Biology subjects

Malik, A.

Publications and source records attributed to Malik, A..

52 records · Page 3Linked to original sources

bioRxiv2021

The impact of mutations on the structural and functional properties of SARS-CoV-2 proteins: A comprehensive bioinformatics analysis

An in-depth analysis of first wave SARS-CoV-2 genome is required to identify various mutations that significantly affect viral fitness. In the present study, we have performed comprehensive in-silico mutational analysis of 3C-like protease (3CLpro), RNA dependent RNA polymerase (RdRp), and spike (S) proteins with the aim of gaining important insights into first wave virus mutations and their functional and structural impact on SARS-CoV-2 proteins. Our integrated analysis gathered 3465 SARS-CoV-2 sequences and identified 92 mutations in S, 37 in RdRp, and 11 in 3CLpro regions. The impact of those mutations was also investigated using various in silico approaches. Among these 32 mutations in S, 15 in RdRp, and 3 in 3CLpro proteins are found to be deleterious in nature and could alter the structural and functional behavior of the encoded proteins. D614G mutation in spike and P323L in RdRp are the globally dominant variants with a high frequency. Most of them have also been found in the binding moiety of the viral proteins which determine their critical involvement in the host-pathogen interactions and drug targets. The findings of the current study may facilitate better understanding of COVID-19 diagnostics, vaccines, and therapeutics.

bioinformatics↗

bioRxiv2021

Physiological and transcriptomic variability indicative of differences in key functions within a single coral colony

Polyps in different locations on individual stony coral colonies experience variation in numerous environmental conditions including flow and light, potentially leading to transcriptional and physiological differences across the colony. Here, we describe high-resolution physiological measurements and differential gene expression from multiple locations within a single colony of Stylophora pistillata, aiming to relate these to environmental gradients across the coral colony. We observed broad transcriptional responses in both the host and photosymbiont in response to height above the substrate, cardinal direction, and, most strongly, location along the branch axis. Specifically, several key physiological processes in the host appear more active toward branch tips including several metabolic pathways, toxin production for prey capture or defense, and biomolecular mechanisms of biomineralization. Further, the increase in gene expression related to these processes toward branch tips is conserved between S. pistillata and Acropora spp. The photosymbiont appears to respond transcriptionally to relative light intensity along the branch and due to cardinal direction. These differential responses were observed across the colony despite its genetic homogeneity and likely inter-polyp communication. While not a classical division of labor, each part of the colony appears to have distinct functional roles related to polyps differential exposure to environmental conditions.

genomics↗

bioRxiv2020

Finding Pathogenic nsSNP's and their structural effect on COPS2 using Molecular Dynamic Approach

COP9 Signalosome Subunit 2 is a highly conserved multiprotein complex which is involved in the cellular process and developmental process. It is one of the essential components in the COP9 Signalosome Complex (CSN). It is also involved in neuronal differentiation interacting with NIF3L1. The gene involved in neuronal differentiation is negatively regulated due to the transcription co-repressor interaction of NIF3L1 with COPS2. In the present study, we have evaluated the outcome for 90 non-synonymous single nucleotide polymorphisms (nsSNPs) in COPS2 gene through computational tools. After the analysis, 4 SNPs (S120C, N144S, Y159H, R173C) were found to be deleterious. The native and mutated structures were prepared using discovery studio and docked to check the interactions with NIF3L1.On the basis of ZDOCK score the top 3 mutations (N144S, Y159H, R173C) were screened out. Further to analyze the effect of amino acid substitution on the molecular structure of protein Molecular Dynamics simulation was carried out. Analysis based on RMSD, RMSF, RG, H-bond showed a significant deviation in the graph, which demonstrated conformation change and instability compared to the wild structure. As it is known mutations in COPS2 gene can disrupt the normal activity of the CSN2 protein which may cause neuronal differentiation. Our results showed N144S, Y159H and R173C mutations are to be more pathogenic and may cause disease

biophysics↗

bioRxiv2020

Ligand Modulation of the Conformational Dynamics of the A2A Adenosine Receptor Revealed by Single-Molecule Fluorescence

