Biology subjects

Mishra, P.

Publications and source records attributed to Mishra, P..

68 records · Page 4Linked to original sources

bioRxiv2020

Isolation, identification and functional characterization of cultivable bacteria from Arabian Sea and Bay of Bengal water samples reveals high diversity

The oxygen minimum zone of the Arabian Sea (AS) and Bay of Bengal (BOB) is rich in organic matter and is an unusual niche. Bacteria present in the oceanic water play an important role in ecology since they are responsible for decomposing, mineralizing of organic matter and in elemental cycling like nitrogen, sulfur, phosphate. This study focuses on culturing bacteria from oxygen minimum zones (OMZ) and non-OMZ regions and their phylogenetic as well as the functional characterization. Genotypic characterization of the isolates using amplified rDNA based 16SrRNA sequencing grouped them into various phylogenetic groups such as alpha-proteobacteria, gamma-proteobacteria and unaffiliated bacteria. The cultivable bacterial assemblages encountered belonged to the genus Halomonas, Marinobacter, Idiomarina, Pshyctobacter and Pseudoalteromonas. Among the enzymatic activities, carbohydrate utilization activity was most predominant (100%) and microorganisms possessed amylase, cellulase, xylanase and chitinase. A large proportion of these bacteria (60%) were observed to be hydrocarbon consuming and many were resistant to ampicillin, chloramphenicol, kanamycin and streptomycin. The high diversity and high percentage of extracellular hydrolytic enzyme activities along with hydrocarbon degradation activity of the culturable bacteria reflects their important ecological role in oceanic biogeochemical cycling. Further assessment confirmed the presence of nitrogen reduction capability in these cultivable bacteria which highlights their importance in oceanic geochemical cycling.

microbiology↗

bioRxiv2020

Sperm-specific COX6B2 Enhances Oxidative Phosphorylation, Proliferation, and Survival in Lung Adenocarcinoma

Cancer testis antigens (CTAs) are genes whose expression is normally restricted to the testis but anomalously activated in cancer. In sperm, a number of CTAs promote energy generation, however whether these proteins contribute to tumor cell metabolism is not understood. Here we describe COX6B2, a sperm-specific component of cytochrome c oxidase (complex IV). COX6B2 is frequently expressed in human lung adenocarcinoma (LUAD) and expression correlates with reduced survival time in patients. COX6B2, but not its somatic isoform COX6B1, enhances activity of complex IV, increasing mitochondrial oxidative phosphorylation (OXPHOS) and NAD+ generation. Consequently, COX6B2-expressing cells display a proliferative advantage, particularly in low oxygen conditions. Conversely, depletion of COX6B2 attenuates OXPHOS and collapses mitochondrial membrane potential leading to cell death or senescence. Furthermore, COX6B2 is both necessary and sufficient for growth of tumors in vivo. Our findings reveal a previously unappreciated, tumor specific metabolic pathway hijacked from one of the most ATP-intensive processes in the animal kingdom: sperm motility.

cancer biology↗

bioRxiv2020

The fly liquid-food electroshock assay (FLEA) reveals opposite roles for neuropeptide F in avoidance of bitterness and shock.

Proper regulation of feeding is important for an organisms well-being and survival. Food intake in Drosophila can be determined in a number of ways, including by measuring the time a flys proboscis interacts with a food source in the fly liquid-food interaction counter (FLIC). Here, we show that electrical current flowing through flies during this interaction is aversive and leads to a reduction in food intake. Based on the FLIC, we engineer a novel assay, the fly liquid-food electroshock assay (FLEA), which allows for current adjustments for each feeding well. Using the FLEA, we show that both external incentives as well as internal motivational state can serve as drivers for flies to overcome higher current (electric shock) to obtain superior food. Unlike similar assays in which bitterness is the aversive stimulus for the fly to overcome, we show that current perception is not discounted as flies become more food-deprived. The FLEA is therefore a novel assay to accurately measure incentive motivation in Drosophila. Using the FLEA, we also show that neuropeptide F is required for proper perception or processing of an electroshock, a novel function for this neuropeptide involved in processing of external and internal stimuli. Significance StatementMany neuropsychiatric disorders, such as depression or addiction, are associated with alterations in motivated behavior. Assays measuring incentive motivation determine how driven an organism is to attain a goal, like food, or how attractive an incentive is. These tests require the animal to put effort into obtaining the reward, which can include physical work or overcoming an aversive stimulus. Such assays for Drosophila feeding have relied on flies overcoming bitterness to obtain their food. However, the perception of bitterness is discounted as flies become food deprived, confounding the interpretation. Here, we developed a novel assay that does not suffer from the same shortcomings and thus allows for more accurate assessments of incentive motivation in this widely used model organism.

