting Z. mauritiana (Lamk.) clones to water stress.

Water stress is one of the major limitations to fruit production worldwide. Identifying suitable indicators, screening techniques and quantifiable traits would facilitate the genetic improvement process for water stress tolerance. In the present study, we evaluated the ability of physiological parameters (Transpiration, E; Fv/Fm; leaf water potential, {psi}leaf; leaf temperature, LT; and, leaf relative water content, RWC) to distinguish between contrasting Z. mauritiana clones subjected to a 30-d drought cycle. Four field-grown clones Seb and Gola (tetraploid) and Q 29 and B 5/4 (diploid) were studied. By 30 d after the onset of water stress treatment, the E, Fv/Fm, {psi}leaf and RWC of drought-stressed plants had declined significantly in all genotypes compared to values of well-watered treatments. However, the reductions were more severe in leaves of diploid clones. Under drought stress, the Seb and Gola, maintained higher E (31.5%), Fv/Fm (6.28%), {psi}leaf; (11.2%), and RWC (9.3 %) than Q 29 and B 5/4 clones. In general, LT of drought-stressed plants was higher (~4{degrees}C) than that of well-watered plants but the relative increase was greater among later than former ones. Under maximum drought stress, LT of Seb and Gola clones was on average 3.0{degrees}C lower than that of Q 29 and B 5/4. Former clones yielded 20% more than later ones, mainly reason being (14.8%) less fruit drop as an effect of water stress. The results indicate that presented parameters can be reliable in screening for water stress tolerance ability, with Fv/Fm, {psi}leaf, RWC and LT having the added advantage of being easily and quickly assessed.

Plant Biology↗

bioRxiv2016

An Electrochemistry Study of Cryoelectrolysis in Frozen Physiological Saline

This is the first quantitative study on the fundamental physical and electrochemical processes that occur during cryoelectrolysis. Cryoelectrolysis is a new minimally invasive tissue ablation surgical technique that combines the processes of electrolysis and solid/liquid phase transformation (freezing). We measured the pH front propagation and the changes in resistance in a tissue simulant made of physiological saline gel with a pH dye as a function of the sample temperature in the high subzero range above the eutectic. Results demonstrate that effective electrolysis can occur in a high subzero freezing milieu and that the propagation of the pH front is only weakly dependent on temperature. These observations are consistent with a mechanism involving ionic movement through the concentrated saline solution channels between ice crystals at subfreezing temperatures above the eutectic. The Joule heating in these microchannel may cause local microscopic melting, the observed weak dependence of pH front propagation on temperature, and the large changes in resistance with time. In addition, we observed that the pH front propagation from the anode is more rapid than from the cathode. The explanation is the electro-osmotic flow from the cathode to the anode. The findings in this paper may be of fundamental value for designing future cryoelectrolytic ablation surgery protocols.

Bioengineering↗

bioRxiv2016

A Saposin deficiency model in Drosophila: lysosomal storage, progressive neurodegeneration, sensory physiological decline and defective calcium homeostasis

Saposin deficiency is a childhood neurodegenerative lysosomal storage disorder (LSD) that can cause premature death within three months of life. Saposins are activator proteins that promote the function of lysosomal hydrolases in the degradation of sphingolipids. There are four saposin proteins in humans, which are encoded by the prosaposin gene. Mutations causing an absence of individual saposins or the whole prosaposin gene lead to distinct LSDs due to the storage of different classes of sphingolipids. The pathological events leading to neuronal dysfunction induced by lysosomal storage of sphingolipids are as yet poorly defined. We have generated and characterised a Drosophila model of saposin deficiency that shows striking similarities to the human diseases. Drosophila saposin-related (dSap-r) mutants show a reduced longevity, progressive neurodegeneration, lysosomal storage, dramatic swelling of neuronal soma, perturbations in sphingolipid catabolism, and sensory physiological deterioration. We have also revealed a genetic interaction with a calcium exchanger (CalX), suggesting that calcium homeostasis may be altered in saposin deficiency. Together these findings support the use of dSap-r mutants in advancing our understanding of the cellular pathology implicated in saposin deficiency and related LSDs.

Neuroscience↗

bioRxiv2017

In vitro assembly of the Rous Sarcoma Virus capsid protein into hexamer tubes under physiological conditions.