G protein-coupled receptors (GPCRs) are the largest class of transmembrane proteins, making them an important target for therapeutics. Activation of these receptors is modulated by orthosteric ligands, which stabilize one or several states within a complex conformational ensemble. The intra-and inter-state dynamics, however, is not well documented. Here, we used single-molecule fluorescence to measure ligand-modulated conformational dynamics of the adenosine A2A Receptor (A2AR) on nanosecond to millisecond timescales. Experiments were performed on detergent-purified A2R in either the ligand-free (apo) state, or when bound to an inverse, partial or full agonist ligand. Single-molecule Forster resonance energy transfer (smFRET) was performed on detergent-solubilized A2AR to resolve active and inactive states via the separation between transmembrane (TM) helices 4 and 6. The ligand-dependent changes of the smFRET distributions are consistent with conformational selection and with inter-state exchange lifetimes [≥] 3 ms. Local conformational dynamics around residue 229 on TM6 was measured using Fluorescence Correlation Spectroscopy (FCS), which captures dynamic quenching due to photoinduced electron transfer (PET) between a covalently-attached dye and proximal aromatic residues. Global analysis of PET-FCS data revealed fast (150-350 ns), intermediate (50-60 s) and slow (200-300 s) conformational dynamics in A2AR, with lifetimes and amplitudes modulated by ligands and a G-protein mimetic (mini-Gs). Most notably, the agonist binding and the coupling to mini-Gs accelerates and increases the relative contribution of the sub-microsecond phase. Molecular dynamics simulations identified three tyrosine residues (Y112, Y288, and Y290) as being responsible for the dynamic quenching observed by PET-FCS and revealed associated helical motions around residue 229 on TM6. This study provides a quantitative description of conformational dynamics in A2AR and supports the idea that ligands bias not only GPCR conformations but also the dynamics within and between distinct conformational states of the receptor.

biophysics↗

bioRxiv2020

An autocrine Vitamin D-driven Th1 shutdown program can be exploited for COVID-19

Pro-inflammatory immune responses are necessary for effective pathogen clearance, but cause severe tissue damage if not shut down in a timely manner1,2. Excessive complement and IFN-{gamma}-associated responses are known drivers of immunopathogenesis3 and are among the most highly induced immune programs in hyper-inflammatory SARS-CoV2 lung infection4. The molecular mechanisms that govern orderly shutdown and retraction of these responses remain poorly understood. Here, we show that complement triggers contraction of IFN-{gamma} producing CD4+ T helper (Th) 1 cell responses by inducing expression of the vitamin D (VitD) receptor (VDR) and CYP27B1, the enzyme that activates VitD, permitting T cells to both activate and respond to VitD. VitD then initiates the transition from pro-inflammatory IFN-{gamma}+ Th1 cells to suppressive IL-10+ Th1 cells. This process is primed by dynamic changes in the epigenetic landscape of CD4+ T cells, generating superenhancers and recruiting c-JUN and BACH2, a key immunoregulatory transcription factor5-7. Accordingly, cells in psoriatic skin treated with VitD increased BACH2 expression, and BACH2 haplo-insufficient CD4+ T cells were defective in IL-10 production. As proof-of-concept, we show that CD4+ T cells in the bronchoalveolar lavage fluid (BALF) of patients with COVID-19 are Th1-skewed and that VDR is among the top regulators of genes induced by SARS-CoV2. Importantly, genes normally down-regulated by VitD were de-repressed in CD4+ BALF T cells of COVID-19, indicating that the VitD-driven shutdown program is impaired in this setting. The active metabolite of VitD, alfacalcidol, and cortico-steroids were among the top predicted pharmaceuticals that could normalize SARS-CoV2 induced genes. These data indicate that adjunct therapy with VitD in the context of other immunomodulatory drugs may be a beneficial strategy to dampen hyperinflammation in severe COVID-19.

