Biology subjects

Easwaran, S.

Publications and source records attributed to Easwaran, S..

4 recordsLinked to original sources

bioRxiv2024

A genome-wide association study implicates the olfactory system in Drosophila melanogaster diapause-associated lifespan extension and fecundity

The effects of environmental stress on animal life are gaining importance. Diapause is a reversible dormancy program triggered by adverse environmental conditions. To characterize the genetic basis of this complex program, we leveraged the Drosophila Genome Reference Panel (DGRP) to conduct a Genome-Wide Association Study (GWAS). We assessed post-diapause and non-diapause fecundity across 193 DGRP lines. GWAS revealed 546 variants, encompassing single nucleotide polymorphisms, insertions, and deletions associated with post-diapause fecundity. We identified 291 candidate diapause-associated genes, 40 of which were previously associated with diapause. Gene network analysis indicated that the diapause-associated genes were primarily linked to neuronal and reproductive system development. Similarly, comparison with other fly GWAS revealed the greatest overlap with olfactory-behavior-associated and fecundity-and-lifespan-associated genes. An RNAi screen of selected candidates identified two neuronal genes, Dip-{gamma} and Scribbler, to be required during recovery for post-diapause fecundity. To complement the genetic analysis, we tested which neurons are required for successful diapause. Although amputation of the antenna had little effect on non-diapause lifespan, it reduced diapause lifespan and post-diapause fecundity. Furthermore, olfactory receptor neurons and temperature-sensing neurons were required for successful diapause recovery. Our results provide insights into the molecular, cellular, and genetic basis of Drosophila adult reproductive diapause.

genetics↗

bioRxiv2024

Altered circadian rhythm, sleep, and rhodopsin 7-dependent shade preference during diapause in Drosophila melanogaster

To survive adverse environments, many animals enter a dormant state such as hibernation, dauer, or diapause. Various Drosophila species undergo adult reproductive diapause in response to cool temperatures and/or short day-length. While it is known that flies are less active during diapause, an in-depth understanding of diapause effects on circadian rhythms and sleep is lacking. Here we show that, in diapause-inducing conditions, Drosophila melanogaster exhibit altered circadian activity profiles, including a severely reduced morning activity peak and an advanced evening activity peak. Consequently, the flies have a single activity peak at a time similar to when non-diapausing flies have a siesta. Temperatures [≤]15 {degrees}C, rather than short day-length, primarily drive the behavior. At cool temperatures, flies also rapidly enter a deep sleep state that lacks the sleep cycles of flies at higher temperatures and requires particularly high levels of stimulation for arousal. Furthermore, we show that at 18-25 {degrees}C, flies prefer to siesta in the shade, a preference that is virtually eliminated at 10 {degrees}C. Resting in the shade is driven by an aversion to blue light, sensed by rhodopsin 7 (Rh7) outside of the eyes. Flies at 10 {degrees}C show neuronal markers of elevated sleep pressure, including increased expression of Bruchpilot and elevated Ca2+ in the R5 ellipsoid body neurons. Therefore sleep pressure might overcome blue light aversion. Thus at temperatures known to cause reproductive arrest, preserve germline stem cells, and extend lifespan, Drosophila melanogaster are prone to deep sleep and exhibit dramatically altered - yet rhythmic - daily activity patterns. Significance statementClimate change is impacting many animals, including insects. In diverse organisms, adverse environments trigger dormancy programs such as hibernation and diapause. Fruit flies undergo diapause to survive winter. Here we develop new methods and show that the same cool temperatures that delay fruit fly reproduction and extend lifespan, also promote deep sleep. Cool flies rapidly fall asleep and are difficult to arouse. Once awake, they immediately fall back to sleep. Whereas in warm environments, midday blue light drives flies to siesta in the shade, in cool temperatures the need to sleep overwhelms light-aversion, reducing shade preference. Animals that adjust their behavior directly to temperature, rather than day length, may be more resilient to a changing climate.