neuroscience↗

bioRxiv2019

Mapping the growth effect of previously hidden ubiquitin alleles using an overexpression based mutational scan

Deep mutational scanning has emerged as a powerful, high throughput approach to determine the growth effect of thousands of alleles at once in bulk competition. However, to date only the growth effect of mutant alleles in isolation has been determined. Building off previous work, we have created a library of all possible single point mutations in ubiquitin and determined the growth effect of mutants overexpressed in the presence of a wild type allele. Using this scan, we explained over half of the previously missing mutants in the single allele scan by showing that they exhibit deleterious effects when co-expressed with wild type ubiquitin. Additionally, unlike the single allele growth effect, these overexpression growth effects were distributed across the entire protein. This overexpression scan methodology can identify likely dominant mutant effects in any essential gene and is highly complementary with traditional deep mutational scanning approaches.

genetics↗

bioRxiv2019

Conjunctive changes in two distinct ion channels induce activity-dependent intrinsic plasticity in rat dentate gyrus granule cells

The dentate gyrus (DG) was the first brain region to provide insights about synaptic and intrinsic plasticity. However, the assessment of intrinsic plasticity in DG has been surprisingly limited. We employed whole-cell patch-clamp recordings to explore the impact of behaviorally relevant theta-modulated burst firing, in the absence of synaptic stimulation, on intrinsic properties of rat DG granule cells. We found that theta burst firing induced a significant reduction in sub-threshold excitability and temporal summation, accompanied by an unexpectedly contrasting enhancement of action potential firing rate. We show that conjunctive changes in HCN, inward-rectifier potassium and persistent sodium channels mediated this form of plasticity, which was dependent on calcium influx through L-type calcium channels and inositol trisphosphate receptors. Our results unveil the expression of conjunctive plasticity in multiple channels, responding to the same activity pattern, establishing a plasticity manifold that could concomitantly mediate encoding and homeostasis in DG engram cells.

neuroscience↗

bioRxiv2019

Microbial diversity of the Arabian Sea in the Oxygen minimum zones by metagenomics approach

Large oxygen depleted areas known as oxygen minimum zones (OMZ) have been observed in the Arabian Sea and recent reports indicate that these areas are expanding at an alarming rate. In marine waters, oxygen depletion may also be related to global warming and the temperature rise, acidification and deoxygenation can lead to major consequences wherein the plants, fish and other biota will struggle to survive in the ecosystem.\n\nThe current study has identified the microbial community structure using NGS based metagenomics analysis in the water samples collected at different depth from the oxygen depleted and non-OMZ areas of Arabian Sea. Environmental variables such as depth, site of collection and oxygen concentration appeared to influence the species richness and evenness among microbial communities in these locations. Our observations clearly indicate that population dynamics of microbes consisting of nitrate reducers accompanied by sulphate reducers and sulphur oxidizers participate in the interconnected geochemical cycles of the OMZ areas. In addition to providing baseline data related to the diversity and microbial community dynamics in oxygen-depleted water in the OMZ; such analysis can provide insight into processes regulating productivity and ecological community structure of the ocean.

microbiology↗

bioRxiv2019

Ion-channel regulation of response decorrelation in a multi-scale model of the dentate gyrus