immunology↗

bioRxiv2020

Interfacial Water Molecules Make RBD of SPIKE Protein and Human ACE2 to Stick Together

A novel coronavirus (SARS-CoV-2; COVID-19) that initially originates from Wuhan province in China has emerged as a global pandemic, an outbreak that started at the end of 2019 which claims 431,192 (Date: 15th June 2020 (https://covid19.who.in) life till now. Since then scientists all over the world are engaged in developing new vaccines, antibodies, or drug molecules to combat this new threat. Here in this work, we performed an in-silico analysis on the protein-protein interactions between the receptor-binding (RBD) domain of viral SPIKE protein and human angiotensin-converting enzyme 2 (hACE2) receptor to highlight the key alteration that happened from SARS-CoV to SARS-CoV-2. We analyzed and compared the molecular differences between these two viruses by using various computational approaches such as binding affinity calculations, computational alanine, and molecular dynamics simulations. The binding affinity calculations show SARS-CoV-2 binds little more firmly to the hACE2 receptor than that of SARS-CoV. Analysis of simulation trajectories reveals that enhanced hydrophobic contacts or the van der Waals interaction play a major role in stabilizing the protein-protein interface. The major finding obtained from molecular dynamics simulations is that the RBD-ACE2 interface is populated with water molecules and interacts strongly with both RBD and ACE2 interfacial residues during the simulation periods. We also emphasize that the interfacial water molecules play a critical role in binding and maintaining the stability of the RBD/hACE2 complex. The water-mediated hydrogen bond by the bridge water molecules is crucial for stabilizing the RBD and ACE2 domains. The structural and dynamical features presented here may serve as a guide for developing new drug molecules, vaccines, or antibodies to combat the COVID-19 pandemic.

biophysics↗

bioRxiv2020

Optimization of skeletal protein preparation for LC-MS/MS sequencing yields additional coral skeletal proteins in Stylophora pistillata

Stony corals generate their calcium carbonate exoskeleton in a highly controlled biomineralization process mediated by a variety of macromolecules including proteins. Fully identifying and classifying these proteins is crucial to understanding their role in exoskeleton formation, yet no optimal method to purify and characterize the full suite of extracted coral skeletal proteins has been established and hence their complete composition remains obscure. Here, we tested four skeletal protein purification protocols using acetone precipitation and ultrafiltration dialysis filters to present a comprehensive scleractinian coral skeletal proteome. We identified a total of 60 proteins in the coral skeleton, 44 of which were not present in previously published stony coral skeletal proteomes. Extracted protein purification protocols carried out in this study revealed that no one method captures all proteins and each protocol revealed a unique set of method-exclusive proteins. To better understand the general mechanism of skeletal protein transportation, we further examined the proteins gene ontology, transmembrane domains, and signal peptides. We found that transmembrane domain proteins and signal peptide secretion pathways, by themselves, could not explain the transportation of proteins to the skeleton. We therefore propose that some proteins are transported to the skeleton via non-traditional secretion pathways.

biochemistry↗

bioRxiv2020

A tentacle for every occasion: comparing the hunting tentacles and sweeper tentacles, used for territorial competition, in the coral Galaxea fascicularis