During a proteolytically-driven maturation process, the ortho-retroviral capsid protein (CA) assembles to form the convex shell that surrounds the viral genome. In some orthoretroviruses, including Rous Sarcoma Virus (RSV), CA carries a short and hydrophobic spacer peptide (SP) at its C-terminus early in the maturation process, which is progressively removed as maturation proceeds. In this work, we show that RSV CA assembles in vitro at physiological temperatures, forming hexamer tubes that effectively model the mature capsid surface. Tube assembly is strongly influenced by electrostatic effects, and is a nucleated process that remains thermodynamically favored at lower temperatures, but is effectively arrested by the large Gibbs energy barrier associated with nucleation. RSV CA tubes are multi-layered, being formed by nested and concentric tubes of capsid hexamers. However the spacer peptide acts as a layering determinant during tube assembly. If only a minor fraction of CA-SP is present, multi-layered tube formation is blocked, and single-layered tubes predominate. This likely prevents formation of biologically aberrant multi-layered capsids in the virion. The generation of single-layered hexamer tubes facilitated 3D helical image reconstruction from cryo-electron microscopy data, revealing the basic tube architecture.

biochemistry↗

bioRxiv2017

How to get the most bang for your buck: the evolution and physiology of nutrition-dependent resource allocation strategies

All organisms utilize resources to grow, survive, and reproduce. The supply of these resources varies widely across landscapes and time, imposing ultimate constraints on the maximal trait values for allocation-related traits. In this review, we address three key questions fundamental to our understanding of the evolution of allocation strategies and their underlying mechanisms. First, we ask: how diverse are flexible resource allocation strategies among different organisms? We find there are many, varied, examples of flexible strategies that depend on nutrition. However, this diversity is often ignored in some of the best-known cases of resource allocation shifts, such as the commonly observed pattern of lifespan extension under nutrient limitation. A greater appreciation of the wide variety of flexible allocation strategies leads directly to our second major question: what conditions select for different plastic allocation strategies? Here, we highlight the need for additional models that explicitly consider the evolution of phenotypically plastic allocation strategies and empirical tests of the predictions of those models in natural populations. Finally, we consider the question: what are the underlying mechanisms determining resource allocation strategies? Although evolutionary biologists assume differential allocation of resources is a major factor limiting trait evolution, few proximate mechanisms are known that specifically support the model. We argue that an integrated framework can reconcile evolutionary models with proximate mechanisms that appear at first glance to be in conflict with these models. Overall, we encourage future studies to 1) mimic ecological conditions in which those patterns evolve, and 2) take advantage of the omic opportunities to produce multi-level data and analytical models that effectively integrate across physiological and evolutionary theory.

evolutionary biology↗

bioRxiv2017

Bacterial physiological adaptations to contrasting edaphic conditions identified using landscape scale metagenomics

Environmental factors relating to soil pH are widely known to be important in structuring soil bacterial communities, yet the relationship between taxonomic community composition and functional diversity remains to be determined. Here, we analyze geographically distributed soils spanning a wide pH gradient and assess the functional gene capacity within those communities using whole genome metagenomics. Low pH soils consistently had fewer taxa (lower alpha and gamma diversity), but only marginal reductions in functional alpha diversity and equivalent functional gamma diversity. However, coherent changes in the relative abundances of annotated genes between pH classes were identified; with functional profiles clustering according to pH independent of geography. Differences in gene abundances were found to reflect survival and nutrient acquisition strategies, with organic-rich acidic soils harboring a greater abundance of cation efflux pumps, C and N direct fixation systems and fermentation pathways indicative of anaerobiosis. Conversely, high pH soils possessed more direct transporter-mediated mechanisms for organic C and N substrate acquisition. These findings show that bacterial functional versatility may not be constrained by taxonomy, and we further identify the range of physiological adaptations required to exist in soils of varying nutrient availability and edaphic conditions.

microbiology↗

bioRxiv2017

Physiological Preparation Of Hair Cells From The Sacculus Of The American Bullfrog (Rana catesbeiana)

