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Results for “Physiology”

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 Read source abstracts and check manuscript versions; preprints are not peer reviewed.

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bioRxiv2017

Individual differences in frequency and topography of slow and fast sleep spindles

Sleep spindles are transient oscillatory waveforms occurring during non-rapid eye movement (NREM) sleep and are implicated in plasticity and memory processes. In humans, spindles can be classified as either slow or fast, but large individual differences in spindle frequency as well as methodological difficulties have hindered progress towards understanding their function. Using two nights of high-density electroencephalography recordings from healthy individuals, we first characterize the individual variability of NREM spectra and demonstrate the difficulty of determining subject-specific spindle frequencies. We then introduce a novel spatial filtering approach that can reliably separate subject-specific spindle activity into slow and fast components that are stable across nights and across N2 and N3 sleep. We then proceed to provide detailed analyses of the topographical expression of individualized slow and fast spindle activity. Group-level analyses conform to known spatial properties of spindles, but also uncover novel differences between sleep stages and spindle classes. Moreover, subject-specific examinations reveal that individual topographies show considerable variability that is stable across nights. Finally, we demonstrate that topographical maps depend nontrivially on the spindle metric employed. In sum, our findings indicate that group-level approaches mask substantial individual variability of spindle dynamics, in both the spectral and spatial domains. We suggest that leveraging, rather than ignoring, such differences may prove useful to further our understanding of the physiology and functional role of sleep spindles.

neuroscience↗

bioRxiv2017

Adaptation to chronic malnutrition leads to reduced dependence on microbiota in Drosophila

Numerous studies have shown that animal nutrition is tightly linked to gut microbiota, especially under nutritional stress. In Drosophila, microbiota are known to promote juvenile growth, development and survival on poor diets, mainly through enhanced digestion leading to changes in hormonal signaling. Here we show that this reliance on microbiota is greatly reduced in replicated Drosophila populations that adapted to a poor larval diet in the course of over 170 generations of experimental evolution. Protein and polysaccharide digestion in these malnutrition-adapted populations became much less dependent on colonization with microbiota. This was accompanied by changes in at least some targets of dFOXO transcription factor, which is a key regulator of cell growth and survival. Our study suggests that some metazoans have retained the evolutionary potential to adapt their physiology such that association with microbiota may become optional rather than essential.

evolutionary biology↗

bioRxiv2017

Connexin43 controls N-cadherin transcription during collective cell migration

Connexins are the primary components of gap junctions, providing direct links between cells in many physiological processes, including cell migration and cancer metastasis. Exactly how cell migration is controlled by gap junctions remains a mystery. To shed light on this, we investigated the role of Connexin43 in collective cell migration during embryo development using the neural crest, an embryonic cell population whose migratory behavior has been likened to cancer invasion. We discovered that Connexin43 is required for contact inhibition of locomotion by directly regulating the transcription of N-cadherin. For this function, the Connexin43 carboxy tail interacts with Basic Transcription Factor 3, which mediates its translocation to the nucleus. Together, they bind to the n-cad promotor regulating n-cad transcription. Thus, we uncover an unexpected, gap junction-independent role for Connexin43 in collective migration that illustrates the possibility that connexins, in general, may be important for a wide variety of cellular processes that we are only beginning to understand.\n\nHighlightsO_LICx43 regulates collective directional migration of neural crest cells\nC_LIO_LICx43 carboxy tail controls cell polarity via n-cad regulation\nC_LIO_LICx43 carboxy tail localises at the nucleus and that depends on BTF3\nC_LIO_LIBTF3 and Cx43 carboxy tail directly interact to bind and regulate n-cad promoter activity\nC_LI

developmental biology↗

bioRxiv2017

Differences in the spatial and temporal patterns of head motion during MRI of adults and infants