The dentate gyrus (DG) is uniquely endowed with multiple forms of biological heterogeneities owing to the expression of adult neurogenesis and sparse connectivity, and has been functionally implicated in response decorrelation and pattern separation. Although channel decorrelation could be achieved through synergistic interactions between these heterogeneities, the impact of individual ion channels on channel decorrelation has not been explored. Here, to systematically assess the cascading impact of molecular-scale (ion channel) perturbations on network-scale outcomes (decorrelation), we first quantified the impact of eliminating individual ion channels on single-cell physiology of heterogeneous populations of granule cells (GCs) and basket cells (BCs). Employing virtual knockout simulations involving both populations, we found that the mapping between ion channels and nine distinct physiological measurements was many-to-many. Next, to assess the impact of ion channel elimination on channel decorrelation, we employed a conductance-based multi-scale network model of the DG. This network was endowed with four distinct forms of heterogeneities (intrinsic, synaptic, structural and afferent), with afferent inputs from the entorhinal cortices driven by virtual arena traversal. We show that individual ion channels expressed in GCs govern DG network excitability, and critically regulate the ability of the network to perform channel decorrelation. The impact of eliminating individual ion channels on network excitability and channel decorrelation was differential and variable, with local heterogeneities playing a pivotal role in determining the strength of such impact. Specifically, in the presence of structurally immature neurons in the DG network, the impact of ion channel elimination on channel decorrelation was considerably lower when compared with a network exclusively constructed with structurally mature neurons. Finally, we show that for any given ion channel knockout, the average percentage change in output correlation was invariant to the specific values of input correlation, across different network configurations endowed with disparate structural and afferent heterogeneities. Our analyses emphasizes that the mapping between components and function is many-to-many across scales, and assign critical roles for biological heterogeneities in conferring multi-scale functional robustness in the face of physiological and pathological perturbations.\n\nSIGNIFICANCE STATEMENTThere are precise sets of computation spanning different scales of analyses that drive behavioral states and responses of an animal. Perturbations to components that drive these computations at one scale could result in a cascading effect that alters physiological properties across several scales. Multi-scale computational models that account for biological heterogeneities at each scale are ideal tools to approach this complex problem, where systematic analyses of such cascades could be rigorously accomplished. Here, we systematically assessed the impact of eliminating individual ion channels, first on neuronal intrinsic properties, and consequently on network excitability and response decorrelation. Our results unveil important roles for biological heterogeneities in conferring multi-scale functional robustness in the face of physiological and pathological perturbations, achieved through many-to-many mappings between constitutive components and physiological outcomes.

neuroscience↗

bioRxiv2019

Heterogeneities in intrinsic excitability and frequency-dependent response properties of granule cells across the blades of the rat dentate gyrus

The dentate gyrus (DG), the input gate to the hippocampus proper, is anatomically segregated into three different sectors, namely the suprapyramidal blade, the crest region and the infrapyramidal blade. Although there are well-established differences between these sectors in terms of neuronal morphology, connectivity patterns and activity levels, differences in electrophysiological properties of granule cells within these sectors have remained unexplored. Here, employing somatic whole-cell patch-clamp recordings from the rat DG, we demonstrate that granule cells in these sectors manifest considerable heterogeneities in their intrinsic excitability, temporal summation, action potential characteristics and frequency-dependent response properties. Across sectors, these neurons showed positive temporal summation of their responses to inputs mimicking excitatory postsynaptic currents, and showed little to no sag in their voltage responses to pulse currents. Consistently, the impedance amplitude profile manifested low-pass characteristics and the impedance phase profile lacked positive phase values at all measured frequencies, voltages and for all sectors. Granule cells in all sectors exhibited class I excitability, with broadly linear firing rate profiles, and granule cells in the crest region fired significantly less action potentials compared to those in the infrapyramidal blade. Finally, we found weak pairwise correlations across the 18 different measurements obtained individually from each of the three sectors, providing evidence that these measurements are indeed reporting distinct aspects of neuronal physiology. Together, our analyses show that granule cells act as integrators of afferent information, and emphasize the need to account for the considerable physiological heterogeneities in assessing their roles in information encoding and processing.

neuroscience↗

bioRxiv2019

Flowering behaviour in Arabis alpina ensures the maintenance of a perennating dormant bud bank