Coral reefs are among the most diverse, complex and densely populated marine ecosystems. To survive, morphologically simple and sessile cnidarians have developed mechanisms to catch prey, deter predators and compete with adjacent corals for space, yet the mechanisms underlying these functions are largely unknown. Here, we characterize the histology, toxic activity and gene expression patterns in two different types of tentacles from the scleractinian coral Galaxea fascilcularis - catch tentacles (CTs), used to catch prey and deter predators, and sweeper tentacles (STs), specialized tentacles used for territorial aggression. STs exhibit more mucocytes and higher expression of mucin genes than CTs, and lack the ectodermal cilia used to deliver food to the mouth and remove debris. STs and CTs also express different sensory g-protein coupled receptors, suggesting they may employ different sensory pathways. Each tentacle type has a different complement of stinging cells (nematocytes), and the expression in the two tentacles of genes encoding structural nematocyte proteins suggests the stinging cells develop within the tentacles. CTs have higher neurotoxic and hemolytic activities, consistent with a role in prey capture, whereas the STs have higher phospholipase A2 activity, which we speculate may have a role in inducing tissue damage during territorial aggression. The toxin genes expressed in each tentacle are also different. These results show that the same organism utilizes two distinct tentacle types, each equipped with a different venom apparatus and toxin composition, for prey capture and defense and for territorial aggression.

ecology↗

bioRxiv2020

Cell polarisation in a bulk-surface model can be driven by both classic and non-classic Turing instability

The GTPase Cdc42 is the master regulator of eukaryotic cell polarisation. During this process the active form of Cdc42 is accumulated at a particular site on the cell membrane called the pole. It is believed that the accumulation of the active Cdc42 resulting in a pole is driven by a combination of activation-inactivation reactions and diffusion. It has been proposed using mathematical modelling that this is the result of diffusion-driven instability, originally proposed by Alan Turing. In this study we developed, analysed and validated a 3D bulk-surface model of the dynamics of Cdc42. We show that the model can undergo both classic and non-classic Turing instability by deriving necessary conditions for which this occurs and conclude that the non-classic case can be viewed as a limit case of the classic case of diffusion driven instability. We thoroughly investigate the parameter space. Using three-dimensional spatio-temporal simulation we predicted pole size and time to polarisation, suggesting that cell polarisation is mainly driven by the reaction strength parameter and that the size of the pole is determined by the relative diffusion.

systems biology↗

bioRxiv2019

The role of ANGIOGENIN, or lack thereof, in the generation of stress-induced tRNA halves and of smaller tRNA fragments that enter Argonaute complexes

Overexpressed Angiogenin (ANG) cleaves tRNA anticodons to produce tRNA halves similar to those produced in response to stress, but it is not clear whether endogenous ANG is essential for producing the stress-induced tRNA halves. It is also not clear whether smaller tRNA fragments (tRFs) are generated from the tRNA halves. Global short RNAseq experiments reveal that ANG over-expression selectively cleaves a subset of tRNAs (tRNAGlu, tRNAGly, tRNALys, tRNAVal, tRNAHis, tRNAAsp and tRNASeC) to produce tRNA halves and 26-30 bases long tRF-5s. Surprisingly, knockout of ANG reveals that the majority of stress-induced tRNA halves except 5 half from tRNAHisGTG and 3 half from tRNAAspGTC are ANG-independent, suggesting there are other RNases that can produce tRNA halves. The 17-25 bases long tRF-3s and tRF-5s that could enter into Argonaute complexes are not induced by ANG overexpression, suggesting that they are generated independently from tRNA halves. Consistent with this, knockout of ANG did not decrease tRF-3 levels or gene-silencing activity. Therefore ANG cleaves specific tRNAs, is not the only RNAse that creates tRNA halves and the shorter tRFs are not generated from the tRNA halves or from independent tRNA cleavage by ANG.

molecular biology↗

bioRxiv2019

Possible co-option of a VEGF-driven tubulogenesis program for biomineralization in echinoderms

