Biology subjects

Goel, S.

Publications and source records attributed to Goel, S..

30 records · Page 2Linked to original sources

bioRxiv2023

NEMO reshapes the protein aggregate interface and promotes aggrephagy by co-condensation with p62

NEMO is a ubiquitin-binding protein which regulates canonical NF-{kappa}B pathway activation in innate immune signaling, cell death regulation and host-pathogen interactions. Here we identified an NF-{kappa}B-independent function of NEMO in proteostasis regulation by promoting autophagosomal clearance of protein aggregates. NEMO-deficient cells accumulate misfolded proteins upon proteotoxic stress and are vulnerable to proteostasis challenges. Moreover, a patient with a mutation in the NEMO gene resulting in defective binding of NEMO to linear ubiquitin chains, developed a widespread mixed brain proteinopathy, including -synuclein, tau and TDP-43 pathology. NEMO amplifies linear ubiquitylation at -synuclein aggregates and promotes the local concentration of p62 into foci. In vitro, NEMO lowers the threshold concentrations required for ubiquitin-dependent phase transition of p62. In summary, NEMO reshapes the aggregate surface for efficient autophagosomal clearance by providing a mobile phase at the aggregate interphase favoring co-condensation with p62.

cell biology↗

bioRxiv2021

Reovirus sensitizes microsatellite stable colorectal cancer to anti-PD-1 treatment via cross-talk in innate and adaptive immune systems

BackgroundMicrosatellite stable (MSS) colorectal cancer (CRC) represents ~85% of all CRCs. These tumors are poorly immunogenic and largely resistant to immunotherapy, necessitating a need to develop new immune enhancing strategies. Oncolytic reovirus has a high propensity to replicate in KRAS mutant tumors which account for ~50% of MSS CRCs. Current study explores the ability of reovirus to potentiate the effect of immune checkpoint inhibition in MSS CRC. MethodsEffectiveness of reovirus infection was quantified through MTT assay for cell viability, and expression of immune-response genes by flow cytometry, RT-qPCR, and microarray. Computational analysis of differentially expressed genes was performed by TAC, DAVID and STRING. Combinatorial approach using anti-PD-1 monoclonal antibody was assessed in ex vivo and in vivo models. Live-cell imaging, tumor volume and survival were measured for quantification of anti-tumor activity. Expression of pattern recognition receptors (PRRs), cell surface and activation markers of immune cells, and PD-1/PD-L1 axis were studied using multi-color flow cytometry, immunoblotting, immunohistochemistry, and immunofluorescence. ResultsReovirus infection exerted growth arrest and expression of immune-response genes in CRCs cell lines in a KRAS-dependent manner. However, microsatellite instability, rather than KRAS status determined immune-repose pathways, functionalities and biological processes post-reovirus infection. Furthermore, reovirus significantly enhanced the anti-tumor activity of anti-human PD-1 [nivolumab] treatment in MSS CRC cell lines ex vivo. Similarly, reovirus increased the activity of anti-mouse PD-1 treatment in the CT26 [MSS, KRASMut], but not the MC38 [MSI, KRASWt] syngeneic mouse model of CRC. Combinatorial treatment has reduced the proliferative index, increased apoptosis and differentially altered PD-L1/PD-1 signaling among CT26 and MC38 tumors. Activation of innate immune system and expression of PRRs and antigen presentation markers were observed under reovirus and anti-PD-1 treatment that additionally reduced immunosuppressive macrophages. This led to an increase in T cell subsets, increase in effector T cell activation, and decrease in exhaustion markers specifically within CT26 microenvironment. ConclusionThe current study systematically evaluates immune characteristics and immune microenvironment of CRC under reovirus/anti-PD-1 combination treatment that proves increased effectiveness among MSS compared to MSI CRCs. This is a promising regimen warranting translation into clinical trials. One Sentence SummaryOncolytic reovirus alters innate and adaptive immune system and potentiates MSS type colorectal cancer to checkpoint inhibition therapy.

cancer biology↗

bioRxiv2021

DeepTrio: Variant Calling in Families Using Deep Learning

Every human inherits one copy of the genome from their mother and another from their father. Parental inheritance helps us understand the transmission of traits and genetic diseases, which often involve de novo variants and rare recessive alleles. Here we present DeepTrio, which learns to analyze child-mother-father trios from the joint sequence information, without explicit encoding of inheritance priors. DeepTrio learns how to weigh sequencing error, mapping error, and de novo rates and genome context directly from the sequence data. DeepTrio has higher accuracy on both Illumina and PacBio HiFi data when compared to DeepVariant. Improvements are especially pronounced at lower coverages (with 20x DeepTrio roughly equivalent to 30x DeepVariant). DeepTrio includes pre-trained models for Illumina WGS, Illumina exome, and PacBio HiFi.