AimHead motion has a profound effect on MRI, and will contaminate comparisons of function or structure between groups that move differently. This work compares adults and infants. Infants might move differently for physical, physiological and cognitive reasons, but so far these differences have not been quantified.\n\nMethodsThe spatial modes and total magnitude of motion in the MRI scanner were measured (N=211). The effects of group (infant vs. adult) and stimulation paradigm (auditory vs. visual) were evaluated.\n\nResultsSpatial modes of motion were found to be distinct between infant and adult groups. Infants had less anterior-posterior translational motion, but greater motion in other dimensions, often with complex multi-axis patterns. In magnitude distribution, sleeping infants often remained more still than adults, but when movement did occur it was more extreme and abrupt. Two groups of adults presented with different stimulation showed similar shapes of motion.\n\nConclusionThe spatial modes and magnitude distribution of motion differed substantially between groups, and must be considered carefully as a confound in comparisons of structure or function. The abruptness and magnitude of movement suggests that for infants relative to adults post-processing strategies such as de-noising are likely to be more effective than prospective motion correction.\n\nKey notesO_LIQuantified the spatial and temporal distribution of motion during MRI in 211 adults and neonates\nC_LIO_LIThe different spatial modes in adults and infants were visualized and statistically contrasted\nC_LIO_LIThe magnitude of motion had \"heavier tails\" in infants, with more still periods, and more large movements, than adults.\nC_LI

neuroscience↗

bioRxiv2017

Metabolic coupling in bacteria

Bacteria frequently engage in cross-feeding interactions that involve an exchange of metabolites with other micro- or macroorganisms. The often obligate nature of these associations, however, hampers manipulative experiments, thus limiting our mechanistic understanding of the ecophysiological consequences that result for the organisms involved. Here we address this issue by taking advantage of a well-characterised experimental model system, in which auxotrophic genotypes of E. coli derive essential amino acid from prototrophic donor cells using intercellular nanotubes. Surprisingly, donor-recipient cocultures revealed that the mere presence of auxotrophic genotypes in coculture was sufficient to increase amino acid production levels in donor cells. Subsequent experiments unravelled that this effect was due to the depletion of amino acid concentrations in the cytoplasm of donor cells, which delayed feedback inhibition of the corresponding amino acid biosynthetic pathway. This finding indicates that in newly established mutualistic associations, an intercellular regulation of exchanged metabolites can simply emerge from the architecture of the underlying biosynthetic pathways, rather than through the evolution of new regulatory mechanisms. Taken together, our results show that a single loss-of-function mutation can physiologically couple the metabolism of two cross-feeding cells in a source-sink-like relationship.

microbiology↗

bioRxiv2017

Managed woodlot revealed a trade-off between edible leaves and timber production in Vitex doniana Sweet (Lamiaceae)

Vitex doniana Sweet is a major wild-harvested tree resource for food in Benin. However, the species is under threats characterised by increasing human pressure on remnant populations. This study represents the first to explore species response to biotic stress. We tested the response of V. doniana to coppicing and fertilization. Two stump heights (20 and 40 cm) in combination with three organic manure rates (0.5; 1 and 1.5 kg per seedling), with eight replicates were tested in a randomised complete block design. We used mixed effect models with pseudoreplication, and the maximum likelihood method to compare effects of fixed factors on sprouting vigour, sprout growth and biomass yield in the short (12 months) and medium (5 years) terms. Results indicated that stump height significantly affected sprouting and all growth parameters, in the short and medium terms. However, there seemed a delayed effect of manure. We found initial seedling growth also an important factor. The hidden effect of stump height on biomass yield is discussed. Findings clearly indicate a trade-off between edible leaves and timber production by managed woodlot. Implications of findings for further investigation of above and below ground biomass dynamics and resources allocation in treated trees are discussed.\n\nHighlightsA clear trade-off between edible leaves and timber production is observed in managed Vitex doniana Sweet woodlot. Coppicing as a biotic stress induced important physiological changes that merit further investigations.

plant biology↗

bioRxiv2017

Synaptic and peptidergic connectome of a neurosecretory centre in the annelid brain

