Biology subjects

Arora, K.

Publications and source records attributed to Arora, K..

35 records · Page 2Linked to original sources

bioRxiv2021

SARS-CoV-2 variants of concern infect the respiratory tract and induce inflammatory response in wild-type laboratory mice

The emergence of new severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) variants of concern poses a major threat to the public health due to possible enhanced virulence, transmissibility and immune escape. These variants may also adapt to new hosts in part through mutations in the spike protein. In this study, we evaluated the infectivity and pathogenicity of SARS-CoV-2 variants of concern in wild-type C57BL/6 mice. Six-week-old mice were inoculated intranasally with a representative virus from the original B.1 lineage or emerging B.1.1.7 and B.1.351 lineages. We also infected a group of mice with a mouse-adapted SARS-CoV-2 (MA10). Viral load and mRNA levels of multiple cytokines and chemokines were analyzed in the lung tissues on day 3 after infection. Our data show that unlike the B.1 virus, the B.1.1.7 and B.1.351 viruses are capable of infecting C57BL/6 mice and replicating at high concentrations in the lungs. The B.1.351 virus replicated to higher titers in the lungs compared to the B.1.1.7 and MA10 viruses. The levels of cytokines (IL-6, TNF-, IL-1{beta}) and chemokine (CCL2) were upregulated in response to the B.1.1.7 and B.1.351 infection in the lungs. Overall, these data indicate a greater potential for infectivity and adaptation to new hosts by emerging SARS-CoV-2 variants.

microbiology↗

bioRxiv2021

N-terminal extensions strengthen hydrophobic inter-subunit interactions between HU's C-terminal domains to frustrate heterodimer formation

HU is a nucleoid-associated protein (NAP) that helps bacterial chromosomal DNA to remain compact. Escherichia coli contains two homologs of HU that are ~ 70 % identical: HU-A and HU-B. The early log phase, late log phase, and stationary phase of E. coli growth are reported to be dominated, respectively, by HU-AA homodimers, HU-BB homo-dimers, and HU-AB heterodimers. Here, we show that the formation of HU-AB heterodimers occurs to a much lower degree in HU chains that have a displaced N-terminus, whether through addition of an N-terminal affinity (polyhistidine) tag, or fusion of a fluorescent protein. A combination of mass spectrometry, spectroscopy, chromatography, and electrophoresis (exploring glutaraldehyde crosslinking of subunits) was used to study the mixing, co-expression, unfolding and refolding of HU-AA and HU-BB homodimers. The data suggests that, in HU polypeptides with N-terminal extension, whereas inter-subunit contacts between the alpha helical N-terminal domains (NTDs) undergo facile unfolding and dissociation, inter-subunit contacts between the beta sheet- and IDR-dominated C-terminal domains (CTDs) fail to do so, due to persistence of hydrophobic inter-subunit interactions between two beta sheets. This persistence causes HU to remain nominally dimeric even after substantive unfolding, and frustrates subunit exchange and heterodimer formation.

biochemistry↗

bioRxiv2020

Neuroinvasion and encephalitis following intranasal inoculation of SARS-CoV-2 in K18-hACE2 mice

Severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) infection can cause neurological disease in humans, but little is known about the pathogenesis of SARS-CoV-2 infection in the central nervous system. Herein, using K18-hACE2 mice, we demonstrate that SARS-CoV-2 neuroinvasion and encephalitis is associated with mortality in these mice. Intranasal infection of K18-hACE2 mice with 105 plaque-forming units of SARS-CoV-2 resulted in 100% mortality by day 6 after infection. The highest virus titers in the lungs were observed at day 3 and declined at days 5 and 6 after infection. In contrast, very high levels of infectious virus were uniformly detected in the brains of all the animals at days 5 and 6. Onset of severe disease in infected mice correlated with peak viral levels in the brain. SARS-CoV-2-infected mice exhibited encephalitis hallmarks characterized by production of cytokines and chemokines, leukocyte infiltration, hemorrhage and neuronal cell death. SARS-CoV-2 was also found to productively infect cells within the nasal turbinate, eye and olfactory bulb, suggesting SARS-CoV-2 entry into the brain by this route after intranasal infection. Our data indicate that direct infection of CNS cells together with the induced inflammatory response in the brain resulted in the severe disease observed in SARS-CoV-2-infected K18-hACE2 mice.