bioinformatics↗

bioRxiv2021

Haplotype-aware variant calling enables high accuracy in nanopore long-reads using deep neural networks

Long-read sequencing has the potential to transform variant detection by reaching currently difficult-to-map regions and routinely linking together adjacent variations to enable read based phasing. Third-generation nanopore sequence data has demonstrated a long read length, but current interpretation methods for its novel pore-based signal have unique error profiles, making accurate analysis challenging. Here, we introduce a haplotype-aware variant calling pipeline PEPPER-Margin-DeepVariant that produces state-of-the-art variant calling results with nanopore data. We show that our nanopore-based method outperforms the short-read-based single nucleotide variant identification method at the whole genome-scale and produces high-quality single nucleotide variants in segmental duplications and low-mappability regions where short-read based genotyping fails. We show that our pipeline can provide highly-contiguous phase blocks across the genome with nanopore reads, contiguously spanning between 85% to 92% of annotated genes across six samples. We also extend PEPPER-Margin-DeepVariant to PacBio HiFi data, providing an efficient solution with superior performance than the current WhatsHap-DeepVariant standard. Finally, we demonstrate de novo assembly polishing methods that use nanopore and PacBio HiFi reads to produce diploid assemblies with high accuracy (Q35+ nanopore-polished and Q40+ PacBio-HiFi-polished).

bioinformatics↗

bioRxiv2021

Improving variant calling using population data and deep learning

Large-scale population variant data is often used to filter and aid interpretation of variant calls in a single sample. These approaches do not incorporate population information directly into the process of variant calling, and are often limited to filtering which trades recall for precision. In this study, we develop population-aware DeepVariant models with a new channel encoding allele frequencies from the 1000 Genomes Project. This model reduces variant calling errors, improving both precision and recall in single samples, and reduces rare homozygous and pathogenic clinvar calls cohort-wide. We assess the use of population-specific or diverse reference panels, finding the greatest accuracy with diverse panels, suggesting that large, diverse panels are preferable to individual populations, even when the population matches sample ancestry. Finally, we show that this benefit generalizes to samples with different ancestry from the training data even when the ancestry is also excluded from the reference panel.

bioinformatics↗

bioRxiv2020

An Extensive Sequence Dataset of Gold-Standard Samples for Benchmarking and Development

Accurate standards and extensive development datasets are the foundation of technical progress. To facilitate benchmarking and development, we sequence 9 samples, covering the Genome in a Bottle truth sets on multiple instruments (NovaSeq, HiSeqX, HiSeq4000, PacBio Sequel II System) and sample preparations (PCR-Free, PCR-Positive) for both whole genome and multiple exome kits. We benchmark pipelines, quantifying strengths and limitations for sequencing and analysis methods. We identify variability within and between instruments, preparation methods, and analytical pipelines, across various sequencing depths. We discuss the relevance of this variability to downstream analyses, and strategies to reduce variability.

genomics↗

bioRxiv2020

precisionFDA Truth Challenge V2: Calling variants from short- and long-reads in difficult-to-map regions

The precisionFDA Truth Challenge V2 aimed to assess the state-of-the-art of variant calling in difficult-to-map regions and the Major Histocompatibility Complex (MHC). Starting with FASTQ files, 20 challenge participants applied their variant calling pipelines and submitted 64 variant callsets for one or more sequencing technologies (~35X Illumina, ~35X PacBio HiFi, and ~50X Oxford Nanopore Technologies). Submissions were evaluated following best practices for benchmarking small variants with the new GIAB benchmark sets and genome stratifications. Challenge submissions included a number of innovative methods for all three technologies, with graph-based and machine-learning methods scoring best for short-read and long-read datasets, respectively. New methods out-performed the 2016 Truth Challenge winners, and new machine-learning approaches combining multiple sequencing technologies performed particularly well. Recent developments in sequencing and variant calling have enabled benchmarking variants in challenging genomic regions, paving the way for the identification of previously unknown clinically relevant variants.

bioinformatics↗

bioRxiv2020

Transcriptional network involving ERG and AR orchestrates Distal-Less Homeobox 1 mediated prostate cancer progression.