Neurosecretory centres in animal brains use peptidergic signalling to influence physiology and behaviour. Understanding neurosecretory centre function requires mapping cell types, synapses, and peptidergic networks. Here we use electron microscopy and gene expression mapping to analyse the synaptic and peptidergic connectome of an entire neurosecretory centre. We mapped 78 neurosecretory neurons in the brain of larval Platynereis dumerilii, a marine annelid. These neurons form an anterior neurosecretory organ expressing many neuropeptides, including hypothalamic peptide orthologues and their receptors. Analysis of peptide-receptor pairs revealed sparsely connected networks linking specific neuronal subsets. We experimentally analysed one peptide-receptor pair and found that a neuropeptide can couple neurosecretory and synaptic brain signalling. Our study uncovered extensive non-synaptic signalling within a neurosecretory centre and its connection to the synaptic brain.

neuroscience↗

bioRxiv2017

Life and death of proteins after protease cleavage: protein degradation by the N-end rule pathway

The activity and abundance of proteins within a cell are controlled precisely to ensure the regulation of cellular and physiological processes. In eukaryotes, this can be achieved by targeting specific proteins for degradation by the ubiquitin-proteasome system. The N-end rule pathway, a subset of the ubiquitin-proteasome system, targets proteins for degradation depending on the identity of a protein N-terminal residue or its post-translational modifications. Here, we discuss the most recent findings on the diversity of N-end rule pathways. We also focus on recently found defensive functions of the N-end rule pathway in plants. We then discuss the current understanding of N-end rule substrate formation by protease cleavage. Finally, we review state-of-the-art proteomics techniques used for N-end rule substrate identification, and discuss their usefulness and limitations for the discovery of the molecular mechanisms underlying the roles of the N-end rule pathway in plants.

plant biology↗

bioRxiv2017

Dissection of MAPK signaling specificity through protein engineering in a developmental context

Mitogen-activated protein kinases (MAPK) signaling affects many processes, some of which have different outcomes in the same cell. In Arabidopsis, activation of a MAPK cascade consisting of YODA, MKK4/5 and MPK3/6 inhibits early stages of stomatal developmental, but this ability is lost at the latest stage when guard mother cells (GMCs) transition to guard cells (GCs). Rather than downregulating cascade components, stomatal precursors must have a mechanism to prevent late stage inhibition because the same MKKs and MPKs mediate other physiological responses. Here, we artificially activated the MAPK cascade using MKK7, another MKK that can modulate stomatal development, and found that inhibition of stomatal development is still possible in GMCs. This suggests that MKK4/5, but not MKK7, are specifically prevented from inhibiting stomatal development. To identify regions of MKKs responsible for cell-type specific regulation, we used a domain swap approach with MKK7 and a battery of in vitro and in vivo kinase assays. We found that N-terminal regions of MKK5 and MKK7 establish specific signal-to-output connections like they do in other organisms, but they do so in combination with previously undescribed modules in the C-terminus. One of these modules encodes the GMC-specific regulation of MKK5, that when swapped with MKK7s, allows MKK5 to mediate robust inhibition of late stomatal development. Because MKK structure is conserved across species, the identification of new MKK specificity modules and signaling rules furthers our understanding of how eukaryotes create specificity in complex biological systems.

plant biology↗

bioRxiv2017

A homozygous missense mutation in ERAL1, encoding a mitochondrial rRNA chaperone, causes Perrault syndrome