Biomineralization is the process in which living organisms use minerals to form hard structures that protect and support them. Biomineralization is believed to have evolved rapidly and independently in different phyla utilizing existing components used for other purposes. The mechanistic understanding of the regulatory networks that drive biomineralization and their evolution is far from clear. The sea urchin skeletogenesis is an excellent model system for studying both gene regulation and mineral uptake and deposition. The sea urchin calcite spicules are formed within a tubular cavity generated by the skeletogenic cells under the control the vascular endothelial growth factor (VEGF) signaling. The VEGF pathway controls tubulogenesis and vascularization across metazoans while its regulation of biomineralization was only observed in echinoderms. Despite the critical role of VEGF signaling in sea urchin spiculogenesis, the downstream program it activates was largely unknown. Here we study the cellular and molecular machinery activated by the VEGF pathway during sea urchin spiculogenesis and reveal multiple parallels to the regulation of tubulogenesis during vertebrate vascularization. Human VEGF rescues sea urchin VEGF knock-down; VEGF-dependent vesicle deposition plays a significant role in both systems and sea urchin VEGF signaling activates hundreds of genes including biomineralization and vascularization genes. Five upstream transcription factors and three signaling genes active in spiculogenesis are homologous to vertebrate factors that regulate vascularization. Overall, our findings suggest that sea urchin spiculogenesis and vertebrate vascularization diverged from a common ancestral tubulogenesis program, broadly adapted for vascularization and specifically co-opted for biomineralization in the echinoderm phylum. Significance statementThe sea urchin calcite spicules and vertebrate blood vessels are quite distinct in their function, yet both have a tubular structure and are controlled by the vascular endothelial growth factor (VEGF) pathway. Here we study the downstream program by which VEGF pathway drives sea urchin spiculogenesis and find remarkable similarities to the control of vertebrate vascularization. The similarities are observed both in the upstream gene regulatory network, in the downstream effector genes and the cellular processes that VEGF signaling controls at the site of the calcite spicule formation. We speculate that sea urchin spiculogenesis and vertebrate vascularization diverged from a common ancestral tubulogenesis program that was co-opted for biomineralization in the echinoderm phylum.

evolutionary biology↗

bioRxiv2019

An EEG study of detection without localisation in change blindness

Previous studies of change blindness have suggested a distinction between detection and localisation of changes in a visual scene. Using a simple paradigm with an array of coloured squares, the present study aimed to further investigate differences in event-related potentials (ERPs) between trials in which participants could detect the presence of a colour change but not identify the location of the change (sense trials), versus those where participants could both detect and localise the change (localise trials). Individual differences in performance were controlled for by adjusting the difficulty of the task in real time. Behaviourally, reaction times for sense, blind, and false alarm trials were distinguishable when comparing across levels of participant certainty. In the EEG data, we found no significant differences in the visual awareness negativity ERP, contrary to previous findings. In the N2pc range, both awareness conditions (localise and sense) were significantly different to trials with no change detection (blind trials), suggesting that this ERP is not dependent on explicit awareness. Within the late parietal positivity range, all conditions were significantly different. These results suggest that changes can be sensed without knowledge of the location of the changing object, and that participant certainty scores can provide valuable information about the perception of changes in change blindness.

neuroscience↗

bioRxiv2018

A novel mutation alters the stability of PapA2 resulting in the complete abrogation of sulfolipids in clinical mycobacterial strains

The analysis of whole genome has revealed specific geographical distribution of Mycobacterium tuberculosis (Mtb) strains across the globe suggestive of unique host dependent adaptive mechanisms in Mtb evolved out of selective pressures. We provide an important correlation of a genome-based mutation to a molecular phenotype across two predominant clinical Mtb lineages of the Indian subcontinent. We have identified a distinct lineage specific mutation-G247C, translating into an alanine to proline conversion in the papA2 gene of Indo oceanic lineage 1 [L1] Mtb strains; restoration of cell wall sulfolipids by simple genetic complementation of papA2 from lineage 3 [L3] or from H37Rv (Lineage 4-L4) attributed the loss of this glycolipid to this specific mutation in Indo oceanic L1 Mtb. Investigation into the structure of Mtb PapA2 revealed a distinct non-ribosomal peptide synthetase (NRPS) C domain conformation with an unconventional presence of Zinc binding motif. Surprisingly, the A83P mutation did not map to either the catalytic centre in the N-terminal subdomain or any of the substrate binding region of the protein. On the contrary, the inherent ability of mutant PapA2 to form insoluble aggregates and molecular simulations with the Wt/ mut PapA2 purports an important role for the surface associated 83rd residue in protein conformation. The present study demonstrates the importance of a critical structural residue in the papA2 protein of Mtb and helps establish a link between observed genomic alteration and its molecular consequence in the successful human pathogen Mtb.