Arabis alpina, similar to woody perennials, has a complex architecture with a zone of axillary vegetative branches and a zone of dormant buds that serve as perennating organs. We show that floral development during vernalization is the key for shaping the dormant bud zone by facilitating a synchronized and rapid growth after vernalization and thereby causing an increase in auxin response and transport and endogenous indole-3-acetic acid (IAA) levels in the stem. Floral development during vernalization is associated with the development of axillary buds in subapical nodes. Our transcriptome analysis indicated that these buds are not dormant during vernalization but only attain sustained growth after the return to warm temperatures. Floral and subapical vegetative branches grow after vernalization and inhibit the development of the buds below. Dormancy in these buds is regulated across the A. alpina life cycle by low temperatures and by apical dominance in a BRANCHED 1-dependent mechanism.

plant biology↗

bioRxiv2019

Development and validation of microsatellite markers from de novo transcriptome assembly of eggplant (Solanum melongena L.) and its putative progenitor S. incanum L. cultivars

An elite cultivar of eggplant, Ramnagar Giant (Solanum melongena L.) and W-4 (S. incanum L.) with contrasting horticultural traits were used as parental lines to develop a mapping population of RILs. To accelerate breeding programs and to develop large scale SSR markers to be used in QTL mapping, RNASeq libraries from different tissues of both the parental plants were deep sequenced and assembled into representation of a high quality de novo transcriptome using Illumina-based Next Generation Sequencing technology. 99.99% of high quality bases were obtained from all the tissues and deposited in TSA database at the NCBI link. Total 3, 156 and 3, 196 SNVs were detected in S. melongena and S. incanum, respectively. In S. melongena, 11, 262 SSR while in S. incanum 11, 829 SSR containing regions were identified. Based on functional annotation, 21, 914 unique genes could be identified for S. melongena, 21,706 unique genes for S. incanum and overall, 60 different transcription factors were identified in both the lines. Further, a total of 536 SSR markers were designed and screened for polymorphism of which, 157 markers produced polymorphism between the parental lines. The polymorphic SSRs shall be used for genotyping of RILs to map QTLs for various horticultural traits in eggplant and identification of candidate genes in response to biotic and abiotic stress.

genomics↗

bioRxiv2018

PUMILIO hyperactivity drives premature aging of Norad-deficient mice

Although numerous long noncoding RNAs (lncRNAs) have been identified, our understanding of their roles in mammalian physiology remains limited. Here we investigated the physiologic function of the conserved lncRNA Norad in vivo. Deletion of Norad in mice results in genomic instability and mitochondrial dysfunction, leading to a dramatic multi-system degenerative phenotype resembling premature aging. Loss of tissue homeostasis in Norad-deficient animals is attributable to augmented activity of PUMILIO proteins, which act as post-transcriptional repressors of target mRNAs to which they bind. Norad is the preferred RNA target of PUMILIO2 (PUM2) in mouse tissues and, upon loss of Norad, PUM2 hyperactively represses key genes required for mitosis and mitochondrial function. Remarkably, enforced Pum2 expression fully phenocopies Norad deletion, resulting in rapid-onset aging-associated phenotypes. These findings provide new insights and open new lines of investigation into the roles of noncoding RNAs and RNA binding proteins in normal physiology and aging.

molecular biology↗

bioRxiv2018

A strain of an emerging Indian pathotype of Xanthomonas oryzae pv. oryzae defeats the rice bacterial blight resistance gene xa13 without inducing a clade III SWEET gene and is nearly identical to a recent Thai isolate

The rice bacterial blight pathogen Xanthomonas oryzae pv. oryzae (Xoo) injects transcription activator-like effectors (TALEs) that bind and activate host susceptibility (S) genes important for disease. Clade III SWEET genes are major S genes for bacterial blight. The resistance genes xa5, which reduces TALE activity generally, and xa13, a SWEET11 allele not recognized by the cognate TALE, have been effectively deployed. However, strains that defeat both resistance genes individually were recently reported in India and Thailand. To gain insight into the mechanism(s), we completely sequenced the genome of one such strain from each country and examined the encoded TALEs. Strikingly, the two strains are clones, sharing nearly identical TALE repertoires, including a TALE known to activate SWEET11 strongly enough to be effective even when diminished by xa5. We next investigated SWEET gene induction by the Indian strain. The Indian strain induced no clade III SWEET in plants harbouring xa13, indicating a pathogen adaptation that relieves dependence on these genes for susceptibility. The findings open a door to mechanistic understanding of the role SWEET genes play in susceptibility and illustrate the importance of complete genome sequence-based monitoring of Xoo populations in developing varieties with effective disease resistance.