Perrault syndrome (PS) is a rare recessive disorder characterized by ovarian dysgenesis and sensorineural deafness. It is clinically and genetically heterogeneous, and previously mutations have been described in different genes, mostly related to mitochondrial proteostasis. We diagnosed three unrelated females with PS and set out to identify the underlying genetic cause using exome sequencing. We excluded mutations in the known PS genes, but identified a single homozygous mutation in the ERAL1 gene (c.707A>T; p.Asn236Ile). Since ERAL1 protein binds to the mitochondrial 12S rRNA and is involved in the assembly of the small mitochondrial ribosomal subunit, the identified variant represented a likely candidate. In silico analysis of a 3D model for ERAL1 suggested that the mutated residue hinders protein-substrate interactions, potentially affecting its function. On a molecular basis, PS skin fibroblasts had reduced ERAL1 protein levels. Complexome profiling of the cells showed an overall decrease in the levels of assembled small ribosomal subunit, indicating that the ERAL1 variant affects mitochondrial ribosome assembly. Moreover, levels of the 12S rRNA were reduced in the patients, and were fully rescued by lentiviral expression of wild type ERAL1. At the physiological level, mitochondrial respiration was markedly decreased in PS fibroblasts, confirming disturbed mitochondrial function. Finally, knockdown of the C. elegans ERAL1 homologue E02H1.2 almost completely blocked egg production in worms, mimicking the compromised fertility in PS-affected women. Our cross-species data in patient cells and worms support the hypothesis that mutations in ERAL1 can cause PS and are associated with changes in mitochondrial metabolism.

genetics↗

bioRxiv2017

The Co-regulation Data Harvester for Tetrahymenathermophila: automated high-throughput geneannotation and functional inference in a microbialeukaryote

Identifying co-regulated genes can provide a useful approach for defining pathway-specific machinery in an organism. To be efficient, this approach relies on thorough genome annotation, which is not available for most organisms with sequenced genomes. Studies in Tetrahymena thermophila, the most experimentally accessible ciliate, have generated a rich transcriptomic database covering many well-defined physiological states. Genes that are involved in the same pathway show significant co-regulation, and screens based on gene co-regulation have identified novel factors in specific pathways, for example in membrane trafficking. However, a limitation has been the relatively sparse annotation of the Tetrahymena genome, making it impractical to approach genome-wide analyses. We have therefore developed an efficient approach to analyze both co-regulation and gene annotation, called the Co-regulation Data Harvester (CDH). The CDH automates identification of co-regulated genes by accessing the Tetrahymena transcriptome database, determines their orthologs in other organisms via reciprocal BLAST searches, and collates the annotations of those orthologs' functions. Inferences drawn from the CDH reproduce and expand upon experimental findings in Tetrahymena. The CDH, which is freely available, represents a powerful new tool for analyzing cell biological pathways in Tetrahymena. Moreover, to the extent that genes and pathways are conserved between organisms, the inferences obtained via the CDH should be relevant, and can be explored, in many other systems.

bioinformatics↗

bioRxiv2017

Whole genome sequences of the raspberry and strawberry pathogens Phytophthora rubi and P. fragariae

Phytophthora rubi and P. fragariae are two closely related oomycete plant pathogens that exhibit strong morphological and physiological similarities, but are specialized to infect different hosts of economic importance, namely raspberry and strawberry. Here, we report the draft genome sequences of these two Phytophthora species as a first step towards understanding the genomic processes underlying plant host adaptation in these pathogens.

genomics↗

bioRxiv2017

CaMK (CMK-1) and O-GlcNAc transferase (OGT-1) modulate mechanosensory responding and habituation in an interstimulus interval-dependent manner in Caenorhabditis elegans