microbiology↗

bioRxiv2020

PRAK-03202: A triple antigen VLP vaccine candidate against SARS CoV-2

The rapid development of safe and effective vaccines against SARS CoV-2 is the need of the hour for the coronavirus outbreak. Here, we have developed PRAK-03202, the worlds first triple antigen VLP vaccine candidate in a highly characterized S. cerevisiae-based D-CryptTM platform, which induced SARS CoV-2 specific neutralizing antibodies in BALB/c mice. Immunizations using three different doses of PRAK-03202 induces antigen specific (Spike, envelope and membrane proteins) humoral response and neutralizing potential. PBMCs from convalescent patients, when exposed to PRAK-03202, showed lymphocyte proliferation and elevated IFN-{gamma} levels suggestive of conservation of epitopes and induction of T helper 1 (Th1)-biased cellular immune responses. These data support the clinical development and testing of PRAK-03202 for use in humans.

immunology↗

bioRxiv2020

Mechanism of bacterial adhesion and embedment in a DNA biofilm matrix:Evidence that binding of outer membrane lipopolysaccharide (LPS) to HU is key

In biofilms, bacteria are embedded within a matrix of extracellular DNA (e-DNA). Since bacterial cells and e-DNA are both negatively-charged, a positively-charged substance must act like a glue to allow bacteria to be embedded within the DNA matrix. Here we show that HU (a highly-abundant, histone-like, nucleoid-associated, DNA-binding protein) facilitates bacterium-bacterium and bacterium-DNA interactions by binding to lipopolysaccharide (LPS), a bacterial outer membrane component. We demonstrate that LPS binds to both the canonical and non-canonical DNA-binding sites on HU. We propose that the hexose sugar-terminal phosphate moieties present in the lipid A head-group of LPS bind to the same lysine/arginine residues that are involved in binding of the pentose sugar-phosphate groups in DNA. Alternate binding of LPS and DNA by HUs DNA-binding sites could allow HU to bind to bacterial cells surfaces and thus elicit bacterium-bacterium and bacterium-DNA interactions in biofilms.

microbiology↗

bioRxiv2020

Somatic variant analysis of linked-reads sequencing data with Lancet

Summary We present a new version of the popular somatic variant caller, Lancet, that supports the analysis of linked-reads sequencing data. By seamlessly integrating barcodes and haplotype read assignments within the colored De Bruijn graph local-assembly framework, Lancet computes a barcode-aware coverage and identifies variants that disagree with the local haplotype structure.Availability and Implementation Lancet is implemented in C++ and is available for academic and non-commercial research purposes as an open-source package at https://github.com/nygenome/lancet.Contact gnarzisi{at}nygenome.orgCompeting Interest StatementThe authors have declared no competing interest.View Full Text

bioinformatics↗

bioRxiv2020

LAYNII: A software suite for layer-fMRI

High-resolution fMRI in the sub-millimeter regime allows researchers to resolve brain activity across cortical layers and columns non-invasively. While these high-resolution data make it possible to address novel questions of directional information flow within and across brain circuits, the corresponding data analyses are challenged by MRI artifacts, including image blurring, image distortions, low SNR, and restricted coverage. These challenges often result in insufficient spatial accuracy of conventional analysis pipelines. Here we introduce a new software suite that is specifically designed for layer-specific functional MRI: LayNii. This toolbox is a collection of command-line executable programs written in C/C++ and is distributed open-source and as pre-compiled binaries for Linux, Windows, and macOS. LayNii is designed for layer-fMRI data that suffer from SNR and coverage constraints and thus cannot be straightforwardly analyzed in alternative software packages. Some of the most popular programs of LayNii contain layerification and columnarization in the native voxel space of functional data as well as many other layer-fMRI specific analysis tasks: layer-specific smoothing, model-based vein mitigation of GE-BOLD data, quality assessment of artifact dominated sub-millimeter fMRI, as well as analyses of VASO data. HighlightsO_LIA new software toolbox is introduced for layer-specific functional MRI: LayNii. C_LIO_LILayNii is a suite of command-line executable C++ programs for Linux, Windows, and macOS. C_LIO_LILayNii is designed for layer-fMRI data that suffer from SNR and coverage constraints. C_LIO_LILayNii performs layerification in the native voxel space of functional data. C_LIO_LILayNii performs layer-smoothing, GE-BOLD deveining, QA, and VASO analysis. C_LI O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=78 SRC="FIGDIR/small/148080v3_ufig1.gif" ALT="Figure 1"> View larger version (19K): org.highwire.dtl.DTLVardef@e12e9org.highwire.dtl.DTLVardef@1fbb564org.highwire.dtl.DTLVardef@41da13org.highwire.dtl.DTLVardef@1545106_HPS_FORMAT_FIGEXP M_FIG Graphical abstract C_FIG