SHORT ABSTRACTThe American bullfrogs sacculus permits direct examination of hair-cell physiology. Here we describe the dissection and preparation of the bullfrogs sacculus for biophysical studies. We show representative experiments from these hair cells, including the calculation of a bundles force-displacement relation and measurement of its spontaneous motion.\n\nLONG ABSTRACTThe study of hearing and balance rests upon insights drawn from biophysical studies of model systems. One such model, the sacculus of the American bullfrog, has become a mainstay of auditory and vestibular research. Studies of this organ have revealed how sensory cells--hair cells--can actively detect signals from the environment. Because of these studies, we now better understand the mechanical gating and localization of a hair cells transduction channels, calciums role in mechanical adaptation, and the identity of hair-cell currents. This highly accessible organ continues to provide insight into the workings of hair cells.\n\nHere we describe the preparation of the bullfrogs sacculus for biophysical studies on its hair cells. We include the complete dissection procedure and provide specific protocols for the preparation of the sacculus in specific contexts. We additionally include representative results using this preparation, including the calculation of a hair bundles force-displacement relation and measurement of a bundles spontaneous motion.

biophysics↗

bioRxiv2017

Mitochondria Are Physiologically Maintained At Close To 50 C

In endothermic species, heat released as a product of metabolism ensures stable internal temperature throughout the organism, despite varying environmental conditions. Mitochondria are major actors in this thermogenic process. Part of the energy released by the oxidation of respiratory substrates drives ATP synthesis and metabolite transport, while a noticeable proportion is released as heat. Using a temperature-sensitive fluorescent probe targeted to mitochondria, we measured mitochondrial temperature in situ under different physiological conditions. At a constant external temperature of 38 {degrees}C, mitochondria were more than 10 {degrees}C warmer when the respiratory chain was fully functional, both in HEK293cells and primary skin fibroblasts. This differential was abolished in cells lacking mitochondrial DNA or by respiratory inhibitors, but preserved or enhanced by expressing thermogenic enzymes such as the alternative oxidase or the uncoupling protein 1. The activity of various RC enzymes was maximal at, or slightly above, 50 {degrees}C. Our study prompts a re-examination of the literature on mitochondria, taking account of the inferred high temperature.

cell biology↗

bioRxiv2017

Physiological Assembly Of Functionally Active 30S Ribosomal Subunits From In Vitro Synthesized Parts

Synthetic ribosomes in vitro can facilitate engineering translation of novel polymers, identifying ribosome biogenesis central components, and paving the road to constructing replicating systems from defined biochemical components. Here, we report functional synthetic Escherichia coli 30S ribosomal subunits constructed using a defined, purified cell free system under physiological conditions. We test hypotheses about key components of natural ribosome biogenesis pathway as required for efficient function - including integration of 16S rRNA modification, cofactors facilitated ribosome assembly and protein synthesis in the same compartment in vitro. We observe ~17% efficiency for fully synthetic 30S and ~70% efficiency from in vitro transcribed 16S rRNA assembled with natural proteins. We observe up to 5 fold improvement over previous crude extracts. We suggest extending the minimal list of components required for central-dogma replication from the 151 gene products previously reported to at least 180 to allow the speed and accuracy of macromolecular synthesis to approach native E. coli values.

biochemistry↗

bioRxiv2017

Aminoglycoside Antibiotics Perturb Physiologically Important MicroRNA Contributing To Drug Toxicity

miRNAs are key non-protein coding regulators of gene expression in various pathophysiological conditions. Targeting miRNA with small molecules offer an unconventional approach, where clinically active compounds with RNA binding activity can be tested for their ability to modulate miRNA levels and thus for drug repositioning. Aminoglycoside antibiotics are highly effective microbicidal RNA binding molecules that bind to prokaryotic rRNA secondary structures. Here, we report that specific subsets of miRNA can be modulated by aminoglycosides. However, ototoxicity (cochlear and vestibular) and nephrotoxicity of multiple origins resulting from prolonged use are a well-known disadvantage of aminoglycosides. Mature non-coding RNAs and their precursors can present off-target sites, by forming secondary structures that resemble ribosomal RNA, thus providing an additional molecular basis for the toxicity of aminoglycosides. Using in vitro, in cellulae and physiological responses, we provide evidence for the direct functional perturbation of the miR- 96 cluster leading to selective cell death in neuromasts- the zebrafish equivalent of cochlear hair cells, by Streptomycin, a prototype aminoglycoside antibiotic, thus contributing to the observed ototoxicity. Our observations, collectively underscore the importance of re- evaluating RNA binding drugs for their off-targeting effects in the context of miRNA and other functional non-coding RNA.