pathology↗

bioRxiv2017

Pervasive contingency and entrenchment in a billion years of Hsp90 evolution

Although many potential mutations within proteins modulate each others effects1-4, the extent to which these epistatic interactions influenced the fitness effects of the sequence changes that actually occurred during historical evolution - and thus made molecular evolution contingent and irreversible - is controversial5-16. We addressed this question directly by precisely measuring the fitness effects in both extant and reconstructed ancestral sequence contexts of all historical amino acid substitutions that occurred during the billion-year evolutionary history of the heat shock protein 90 (Hsp90) ATPase domain beginning from a deep eukaryotic ancestor to modern Saccharomyces cerevisiae. We find a pervasive influence of epistasis on historical sequence evolution: the majority of the 98 derived states that evolved during history were deleterious at times before they happened, and the vast majority also became subsequently entrenched6, with the ancestral state becoming deleterious after its substitution. A few of these epistatic interactions were of massive fitness consequence, but the majority were of small but evolutionarily relevant effect size. We find that both the large- and small-effect epistasis were largely caused by specific interactions among sites rather than a general permissive or restrictive effect17. Our results highlight how epistasis continually opens and closes windows of mutational opportunity over evolutionary timescales, producing histories and biological states that reflect, in significant part, the transient internal constraints imposed by a proteins fleeting sequence states.

evolutionary biology↗

bioRxiv2017

Heterogeneities in afferent connectivity dominate local heterogeneities in the emergence of response decorrelation in the dentate gyrus

The ability of a neuronal population to effectuate response decorrelation has been identified as an essential prelude to efficient neural encoding. To what extent are diverse forms of local and afferent heterogeneities essential in accomplishing such response decorrelation in the dentate gyrus (DG)? Here, we incrementally incorporated four distinct forms of biological heterogeneities into conductance-based network models of the DG and systematically delineate their relative contributions to response decorrelation. We incorporated intrinsic heterogeneities by stochastically generating several electrophysiologically-validated basket and granule cell models that exhibited significant parametric variability, and introduced synaptic heterogeneities through randomized local synaptic strengths. In including adult neurogenesis, we subjected the valid model populations to randomized structural plasticity and matched neuronal excitability to electrophysiological data. We assessed networks comprising different combinations of these three local heterogeneities with identical or heterogeneous afferent inputs from the entorhinal cortex. We found that the three forms of local heterogeneities were independently and synergistically capable of mediating significant response decorrelation when the network was driven by identical afferent inputs. Strikingly, however, when we incorporated afferent heterogeneities into the network to account for the unique divergence in DG afferent connectivity, the impact of all three forms of local heterogeneities were significantly suppressed by the dominant role of afferent heterogeneities in mediating response decorrelation. Our results unveil a unique convergence of cellular- and network-scale degeneracy in the emergence of response decorrelation in the DG, and constitute a significant departure from the literature that assigns a critical role for local network heterogeneities in input discriminability.\n\nSIGNIFICANCE STATEMENTThe olfactory bulb and the dentate gyrus (DG) networks assimilate new neurons in adult rodents, with adult neurogenesis postulated to subserve efficacious information transfer by reducing correlations in neuronal responses to afferent inputs. Heterogeneities emerging from the lateral dendro-dendritic synapses, mediated by locally-projecting neurogenic inhibitory granule cells, are known to play critical roles in channel decorrelation in the olfactory bulb. However, the contributions of different heterogeneities in mediating response decorrelation in DG, comprising neurogenic excitatory granule cells projecting beyond DG and endowed with uniquely divergent afferent inputs, have not been delineated. Here, we quantitatively demonstrate the dominance of afferent heterogeneities, over multiple local heterogeneities, in the emergence of response decorrelation in DG, together unveiling cross-region degeneracy in accomplishing response decorrelation.

neuroscience↗