molecular biology↗

bioRxiv2018

The role of land use types and water chemical properties in structuring the microbiome of a connected lakes system

Lakes and other freshwater bodies are intimately connected to the surrounding land, yet to what extent land-use affects the quality of freshwater and the microbial communities living in various freshwater environments is largely unknown. We address this question through an analysis of the land use surrounding 46 inter-connected lakes located within 7 different drainage basins in northern Germany, and the microbiomes of these lakes during early summer. Lake microbiome structure was not determined by the specific drainage basin or by basin size, and bacterial distribution did not seem to be limited by distance. Instead, land use within the drainage basin could predict, to some extent, NO2+NO3 concentrations in the water, which (together with temperature, chlorophyll a and total phosphorus) affected water microbiome structure. Land use directly surrounding the water bodies, however, had little observable effects on water quality or the microbiome. Several microbial lineages, including environmentally important Cyanobacteria and Verrucomicrobia, were differentially partitioned between the lakes. As the amount of available data on land use (e.g. from remote sensing) increases, identifying relationships between land use, aquatic microbial communities and their effect on water quality will be important to better manage freshwater resources worldwide, e.g. by systemically identifying water bodies prone to ecological changes or the presence of harmful organisms.

microbiology↗

bioRxiv2018

Matrin 3-dependent neurotoxicity is modified by nucleic acid binding and nucleocytoplasmic localization

Abnormalities in nucleic acid processing are associated with the development of amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD). Mutations in Matrin 3 (MATR3), a poorly understood DNA- and RNA-binding protein, cause familial ALS/FTD, and MATR3 pathology is a feature of sporadic disease, suggesting that MATR3 dysfunction is integrally linked to ALS pathogenesis. Using a primary neuron model to assess MATR3-mediated toxicity, we noted that neurons were bidirectionally vulnerable to MATR3 levels, with pathogenic MATR3 mutants displaying enhanced toxicity. MATR3s zinc finger domains partially modulated toxicity, but elimination of its RNA recognition motifs had no effect on neuronal survival, instead facilitating its self-assembly into liquid-like droplets. In contrast to other RNA-binding proteins associated with ALS, cytoplasmic MATR3 redistribution mitigated neurodegeneration, suggesting that nuclear MATR3 mediates toxicity. Our findings offer a foundation for understanding MATR3-related neurodegeneration and how nucleic acid binding functions, localization, and pathogenic mutations drive sporadic and familial disease.

cell biology↗

bioRxiv2017

Global Gene Repression By Dicer-Independent tRNA Fragments

tRNA derived RNA fragments (tRFs) is an emerging group of small RNAs as abundant as miRNAs, and yet their roles are not well understood. Here, we focus on endogenous tRFs (18-22 bases) derived from 3 end of human mature tRNAs (tRF-3) and their functions in gene repression. tRF-3 levels increase upon parental tRNA over-expression or tRNA induction by c-Myc oncogene activation. Elevated tRF-3 levels lead to repression of target genes with a sequence complementary to the tRF-3 in the 3 UTR. The tRF-3-mediated repression is Dicer-independent, Argonaute-dependent and the targets are recognized by 5 seed sequence rules similar to miRNAs. Furthermore, tRF-3s associate with GW proteins in P-bodies. RNA-seq identifies the endogenous target genes of tRF-3s that are specifically repressed upon tRF-3 induction. Overall, our analysis shows Dicer-independent tRF-3s, generated upon tRNA upregulation such as c-Myc overexpression, regulate gene expression globally through Argounate via seed sequence matches.

molecular biology↗