Nearly half of the prostate cancer (PCa) cases show elevated levels of ERG oncoprotein due to TMPRSS2-ERG gene fusion. Here, we demonstrate ERG mediated upregulation of Distal-less homeobox-1 (DLX1), an established PCa biomarker. Using series of functional assays, we show DLX1 elicits oncogenic properties in prostate epithelial cells, and abrogating its function leads to reduced tumor burden in mouse xenografts. Clinically, [~]60% of the PCa patients exhibit high DLX1 levels, while [~]50% of these cases also harbor elevated ERG associated with aggressive disease and poor survival probability. Mechanistically, we show that ERG gets recruited onto DLX1 promoter and interacts with its enhancer-bound androgen receptor (AR) and FOXA1 to regulate DLX1 expression in TMPRSS2-ERG positive cases. Alternatively, in ERG-negative cases, DLX1 is regulated by AR/AR-V7 and FOXA1. Importantly, BET bromodomain inhibitors disrupt the transcriptional regulators of DLX1 and its associated oncogenic properties, signifying their efficacy in treatment of DLX1-positive PCa patients.

cancer biology↗

bioRxiv2019

De novo transcriptome profiling of mustard aphid (Lipaphis erysimi) and differential expression of transcripts associated with developmental stages, feeding and non-feeding conditions

Lipaphis erysimi is a Brassicaceae specialist aphid, which causes significant losses in yield and/or reduction of oil content of vegetable and oilseed brassicas and is a major pest in the Indian subcontinent. This study reports the de novo transcriptome of L. erysimi for the first time. We also present a comparative analysis of nymphs and adult transcriptomes to study the differential expression profiles associated with different developmental stages as well as different feeding conditions. For this, RNA-seq was performed on three different biological samples adults, nymphs (with all nymph stages pooled) and adults starved for 3 hours (referred to as Adult Feeding, AF; Nymphs Feeding, NF, and Adult Starved_3 hr, ANF samples henceforth). A final transcriptome comprising 52,652 transcripts of 1064bp average length and N50 value of 1806 bp was generated. A total of 27,112 transcripts were annotated with insect proteins from SwissProt, of which 4128 transcripts were components of 165 KEGG pathways. A total of 17,296 transcripts were classified based on their Gene Ontology. Potential transcripts for host selection, detoxification, salivary proteins and effectors, molecular chaperones and developmental genes were identified. A total of 23,532 transcripts that remained unannotated were subjected to BLAST against aphid sequences available at AphidBase and a total of 3091 transcripts had hits with sequences of other aphids in the database, out of which 1380 had protein hits. A total of 20441 found to share no homology to any sequence available in the public domain and could therefore represent novel aphid genes or sequences that are unique to L. erysimi. This is an exploratory study with no biological replicates. However, the significant repertoire of feeding- and development-related genes and their differential expression profiles generated in this study adds to the limited data available on L. erysimi and it would facilitate studies on the molecular basis of aphid feeding and development. This could also allow identification of novel target genes for development of RNAi-based aphid control methods.

systems biology↗

bioRxiv2019

Membrane integration and topology of RIFIN and STEVOR proteins of the Plasmodium falciparum parasite

The malarial parasite Plasmodium, infects red blood cells by remodeling them and transporting its own proteins to their cell surface. These proteins trigger adhesion of infected cells to uninfected cells (rosetting), and to the vascular endothelium, obstructing blood flow and contributing to pathogenesis. RIFINs (P. falciparum-encoded repetitive interspersed families of polypeptides) and STEVORs (subtelomeric variable open reading frame), are two classes of proteins that are involved in rosetting. Here we study the membrane insertion and topology of three RIFIN and two STEVOR proteins, employing a well-established assay that uses N-linked glycosylation of sites within the protein as a measure to assess the topology a protein adopts when inserted into the ER membrane. Our results indicate that all the proteins tested assume an overall topology of Ncyt-Ccyt, with predicted transmembrane helices TM1 and TM3 integrated into the ER membrane. We also show that the segments predicted as TM2 do not reside in the membrane. Our conclusions are consistent with other recent topology studies on RIFIN and STEVOR proteins.

biochemistry↗

bioRxiv2018

Polycomb Group protein EZH2-mediated transcriptional repression of microRNA-338/-421 drives SPINK1-positive prostate cancer