The ability to learn is an evolutionarily conserved adaptation that remains incompletely understood. Genetically tractable model organisms facilitate mechanistic explanations of learning that span genetic, neural circuit, and behavioural levels. Many aspects of neural physiology, including processes that underlie learning (e.g. neurotransmitter release and long-lasting changes in synaptic strength), are regulated by brief and local changes in [m] levels of free intracellular Ca2+. On this scale, changes in [Ca2+] activate many Ca2+-sensors, including the Ca2+/calmodulin-dependent kinases (CaMKs). Here we reveal that the Caenorhabditis elegans ortholog of CaMK1/4, CMK-1, functions in primary sensory neurons to regulate responses to mechanical stimuli and behavioral plasticity, specifically habituation, a conserved form of non-associative learning. The habituation phenotypes of cmk-1 mutants were dependent on interstimulus interval (ISI), such that CMK-1 slows habituation at short ISIs, but promotes it at long ISIs. We predicted potential CaMK phosphorylation targets from catalytic site analysis of the human and C. elegans CaMKs and mutant analysis of these candidates implicated O-linked N-acetylglucosamine (O-GlcNAc) transferase, OGT-1, in mechanosensitivity and learning. Cell specific rescue and knockdown experiments showed that both CMK-1 and OGT-1 function cell autonomously in mechanosensory neurons to modulate learning. Interestingly, despite their similar mutant phenotypes, detailed behavioral analysis of double mutants demonstrated that CMK-1 and OGT-1 act in parallel genetic pathways. Our research identifies CMK-1 and OGT-1 as co-expressed yet independent regulators of mechanosensitivity and learning.

animal behavior and cognition↗

bioRxiv2017

Compositionally distinct nuclear pore complexes of functionally distinct dimorphic nuclei in ciliate Tetrahymena

SUMMARY STATEMENTOur study demonstrates compositional and structural differences of the nuclear pore complex between the functionally differentiated macronucleus and micronucleus within a single cytoplasm of ciliated protozoa.\n\nABSTRACTThe nuclear pore complex (NPC), a gateway for nucleocytoplasmic trafficking, is composed of about 30 different proteins called nucleoporins. It remains unknown whether the NPCs within a species are homogeneous or vary depending on the cell type, or physiological condition. Here, we present evidence for compositionally distinct NPCs that form within a single cell in a binucleated ciliate. In Tetrahymena thermophila, each cell contains both a transcriptionally-active macronucleus (MAC) and a germline micronucleus (MIC). By combining in silico analysis, mass spectrometry analysis for immuno-isolated proteins, and subcellular localization analysis of GFP fused proteins, we identified numerous novel components of MAC and MIC NPCs. Core members of the Nup107-160 scaffold complex were enriched in MIC NPCs. Strikingly, two paralogs of Nup214 and of Nup153 localized exclusively to either MAC or MIC NPCs. Furthermore, the transmembrane components Pom121 and Pom82 localize exclusively to MAC and MIC NPCs, respectively. Our results argue that functional nuclear dimorphism in ciliates is likely to depend on compositional and structural specificity of NPCs.

cell biology↗

bioRxiv2017

Live Tracking Of Moving Samples In Confocal Microscopy For Vertically Grown Plant Roots

Roots navigate through soil integrating environmental signals to orient their growth. The Arabidopsis root is a widely used model for developmental, physiological and cell biological studies. Live imaging greatly aids these efforts, but the horizontal sample position and continuous root tip displacement present significant difficulties. Here, we develop a confocal microscope setup for vertical sample mounting and integrated directional illumination. We present TipTracker - a custom software for automatic tracking of diverse moving objects usable on various microscope setups. Combined, this enables observation of root tips growing along the natural gravity vector over prolonged periods of time, as well as the ability to induce rapid gravity or light stimulation. We also track migrating cells in the developing zebrafish embryo, demonstrating the utility of this system in the acquisition of high resolution data sets of dynamic samples. We provide detailed descriptions of the tools enabling the easy implementation on other microscopes.

plant biology↗

bioRxiv2017

Calcium oscillations in wounded fibroblast monolayers are spatially regulated through substrate mechanics

