Biology subjects

Shukla, H.

Publications and source records attributed to Shukla, H..

10 recordsLinked to original sources

bioRxiv2026

Polymorphic 3D genome architecture mediated by transposable elements

The three-dimensional (3D) folding of the genome plays a crucial role in genome regulation. However, how 3D genome structure varies between individuals and consequently influences genome function and evolution remains poorly understood. One potential source of this variation is transposable elements (TEs), genomic parasites whose location and composition vary between species and individuals. Hosts typically silence TEs through enriching them with repressive epigenetic marks, turning euchromatic TEs into heterochromatin islands, which were shown to spatially interact with pericentromeric heterochromatin (PCH). Because most TE insertions are present in only a few individuals within Drosophila populations, we asked whether polymorphism in the presence/absence of TEs drives varying 3D structures through TE-PCH spatial interactions. We performed deep-coverage Hi-C of two wild-type Drosophila strains and developed a Hi-C analysis framework enabling allelic comparisons of spatial interactions with PCH. Supporting our hypothesis, nearly 40% of strain-specific euchromatic TEs cause their adjacent euchromatic regions to be spatially closer to PCH than TE-free homologous alleles. These interactions are not limited to specific TE families, and, surprisingly, telomere-proximal TEs show a similar propensity as centromere-proximal TEs to enhance PCH interactions. The most defining feature of TEs involved in PCH interactions is H3K9me3 enrichment, revealing a chromatin-based mechanism for TE-mediated 3D genome organization broadly applicable across TE families and genome locations. Importantly, TEs involved in PCH interactions reduce the expression of adjacent genes and are evolutionarily young, indicating stronger selection against them. Our study reveals a previously uncharacterized mechanism by which TEs influence the function and evolution of host genomes by generating polymorphic 3D genome organization.

genomics↗

bioRxiv2025

Preclinical evaluation of Targeted IL-1β Knockdown via CD44-Immunoliposomes: A Nano-therapy against the Inflammatory Microenvironment

Chronic inflammation, characterized by the infiltration of macrophages and the heightened release of pro-inflammatory cytokines, is the underlying cause of the pathogenesis of many critical diseases. Therapeutic interventions for controlling inflammation via gene knockdown of inflammatory mediators have emerged as a promising approach for regulating uncontrolled inflammation. This study explores the potential of siIL-1{beta}-anti-CD44-Liposomes (SIL) as a potent anti-inflammatory therapy against pro-inflammatory RAW264.7 macrophages via gene specific knockdown of IL-1{beta} mRNA through RNAi, and the subsequent down-regulation of the pro-inflammatory cytokine loop. The designed SIL exhibited a uniform size of 131.1 {+/-} 0.5 nm with a quasi-spherical morphology and sustained release of siIL-1{beta} within 24 hours. The reduction in pro-inflammatory cytokines like IL-1{beta}, TNF-, and IL-6 and inflammatory enzymes iNOS and COX-2; and the simultaneous increase in the anti-inflammatory cytokine IL-4, is indicative of the formulations therapeutic efficacy in reducing inflammation at a cellular level. The effects of SIL on the Macrophage-T cell crosstalk also uncovers the liposomes efficacy in reducing cytokine-mediated T cell effector functions. The nuanced effects of siIL-1{beta}-anti-CD44-Liposomes on in-vivo model of chronic inflammation underscore their potential for precise therapeutic interventions in inflammatory conditions, with multifaceted anti-inflammatory effects on tissue levels and cytokine levels. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=147 SRC="FIGDIR/small/683015v1_ufig1.gif" ALT="Figure 1"> View larger version (61K): org.highwire.dtl.DTLVardef@f4d0f9org.highwire.dtl.DTLVardef@c3c24aorg.highwire.dtl.DTLVardef@8c115org.highwire.dtl.DTLVardef@bc863d_HPS_FORMAT_FIGEXP M_FIG Schematic representation of the study: CD44 receptors are elevated in LPS activated macrophages, which can be targeted by using anti-CD44 Liposomes, for RNA therapy. IL-1 knockdown via siRNA leads to lower inflammatory nature of macrophages, compromising its the antigen presentation and T-cell activation. This lowers the cytokine storm in inflammatory milieu and lower tissue damage can be achieved. Created with BioRender.com. C_FIG

pharmacology and toxicology↗

bioRxiv2025

Tumor Organoids Modeling Reveals Timed Responses and Interplay of Radiotherapy and Chemotherapy in Pancreatic Cancer