PurposeSerine Peptidase Inhibitor, Kazal type-1 (SPINK1) overexpression defines the second most recurrent and aggressive prostate cancer (PCa) subtype. However, the underlying molecular mechanism and pathobiology of SPINK1 in PCa remains largely unknown.\n\nExperimental DesignMicroRNA-prediction tools were employed to examine the SPINK1-3UTR for miRNAs binding. Luciferase reporter assays were performed to confirm the SPINK1-3UTR binding of shortlisted miR-338-5p/miR-421. Further, miR-338-5p/-421 overexpressing cancer cells (SPINK1-positive) were evaluated for oncogenic properties using cell-based functional assays and mice xenograft model. Global gene expression profiling was performed to unravel the biological pathways altered by miR-338-5p/-421. Immunohistochemistry and RNA in-situ hybridization was carried-out on PCa patients tissue microarray for SPINK1 and EZH2 expression respectively. Chromatin immunoprecipitation assay was performed to examine EZH2 occupancy on the miR-338-5p/-421 regulatory regions. Bisulfite sequencing and methylated DNA-immunoprecipitation was performed on PCa cell lines and patients specimens.\n\nResultsWe established a critical role of miRNA-338-5p/-421 in post-transcriptional regulation of SPINK1. Ectopic expression of miRNA-338-5p/-421 in SPINK1-positive PCa cells abrogate oncogenic properties including cell-cycle progression, stemness and drug resistance, and show reduced tumor burden and distant metastases in mice model. Importantly, we show SPINK1-positive PCa patients exhibit increased EZH2 expression, suggesting its role in miRNA-338-5p/-421 epigenetic silencing. Furthermore, presence of CpG dinucleotide DNA methylation marks on the regulatory regions of miR-338-5p/-421 in SPINK1-positive PCa cells and patients specimens confirms epigenetic silencing.\n\nConclusionOur findings revealed that miRNA-338-5p/-421 are epigenetically silenced in SPINK1-positive PCa, while restoring the expression of these miRNAs using epigenetic drugs or synthetic mimics could abrogate SPINK1-mediated oncogenesis.\n\nTRANSLATIONAL IMPACTWe establish a regulatory model involving the functional interplay between SPINK1, miRNA-338-5p/miRNA-421 and EZH2, thereby, revealing hitherto unknown mechanism of SPINK1 up-regulation in SPINK1-positive subtype. Our findings provide a strong rationale for the development of potential therapeutic strategies for SPINK1-positive malignancies. We demonstrate that restoring miRNA-338-5p/miRNA-421 expression using epigenetic drugs including DNMTs inhibitors in combination with HDACs or HKMTs inhibitors or miRNA synthetic mimics in SPINK1-positive prostate cancer abrogate SPINK1-mediated oncogenicity. The major findings of this study will not only advance the prostate cancer field, but will also be valuable for treatment and disease management of other SPINK1-positive malignancies.

cancer biology↗

bioRxiv2018

Analysis of genetic control and QTL mapping of essential wheat grain quality traits in a recombinant inbred population

Wheat cultivars are genetically crossed for improving end use quality for apt traits as per need of baking industry and broad consumers preferences. The processing and baking qualities of bread wheat underlie into genetic make-up of a variety and influence by environmental factors and their interactions. WL711 and C306 derived recombinant inbred lines (RILs) population of 206 was used for phenotyping of quality related traits in three different environmental conditions. The genetic analysis of quality traits showed considerable variation for measurable quality traits with normal distribution and transgressive segregation across the years. From the 206 RIL, few RILs found to be superior to those of the parental cultivars for key quality traitsindicating their potential usefor improvement of end use quality and also suggestingprobability of finding new alleles and allelic combinations from the RIL population. A genetic linkage map including 346 markers was constructed withtotal map distance of 4526.8cM andinterval distance between adjacent markersof 12.9cM. Mapping analysis identified 38 putative QTLs for 13 quality related traits with QTLs explaining 7.9% - 16.8% phenotypic variation spanning over 14 chromosomes i.e. 1A, 1B, 1D, 2A, 2D, 3B, 3D, 4A, 4B, 4D, 5D, 6A, 7A and 7B. Major novel QTLs regions for quality traits have been identified on several chromosome in studied RIL population posing their potential role in marker assisted selection for better bread making quality after validation.

genetics↗