cell biology↗

bioRxiv2017

Physiological tolerance of the early life history stages of fresh water prawn (Macrobrachium rosenbergii De Man, 1879) to environmental stress

Global climate change is transforming life on earth, causing widespread effects on all ecosystems. Among marine ecosystems, estuaries are considered as nursery grounds for marine and fresh water species. M. rosenbergii, a euryhaline species, migrate to the estuaries for breeding and spawning. The subsequent larval rearing takes place by experiencing variations in temperature and salinity conditions. The present study examines the effect of different temperature and salinity on the larval development and survival by observations on stored yolk utilization, cardiac performance, as well as changes in the rate of growth in body appendages and larval activity. The larvae showed 100% mortality at higher temperature (33.5 {+/-} 0.5 {degrees}C) in all the salinity conditions (12 PPT, 15 PPT, and 20 PPT). The survival rate varied between 76-96 % on exposure to lesser temperature conditions. Likewise, the post-embryonic yolk lasted for 4 days at ambient temperature (29 {degrees}C); whereas, at 33.5 {+/-} 0.5 {degrees}C, it lasted only for 2-3 days. There was an increase in total length of larvae, when exposed to higher temperature and salinity, independently or in combination, but at 33.5 {+/-} 0.5 {degrees}C under all salinity conditions the larvae died on the 5th day. For the cardiac performance, larval heart beat (fH) significantly increased for higher temperature and salinity conditions (20 PPT; 33.5 {degrees}C) and lowered at ambient condition 12 PPT; 29{degrees}C. Larval stroke volume Vs, and Cardiac output Q{square} were higher in ambient conditions and lowest in higher temperature and salinity conditions. However, temperature and salinity together did not show any significant effect on cardiac performance. On the other hand, the larval activity decreased significantly at higher temperature and salinity conditions, compared to ambient conditions but the interactive effect did not show any change. Thus, the physiological responses to temperature and salinity by the early life stages of M. rosenbergii could restrain the tolerance capability of the organism, thereby interfering in the successful completion of the larval development under the altered climatic conditions.

developmental biology↗

bioRxiv2017

Characterization of a set of abdominal neuroendocrine cells that regulate stress physiology using colocalized diuretic peptides in Drosophila

Multiple neuropeptides are known to regulate water and ion balance in Drosophila melanogaster. Several of these peptides also have other functions in physiology and behavior. Examples are corticotropin-releasing factor-like diuretic hormone (diuretic hormone 44; DH44) and leucokinin (LK), both of which induce fluid secretion by Malpighian tubules (MTs), but also regulate stress responses, feeding, circadian activity and other behaviors. Here, we investigated the functional relations between the LK and DH44 signaling systems. DH44 and LK peptides are only colocalized in a set of abdominal neurosecretory cells (ABLKs). Targeted knockdown of each of these peptides in ABLKs leads to increased resistance to desiccation, starvation and ionic stress. Food ingestion is diminished by knockdown of DH44, but not LK, and water retention is increased by LK knockdown only. Thus, the two colocalized peptides display similar systemic actions, but differ with respect to regulation of feeding and body water retention. We also demonstrated that DH44 and LK have additive effects on fluid secretion by MTs. It is likely that the colocalized peptides are coreleased from ABLKs into the circulation and act on the tubules where they target different cell types and signaling systems to regulate diuresis and stress tolerance. Additional targets seem to be specific for each of the two peptides and subserve regulation of feeding and water retention. Our data suggest that the ABLKs and hormonal actions are sufficient for many of the known DH44 and LK functions, and that the remaining neurons in the CNS play other functional roles.

neuroscience↗

bioRxiv2017

Genome downsizing, physiological novelty, and the global dominance of flowering plants