PurposeModel the therapeutic effects of chemotherapy, radiotherapy, and chemoradiotherapy to study the heterogeneous responses of patient-derived tumor organoids (PDTOs). MethodsWe proposed novel mathematical forecasting frameworks based on logistic growth ordinary differential equations (ODEs) and the characteristics of tumor responses to model the therapeutic effects of chemo- and radiation-induced killing. To validate the models, we cultured PDTOs from different patients and treated them with radiation (4 Gy and 8 Gy), chemotherapy (FOLFIRINOX at a previously determined organoid-specific IC50 dose), and combined regimens, respectively. The diameters of 20-40 organoids per patient were tracked and measured using brightfield images up to 7 or 9 days following each treatment to capture organoid growth dynamics data, which was used for model fitting. The accuracy of the modeling was evaluated by the average normalized mean squared errors (NMSE) of data fitting. ResultsThe proposed mathematical modeling frameworks accurately captured the observed growth dynamics of three PDTO samples after chemotherapy, radiotherapy, and chemoradiotherapy, as reflected by the average NMSE of data fitting, which are all close to zero (less than 0.0045). The fitted parameters, including killing strength, effect window, and peak killing timing, revealed significant heterogeneities in treatment responses across different PDTOs. Chemotherapy shows pronounced effectiveness in the early stage, while radiotherapy exhibits the effect later in the first week, and a significantly stronger secondary response occurs one week after radiation. Chemoradiotherapy combines the strengths of both modalities, producing pronounced effects in both response phases, with modeling results suggesting that the radiation-induced killing effect may play a dominant role in the synergistic interaction. ConclusionsOur modeling frameworks demonstrated high accuracy in modeling the heterogeneous therapeutic responses of PDTOs and provided insights into the dynamic killing effects and interplay between chemotherapy and radiotherapy. The modeling of therapeutic responses of PDTOs provides a valuable tool for optimizing treatment regimens and informing clinical trial design, thereby improving the efficacy of personalized medicine.

cancer biology↗

bioRxiv2024

Genetic variation in recalcitrant repetitive regions of the Drosophila melanogaster genome

Many essential functions of organisms are encoded in highly repetitive genomic regions, including histones involved in DNA packaging, centromeres that are core components of chromosome segregation, ribosomal RNA comprising the protein translation machinery, telomeres that ensure chromosome integrity, piRNA clusters encoding host defenses against selfish elements, and virtually the entire Y chromosome. These regions, formed by highly similar tandem arrays, pose significant challenges for experimental and informatic study, impeding sequence-level descriptions essential for understanding genetic variation. Here, we report the assembly and variation analysis of such repetitive regions in Drosophila melanogaster, offering significant improvements to the existing community reference assembly. Our work successfully recovers previously elusive segments, including complete reconstructions of the histone locus and the pericentric heterochromatin of the X chromosome, spanning the Stellate locus to the distal flank of the rDNA cluster. To infer structural changes in these regions where alignments are often not practicable, we introduce landmark anchors based on unique variants that are putatively orthologous. These regions display considerable structural variation between different D. melanogaster strains, exhibiting differences in copy number and organization of homologous repeat units between haplotypes. In the histone cluster, although we observe minimal genetic exchange indicative of crossing over, the variation patterns suggest mechanisms such as unequal sister chromatid exchange. We also examine the prevalence and scale of concerted evolution in the histone and Stellate clusters and discuss the mechanisms underlying these observed patterns.

genomics↗

bioRxiv2022

Near-chromosomal de novo assembly of Bengal tiger genome reveals genetic hallmarks of apex-predation