neuroscience↗

bioRxiv2020

Multi-Antigenic Virus-like Particle of SARS CoV-2 produced in Saccharomyces cerevisiae as a vaccine candidate

Spike, Envelope and Membrane proteins from the SARS CoV-2 virus surface coat are important vaccine targets. We hereby report recombinant co-expression of the three proteins (Spike, Envelope and Membrane) in a engineered Saccharomyces cerevisiae platform (D-Crypt) and their self-assembly as Virus-like particle (VLP). This design as a multi-antigenic VLP for SARS CoV-2 has the potential to be a scalable vaccine candidate. The VLP is confirmed by transmission electron microscopy (TEM) images of the SARS CoV-2, along with supportive HPLC, Dynamic Light Scattering (DLS) and allied analytical data. The images clearly outline the presence of a "Corona" like morphology, and uniform size distribution.

biochemistry↗

bioRxiv2020

The FDA-approved gold drug Auranofin inhibits novel coronavirus (SARS-COV-2) replication and attenuates inflammation in human cells

SARS-COV-2 has recently emerged as a new public health threat. Herein, we report that the FDA-approved gold drug, auranofin, inhibits SARS-COV-2 replication in human cells at low micro molar concentration. Treatment of cells with auranofin resulted in a 95% reduction in the viral RNA at 48 hours after infection. Auranofin treatment dramatically reduced the expression of SARS-COV-2-induced cytokines in human cells. These data indicate that auranofin could be a useful drug to limit SARS-CoV-2 infection and associated lung injury due to its anti-viral, anti-inflammatory and anti-ROS properties. Auranofin has a well-known toxicity profile and is considered safe for human use.

microbiology↗

bioRxiv2020

The Neuroprotective Beta Amyloid Hexapeptide Core Reverses Deficits in Synaptic Plasticity in the 5xFAD APP/PS1 Mouse Model

Alzheimers disease (AD) is the most common cause of dementia in the aging population. Evidence implicates elevated soluble oligomeric A{beta} as one of the primary triggers during the prodromic phase leading to AD, effected largely via hyperphosphorylation of the microtubule-associated protein tau. At low, physiological levels (pM-nM), however, oligomeric A{beta} has been found to regulate synaptic plasticity as a neuromodulator. Through mutational analysis, we found a core hexapeptide sequence within the N-terminal domain of A{beta} (N-A{beta}core) accounting for its physiological activity, and subsequently found that the N-A{beta}core peptide is neuroprotective. Here, we characterized the neuroprotective potential of the N-A{beta}core against dysfunction of synaptic plasticity assessed in ex vivo hippocampal slices from 5xFAD APP/PS1 mice, specifically hippocampal long-term potentiation (LTP) and long-term depression (LTD). The N-A{beta}core was shown to reverse impairment in synaptic plasticity in hippocampal slices from 5xFAD APP/PS1 model mice, both for LTP and LTD. The reversal by the N-A{beta}core correlated with alleviation of downregulation of hippocampal AMPA-type glutamate receptors in preparations from 5xFAD mice. The action of the N-A{beta}core depended upon a critical di-histidine sequence and involved the PI3 kinase pathway via mTOR. Together, the present findings indicate that the non-toxic N-A{beta}core hexapeptide is not only neuroprotective at the cellular level but is able to reverse synaptic dysfunction in AD-like models, specifically alterations in synaptic plasticity.

neuroscience↗

bioRxiv2019

Novel patterns of complex structural variation revealed across thousands of cancer genome graphs