During the Cretaceous (145-66 Ma), early angiosperms rapidly diversified, eventually outcompeting the ferns and gymnosperms previously dominating most ecosystems. Heightened competitive abilities of angiosperms are often attributed to higher rates of transpiration facilitating faster growth. This hypothesis does not explain how angiosperms were able to develop leaves with smaller, but densely packed stomata and highly branched venation networks needed to support increased gas exchange rates. Although genome duplication and reorganization have likely facilitated angiosperm diversification, here we show that genome downsizing facilitated reductions in cell size necessary to construct leaves with a high density stomata and veins. Rapid genome downsizing during the early Cretaceous allowed angiosperms to push the frontiers of anatomical trait space. In contrast, during the same time period ferns and gymnosperms exhibited no such changes in genome size, stomatal size, or vein density. Further reinforcing the effect of genome downsizing on increased gas exchange rates, we found that species employing water-loss limiting crassulacean acid metabolism (CAM) photosynthesis, have significantly larger genomes than C3 and C4 species. By directly affecting cell size and gas exchange capacity, genome downsizing brought actual primary productivity closer to its maximum potential. These results suggest species with small genomes, exhibiting a larger range of final cell size, can more finely tune their leaf physiology to environmental conditions and inhabit a broader range of habitats.

ecology↗

bioRxiv2019

Physiological and morphological correlates of extreme acid tolerance in larvae of the acidophilic amphibian Litoria cooloolensis (Anura: Pelodryadidae)

In the coastal sandy lowlands of east Australia, several anuran species including the Cooloola Sedgefrog Litoria cooloolensis, show remarkable tolerance to dilute highly acidic waters as low as pH 3.5. To investigate the physiological and morphological underpinnings of acid tolerance in L. cooloolensis larvae, we compared Na+ balance, uptake and efflux rates, and gill and skin morphology in larvae reared in circum-neutral (pH 6.5) and pH 3.5 water. We hypothesised that acute exposure to pH 3.5 water would cause an initial loss of ionic homeostasis in larvae, but with chronic exposure larvae would restore Na+ balance. Net Na+ flux rates were not significantly different from zero in larvae reared at pH 3.5 and in acid-na&iumlve animals maintained in pH 6.5 water. Animals reared at pH 6.5 and acutely exposed to pH 3.5 exhibited a net loss of Na+, due to a significant inhibition of Na+ uptake. In contrast, L. cooloolensis larvae reared at pH 3.5 maintained Na+ balance at pH 3.5 and did not exhibit inhibition of Na+ uptake at this pH. Investigation of Na+ transport kinetics and the morphology of the gills and integument suggests tolerance of L. cooloolensis larvae to low pH may be attributed to a high capacity for branchial Na+ uptake, increased tight junction length and elevated mucus production in the gills and integument. These factors confer resistance to acid damage and disruption of ionic homeostasis which would otherwise result in the death of larvae exposed to waters of pH 4.0 and less.

zoology↗

bioRxiv2020

Spatially organized genomic and physiological heterogeneity of the olfactory bulb mitral cell layer

Sensory inputs to the dorsal vs ventral olfactory bulb derive from distinct receptor families, and drive distinct behaviors. To address whether second-order OB neurons and circuits exhibit matching heterogeneity for input-specific readout, we clustered spatial expression profiles of >2,000 genes from the mitral cell layer (MCL). We observed clear dorsal and ventral clusters, together with dorsoventral differences in mitral cell physiology. Bulbar circuits may therefore be tuned for zone-specific computation.

neuroscience↗

bioRxiv2020

Physiological significance of proteolytic processing of Reelin revealed by cleavage-resistant Reelin knock-in mice

Reelin is a secreted protein that plays versatile roles in neuronal development and function, and hypoactivity of Reelin is implicated in many neuropsychiatric disorders. The strength of Reelin signaling is regulated by proteolytic processing, but its importance in vivo is not yet fully understood. Here, we generated Reelin knock-in (PA-DV KI) mice in which the key cleavage site of Reelin was abolished by mutation. As expected, the cleavage of Reelin was severely abrogated in the cerebral cortex and hippocampus of PA-DV KI mice. The amount of Dab1, whose degradation is induced by Reelin signaling, decreased in these tissues, indicating that the signaling strength of Reelin was augmented. The brains of PA-DV KI mice were largely structurally normal, but unexpectedly, the hippocampal layer was disturbed. This phenotype was ameliorated in hemizygote PA-DV KI mice, indicating that excess Reelin signaling is detrimental to hippocampal layer formation. The neuronal dendrites of PA-DV KI mice had more branches and were elongated compared to wild-type mice. These results present the first direct evidence of the physiological importance of Reelin cleavage and suggest that inhibition of Reelin cleavage would counteract neuropsychiatric disorders without causing severe systemic side effects.

neuroscience↗