The tiger, a poster child for conservation, remains an endangered apex predator. Continued survival and recovery will require a comprehensive understanding of their genetic diversity and the use of such information for population management. A high-quality tiger genome assembly will be an important tool for conservation genetics, especially for the Indian tiger, the most abundant subspecies in the wild. Here, we present high-quality near-chromosomal genome assemblies of a female and a male wild Indian tiger (Panthera tigris tigris). Our assemblies had a scaffold N50 of >140{square}Mb, with 19{square}scaffolds, corresponding to the 19 numbered chromosomes, containing 95% of the genome. Our assemblies also enabled detection of longer stretches of runs of homozygosity compared to previous assemblies which will improve estimates of genomic inbreeding. Comprehensive genome annotation identified 26,068 protein-coding genes, including several gene families involved in key morphological features such as the teeth, claws, vision, olfaction, taste and body stripes. We also identified 301 microRNAs, 365 small nucleolar RNAs, 632 tRNAs and other noncoding RNA elements, several of which are predicted to regulate key biological pathways that likely contribute to tigers apex predatory traits. We identify signatures of positive selection in the tiger genome that are consistent with the Panthera lineage. Our high-quality genome will enable use of non-invasive samples for comprehensive assessment of genetic diversity, thus supporting effective conservation and management of wild tiger populations.

genomics↗

bioRxiv2022

Species-specific chromatin landscape determines how transposable elements shape genome evolution

Transposable elements (TEs) are selfish genomic parasites that increase their copy number at the expense of host fitness. The "success," or genome-wide abundance, of TEs differs widely between species. Deciphering the causes for this large variety in TE abundance has remained a central question in evolutionary genomics. We previously proposed that species-specific TE abundance could be driven by the inadvertent consequences of host-direct epigenetic silencing of TEs--the spreading of repressive epigenetic marks from silenced TEs into adjacent sequences. Here, we compared this TE-mediated "epigenetic effect" in six species in the Drosophila melanogaster subgroup to dissect step-by-step the role of such effect in determining genomic TE abundance. We found that TE-mediated spreading of repressive marks is prevalent and substantially varies across and even within species. While this TE-mediated effect alters the epigenetic states of adjacent genes, we surprisingly discovered that the transcription of neighboring genes could reciprocally impact this spreading. Importantly, our multi- species analysis provides the power and appropriate phylogenetic resolution to connect species-specific host chromatin regulation, TE-mediated epigenetic effects, the strength of natural selection against TEs, and genomic TE abundance unique to individual species. Our findings point towards the importance of host chromatin landscapes in shaping genome evolution through the epigenetic effects of a selfish genetic parasite.

genomics↗

bioRxiv2021

Preclinical Efficacy of IMM-BCP-01, a Highly Active Patient-Derived Anti-SARS-CoV-2 Antibody Cocktail

Using an unbiased interrogation of the memory B cell repertoire of convalescent COVID-19 patients, we identified human antibodies that demonstrated robust antiviral activity in vitro and efficacy in vivo against all tested SARS-CoV-2 variants. Here, we describe the pre-clinical characterization of an antibody cocktail, IMM-BCP-01, that consists of three unique, patient-derived recombinant neutralizing antibodies directed at non-overlapping surfaces on the SARS-CoV-2 spike protein. Two antibodies, IMM20184 and IMM20190 directly block spike binding to the ACE2 receptor. Binding of the third antibody, IMM20253, to its unique epitope on the outer surface of RBD, alters the conformation of the spike trimer, promoting release of spike monomers. These antibodies decreased SARS-CoV-2 infection in the lungs of Syrian golden hamsters, and efficacy in vivo efficacy was associated with broad antiviral neutralizing activity against multiple SARS-CoV-2 variants and robust antiviral effector function response, including phagocytosis, ADCC, and complement pathway activation. Our pre-clinical data demonstrate that the three antibody cocktail IMM-BCP-01 shows promising potential for preventing or treating SARS-CoV-2 infection in susceptible individuals. One sentence summaryIMM-BCP-01 cocktail triggers Spike Trimer dissociation, neutralizes all tested variants in vitro, activates a robust effector response and dose-dependently inhibits virus in vivo.

immunology↗

bioRxiv2021

Genomic evidence for inbreeding depression and purging of deleterious genetic variation in Indian tigers