Cancer genomes often harbor hundreds of somatic DNA rearrangement junctions, many of which cannot be easily classified into simple (e.g. deletion, translocation) or complex (e.g. chromothripsis, chromoplexy) structural variant classes. Applying a novel genome graph computational paradigm to analyze the topology of junction copy number (JCN) across 2,833 tumor whole genome sequences (WGS), we introduce three complex rearrangement phenomena: pyrgo, rigma, and tyfonas. Pyrgo are "towers" of low-JCN duplications associated with early replicating regions and superenhancers, and are enriched in breast and ovarian cancers. Rigma comprise "chasms" of low-JCN deletions at late-replicating fragile sites in esophageal and other gastrointestinal (GI) adenocarcinomas. Tyfonas are "typhoons" of high-JCN junctions and fold back inversions that are enriched in acral but not cutaneous melanoma and associated with a previously uncharacterized mutational process of non-APOBEC kataegis. Clustering of tumors according to genome graph-derived features identifies subgroups associated with DNA repair defects and poor prognosis.

genomics↗

bioRxiv2019

Deep sequencing of 3 cancer cell lines on 2 sequencing platforms

To test the performance of a new sequencing platform, develop an updated somatic calling pipeline and establish a reference for future benchmarking experiments, we sequenced 3 common cancer cell lines along with their matched normal cell lines to great sequencing depths (up to 278X coverage) on both Illumina HiSeqX and NovaSeq sequencing instruments. Somatic calling was generally consistent between the two platforms despite minor differences at the read level. We designed and implemented a novel pipeline for the analysis of tumor-normal samples, using multiple variant callers. We show that coupled with a high-confidence filtering strategy, it improves the accuracy of somatic calls. We also demonstrate the utility of the dataset by creating an artificial purity ladder to evaluate the somatic pipeline and benchmark methods for estimating purity and ploidy from tumor-normal pairs. The data and results of the pipeline are made accessible to the cancer genomics community.

genomics↗

bioRxiv2018

Genetic mechanisms of primary chemotherapy resistance in pediatric acute myeloid leukemia: A report from the TARGET initiative

Acute myeloid leukemias (AML) are characterized by mutations of tumor suppressor and oncogenes, involving distinct genes in adults and children. While certain mutations have been associated with the increased risk of AML relapse, the genomic landscape of primary chemotherapy resistant AML is not well defined. As part of the TARGET initiative, we performed whole-genome DNA and transcriptome (RNA and miRNA) sequencing analysis of pediatric AML with failure of induction chemotherapy. We identified at least three genetic groups of patients with induction failure, including those with NUP98 rearrangements, somatic mutations of WT1 in the absence of NUP98 mutations, and additional recurrent variants including those in KMT2C and MLLT10. Comparison of specimens before and after chemotherapy revealed distinct and invariant gene expression programs. While exhibiting overt therapy resistance, these leukemias nonetheless showed diverse forms of clonal evolution upon chemotherapy exposure. This included selection for mutant alleles of FRMD8, DHX32, PIK3R1, SHANK3, MKLN1, as well as persistence of WT1 and TP53 mutant clones, and elimination or contraction of FLT3, PTPN11, and NRAS mutant clones. These findings delineate genetic mechanisms of primary chemotherapy resistance in pediatric AML, which should inform improved approaches for its diagnosis and therapy.

cancer biology↗

bioRxiv2018

PP-2, a src-kinase inhibitor, is a potential corrector for F508del-CFTR in cystic fibrosis

Cystic fibrosis (CF) is an autosomal recessive disorder caused by mutations in the CF transmembrane conductance regulator (CFTR) gene. The most common mutation in CF, an in-frame deletion of phenylalanine 508, leads to a trafficking defect and endoplasmic reticulum retention of the protein where it becomes targeted for degradation. Successful clinical deployments of ivacaftor and ivacaftor/lumacaftor combination have been an exciting translational development in treating CF. However, their therapeutic effects are variable between subjects and remain insufficient. We used the Library of Integrated Network-based Cellular Signatures (LINCS) database as our chemical pool to screen for candidates. For in silico screening, we integrated connectivity mapping and CF systems biology to identify candidate therapeutic compounds for CF. Following in silico screening, we validated our candidate compounds with (i) an enteroid-based compound screening assay using CF ({Delta}F508/{Delta}F508-CFTR) patient-derived enteroids, (ii) short-circuit current analysis using polarized CF primary human airway epithelial cells and (iii) Western blots to measure F508-del-CFTR protein maturation. We identified 184 candidate compounds with in silico screening and tested 24 of them with enteroid-based forskolin-induced swelling (FIS) assay. The top hit compound was PP2, a known src-kinase inhibitor that induced swelling in enteroid comparable to known CF corrector (lumacaftor). Further validation with Western blot and short-circuit current analysis showed that PP-2 could correct mutant CFTR mis-folding and restore CFTR-mediated transmembrane current. We have identified PP2, a known src-kinase inhibitor, as a novel corrector of {Delta}F508-CFTR. Based on our studies and previous reports, src kinase inhibition may represent a novel paradigm of multi-action therapeutics - corrector, anti-inflammatory, and anti-infective - in CF.