neuroscience↗

bioRxiv2021

A Rapid Bead-Based Assay For Screening Of SARS-CoV-2 Neutralising Antibodies

Quantitative determination of neutralizing antibodies against Severe Acute Respiratory Syndrome Corona Virus-2 (SARS-CoV-2) is paramount in immunodiagnostics, vaccine efficacy testing, and immune response profiling among the vaccinated population. Cost-effective, rapid, easy-to-perform assays are essential to support the vaccine development process and immunosurveillance studies. Here, we describe a bead-based screening assay for S1-neutralization using recombinant fluorescent proteins of hACE2 and SARS-CoV2-S1, immobilized on solid beads employing nanobodies /metal-affinity tags. Nanobody-mediated capture of SARS-CoV-2 - Spike (S1) on agarose beads served as the trap for soluble recombinant ACE2-GFPSpark, inhibited by neutralizing antibody. The first approach demonstrates single-color fluorescent imaging of ACE2-GFPspark binding to His-tagged S1-Receptor Binding Domain (RBD-His) immobilized beads. The second approach is dual-color imaging of soluble ACE2-GFPSpark to S1-Orange Fluorescent Protein (S1-OFPSpark) beads. Both methods showed a good correlation with the gold standard pseudovirion assay and can be adapted to any fluorescent platforms for screening. Life-time imaging of the ACE2-GFPSpark confirmed the interaction of ACE2 and S1-OFPSpark on beads. The self-renewable source of secreted recombinant proteins from stable cells and its direct use without necessitating purification renders the platform a cost-effective and rapid one than the popular pseudovirion assay and live virus-based assays. Any laboratory with minimum expertise can rapidly perform this bead assay for neutralizing antibody detection using stable engineered cells.

molecular biology↗

bioRxiv2020

Structural and Functional Implications of Non-synonymous Mutations in the Spike protein of 2,954 SARS-CoV-2 Genomes

Protein-protein interactions between virus and host are crucial for infection. SARS-CoV-2, the causative agent of COVID-19 pandemic is an RNA virus prone to mutations. Formation of a stable binding interface between the Spike (S) protein Receptor Binding Domain (RBD) of SARS-CoV-2 and Angiotensin-Converting Enzyme 2 (ACE2) of host actuates viral entry. Yet, how this binding interface evolves as virus acquires mutations during pandemic remains elusive. Here, using a high fidelity bioinformatics pipeline, we analysed 31,403 SARS-CoV-2 genomes across the globe, and identified 444 non-synonymous mutations that cause 49 distinct amino acid substitutions in the RBD. Molecular phylogenetic analysis suggested independent emergence of these RBD mutants during pandemic. In silico structure modelling of interfaces induced by mutations on residues which directly engage ACE2 or lie in the near vicinity revealed molecular rearrangements and binding energies unique to each RBD mutant. Comparative structure analysis using binding interface from mouse that prevents SARS-CoV-2 entry uncovered minimal molecular determinants in RBD necessary for the formation of stable interface. We identified that interfacial interaction involving amino acid residues N487 and G496 on either ends of the binding scaffold are indispensable to anchor RBD and are well conserved in all SARS-like corona viruses. All other interactions appear to be required to locally remodel binding interface with varying affinities and thus may decide extent of viral transmission and disease outcome. Together, our findings propose the modalities and variations in RBD-ACE2 interface formation exploited by SARS-CoV-2 for endurance. ImportanceCOVID-19, so far the worst hit pandemic to mankind, started in January 2020 and is still prevailing globally. Our study identified key molecular arrangements in RBD-ACE2 interface that help virus to tolerate mutations and prevail. In addition, RBD mutations identified in this study can serve as a molecular directory for experimental biologists to perform functional validation experiments. The minimal molecular requirements for the formation of RBD-ACE2 interface predicted using in silico structure models may help precisely design neutralizing antibodies, vaccines and therapeutics. Our study also proposes the significance of understanding evolution of protein interfaces during pandemic.

evolutionary biology↗