Increasing habitat fragmentation leads to wild populations becoming small, isolated, and threatened by inbreeding depression. However, small populations may be able to purge recessive deleterious alleles as they become expressed in homozygotes, thus reducing inbreeding depression and increasing population viability. We used whole genomes sequencing from 57 tigers to estimate individual inbreeding and mutation loads in a small-isolated, and two large-connected populations in India. As expected, the small-isolated population had substantially higher average genomic inbreeding (FROH=0.57) than the large-connected (FROH=0.35 and FROH=0.46) populations. The small-isolated population had the lowest loss-of-function mutation load, likely due to purging of highly deleterious recessive mutations. The large populations had lower missense mutation loads than the small-isolated population, but were not identical, possibly due to different demographic histories. While the number of the loss-of-function alleles in the small-isolated population was lower, these alleles were at high frequencies and homozygosity than in the large populations. Together, our data and analyses provide evidence of (a) high mutation load; (b) purging and (c) the highest predicted inbreeding depression, despite purging, in the small-isolated population. Frequency distributions of damaging and neutral alleles uncover genomic evidence that purifying selection has removed part of the mutation load across Indian tiger populations. These results provide genomic evidence for purifying selection in both small and large populations, but also suggest that the remaining deleterious alleles may have inbreeding associated fitness costs. We suggest that genetic rescue from sources selected based on genome-wide differentiation should offset any possible impacts of inbreeding depression. Significance statementHabitat fragmentation is sequestering species into small and isolated populations with high chances of extinction. But are small and isolated populations at risk of extinction from inbreeding depression, or does inbreeding and purging of deleterious alleles reduce such threat? Using whole genomes from several wild Indian tiger populations, we provide evidence supporting purging of highly deleterious variants in a small isolated population. However, our analyses also indicate that the remaining highly deleterious alleles are at high frequencies, suggesting continued inbreeding depression despite some successful purging. We discuss the implications of our results for conservation, including possible genomics-informed genetic rescue strategies.

evolutionary biology↗

bioRxiv2019

Metabarcoding for parallel identification of species, sex and diet: an application to the conservation of globally-threatened Gyps vultures

An understanding of the factors affecting the diet composition of critically endangered Gyps vultures in the Indian subcontinent has important applications to the design of effective conservation strategies. After suffering a massive decline (~99%) in numbers caused by veterinary use of the nephrotoxic drug diclofenac, vultures now persist in very low numbers, mostly concentrated within or near National Parks. This spatial association might be attributed to the availability of wild ungulate carcasses free from toxic veterinary drugs. Hence, quantification of vulture diets and the factors affecting them is critical to test this hypothesis. Here, we describe and validate a robust \"field-to-benchtop-to-desktop\" metabarcoding workflow for assessing the species- and sex-specific diet of these obligate scavengers from non-invasively collected faecal samples. Seven metabarcodes targeting mitochondrial and nuclear genes were used to simultaneously determine the vulture species identity, sex and species consumed. The amplicons were sequenced using an Illumina Miseq platform. We included controls and three replicates per sample to establish a series of non-arbitrary thresholds to filter the sequence data and eliminate cross-contamination, PCR/sequencing errors and false positives. Using this strategy enabled identification of species and sex for all samples. Diet species-specific sequences could be identified with high taxonomic resolution for 93% of samples. Out of the seven metabarcodes, just four were sufficient to meet the objectives. From this preliminary analysis, domestic livestock seem to be the most frequently consumed diet item across samples from inside and outside protected habitats. Our method provides a rapid and reliable tool for describing large-scale variation in consumption of domestic versus wild species in the diet of these species, paving the way for a better understanding of the role protected areas play in persistence and recovery of the remaining Gyps vulture populations in the wild.

genetics↗

bioRxiv2018

Draft genome of the liver fluke Fasciola gigantica

Fascioliasis is a neglected food-borne disease caused by liver flukes (genus Fasciola) and affects more than 200 million people worldwide. Despite technological advances, little is known about the molecular biology and biochemistry of the fluke. We present the draft genome of Fasciola gigantica for the first time. The assembled draft genome has a size of ~1.04 Gb with an N50 of 129 kb. A total of 20,858 genes were predicted. The de novo repeats identified in the draft genome were 46.85%. In pathway analysis, all the genes of glycolysis, Krebs cycle and fatty acid metabolism were found to be present, but the key genes for fatty acid production in fatty acid biosynthesis were missing. This indicates that the fatty acid required for the survival of the fluke may be acquired from the host bile. The genomic information will provide a comprehensive resource to facilitate the development of novel interventions for fascioliasis control.

bioinformatics↗