systems biology↗

bioRxiv2018

Structure of Zip2:Spo16, a conserved XPF:ERCC1-like complex critical for meiotic crossover formation

In eukaryotic meiosis, generation of haploid gametes depends on the formation of inter-homolog crossovers, which enable the pairing, physical linkage, and eventual segregation of homologs in the meiosis I division. A class of conserved meiosis-specific proteins, collectively termed ZMMs, are required for formation and spatial control of crossovers throughout eukaryotes. Here, we show that three S. cerevisiae ZMM proteins - Zip2, Zip4, and Spo16 - interact with one another and form a DNA-binding complex critical for crossover formation and control. We determined the crystal structure of a Zip2:Spo16 subcomplex, revealing a heterodimer structurally related to the XPF:ERCC1 endonuclease complex. Zip2:Spo16 lacks an endonuclease active site, but binds specific DNA structures found in early meiotic recombination intermediates. Mutations in multiple DNA-binding surfaces on the Zip2:Spo16 complex severely compromise DNA binding, supporting a model in which the complexs central and HhH domains cooperate to bind DNA. Overall, our data support a model in which the Zip2:Zip4:Spo16 complex binds and stabilizes early meiotic recombination intermediates, then coordinates additional factors to promote crossover formation and license downstream events including synaptonemal complex assembly.

biochemistry↗

bioRxiv2017

Lancet: genome-wide somatic variant calling using localized colored DeBruijn graphs

Reliable detection of somatic variations is of critical importance in cancer research. Lancet is an accurate and sensitive somatic variant caller which detects SNVs and indels by jointly analyzing reads from tumor and matched normal samples using colored DeBruijn graphs. Extensive experimental comparison on synthetic and real whole-genome sequencing datasets demonstrates that Lancet has better accuracy, especially for indel detection, than widely used somatic callers, such as MuTect, MuTect2, LoFreq, Strelka, and Strelka2. Lancet features a reliable variant scoring system which is essential for variant prioritization and detects low frequency mutations without sacrificing the sensitivity to call longer insertions and deletions empowered by the local assembly engine. In addition to genome-wide analysis, Lancet allows inspection of somatic variants in graph space, which augments the traditional read alignment visualization to help confirm a variant of interest. Lancet is available as an open-source program at https://github.com/nygenome/lancet.

bioinformatics↗

bioRxiv2017

Human PGBD5 DNA transposase promotes site-specific oncogenic mutations in rhabdoid tumors

Genomic rearrangements are a hallmark of childhood solid tumors, but their mutational causes remain poorly understood. Here, we identify the piggyBac transposable element derived 5 (PGBD5) gene as an enzymatically active human DNA transposase expressed in the majority of rhabdoid tumors, a lethal childhood cancer. Using assembly-based whole-genome DNA sequencing, we observed previously unknown somatic genomic rearrangements in primary human rhabdoid tumors. These rearrangements were characterized by deletions and inversions involving PGBD5-specific signal (PSS) sequences at their breakpoints, with some recurrently targeting tumor suppressor genes, leading to their inactivation. PGBD5 was found to be physically associated with human genomic PSS sequences that were also sufficient to mediate PGBD5-induced DNA rearrangements in rhabdoid tumor cells. We found that ectopic expression of PGBD5 in primary immortalized human cells was sufficient to promote penetrant cell transformation in vitro and in immunodeficient mice in vivo. This activity required specific catalytic residues in the PGBD5 transposase domain, as well as end-joining DNA repair, and induced distinct structural rearrangements, involving PSS-associated breakpoints, similar to those found in primary human rhabdoid tumors. This defines PGBD5 as an oncogenic mutator and provides a plausible mechanism for site-specific DNA rearrangements in childhood and adult solid tumors.

cancer biology↗