The maintenance of tissue integrity is essential for the life of multicellular organisms. Healing of a skin wound is a paradigm for how various cell types localize and repair tissue perturbations in an orchestrated fashion. To investigate biophysical mechanisms associated with wound localization, we focus on a model system consisting of a fibroblast monolayer on an elastic substrate. We find that the creation of an edge in the monolayer causes cytosolic calcium oscillations throughout the monolayer. The oscillation frequency increases with cell density, which shows that wound-induced calcium oscillations occur collectively. Inhibition of myosin II reduces the number of oscillating cells, demonstrating a coupling between actomyosin activity and calcium response. The spatial distribution of oscillating cells depends on the stiffness of the substrate. For soft substrates with a Youngs modulus E ~ 360 Pa, oscillations occur on average within 0.2 mm distance from the wound edge. Increasing substrate stiffness leads to an average localization of oscillations away from the edge (up to ~0.6 mm). In addition, we use traction force microscopy to determine stresses between cells and substrate. We find that an increase of substrate rigidity leads to a higher traction magnitude. For E < ~2 kPa, the traction magnitude is strongly concentrated at the monolayer edge, while for E > ~8 kPa, traction magnitude is on average almost uniform beneath the monolayer. Thus, the spatial occurrence of calcium oscillations correlates with the cell-substrate traction. Overall, the experiments with fibroblasts demonstrate a collective, chemomechanical localization mechanism at the edge of a wound with a potential physiological role.

biophysics↗

bioRxiv2017

Breathlessness And The Body: Neuroimaging Evidence For The Inferential Leap

Breathlessness debilitates millions of people with chronic illness. Mismatch between breathlessness severity and objective disease markers is common and poorly understood. Traditionally, sensory perception was conceptualised as a stimulus-response relationship, although this cannot explain how conditioned symptoms may occur in the absence of physiological signals from the lungs or airways. A Bayesian model is now proposed in which the brain generates sensations based on expectations learned from past experiences (priors), which are then checked against incoming afferent signals. In this model, psychological factors may act as moderators. They may either alter priors, or change the relative attention towards incoming sensory information, leading to more variable interpretation of an equivalent afferent input.\n\nIn the present study we conducted a preliminary test of this model in a supplementary analysis of previously published data (Hayen 2017). We hypothesised that individual differences in psychological traits (anxiety, depression, anxiety sensitivity) would correlate with the variability of subjective evaluation of equivalent breathlessness challenges. To better understand the resulting inferential leap in the brain, we explored whether these behavioural measures correlated with activity in areas governing either prior generation or sensory afferent input.\n\nBehaviorally, anxiety sensitivity was found to positively correlate with each subjects variability of intensity and unpleasantness during mild breathlessness, and with unpleasantness during strong breathlessness. In the brain, anxiety sensitivity was found to positively correlate with activity in the anterior insula during mild breathlessness, and negatively correlate with parietal sensorimotor areas during strong breathlessness.\n\nOur findings suggest that anxiety sensitivity may reduce the robustness of this Bayesian sensory perception system, increasing the variability of breathlessness perception and possibly susceptibility to symptom misinterpretation. These preliminary findings in healthy individuals demonstrate how differences in psychological function influence the way we experience bodily sensations, which might direct us towards better understanding of symptom mismatch in clinical populations.

neuroscience↗

bioRxiv2017

Intracellular production of hydrogels and synthetic RNA granules by multivalent enhancers

Non-membrane bound, hydrogel-like entities, such as RNA granules, nucleate essential cellular functions through their unique physico-chemical properties. However, these intracellular hydrogels have not been as extensively studied as their extracellular counterparts, primarily due to technical challenges in probing these materials in situ. Here, by taking advantage of a chemically inducible dimerization paradigm, we developed iPOLYMER, a strategy for rapid induction of protein-based hydrogels inside living cells. A series of biochemical and biophysical characterizations, in conjunction with computational modeling, revealed that the polymer network formed in the cytosol resembles a physiological hydrogel-like entity that behaves as a size-dependent molecular sieve. We studied several properties of the gel and functionalized it with RNA binding motifs that sequester polyadenine-containing nucleotides to synthetically mimic RNA granules. Therefore, we here demonstrate that iPOLYMER presents a unique and powerful approach to synthetically reconstitute hydrogel-like structures including RNA granules in intact cells.

synthetic biology↗