Biology subjects

Karmakar, S.

Publications and source records attributed to Karmakar, S..

At least 19 recordsLinked to original sources

bioRxiv2026

From Wandering Thoughts to Mental Stillness: A Minimal Computational Statistical-Mechanical Model of Driven Relaxation

Reduced spontaneous thought during meditation is often described phenomenologically, but a mechanistic account should specify how a dynamical system can enter and persist in such a regime. Here we formulate a minimal statistical-mechanical model in which a dimensionless coordinate, q, represents a coarse-grained propensity for spontaneous-thought activity, and a slowly relaxing variable, r of t, represents the history-dependent effect of an attentional manipulation. The model combines a tilted double-well landscape with Langevin noise, allowing a high-q and a low-q dynamical regime to coexist. We numerically integrate the model under a controlled focusing protocol followed by removal of the drive and quantify low-state occupancy, first-passage times, dwell times, occupancy entropy, and transition entropy. For the chosen dimensionless parameterization, increasing the drive amplitude produces a sharp crossover in low-state occupancy near the deterministic spinodal threshold, A-sub-sp, approximately equal to 0.0148. At a noise intensity of D equal to 0.01, the probability of occupying the low-q regime at drive removal increases from 0.094 at A equal to zero to 0.994 at A equal to 0.02, while the mean first-passage time decreases from 48.0 to 13.5 model time units. Noise produces a non-monotonic effect: for A equal to 0.01, low-state occupancy at drive removal is only 0.033 at D equal to 0.0025, rises to 0.842 at D equal to 0.02, and falls to 0.718 at D equal to 0.04. Strong driving also reduces the effective number of occupied states while changing transition entropy much less, illustrating why a single entropy measure cannot identify thoughtlessness. These results do not establish that human neural dynamics obey this model. Instead, they show that a driven stochastic bistable system can generate the qualitative sequence of focusing, switching, relaxation, and metastable low-activity residence, and they provide quantitative predictions that can be tested against time-resolved neural recordings and independent reports of subjective experience.

animal behavior and cognition↗

bioRxiv2026

The Core Promoter has an active role in interpreting patterning information

Precise spatiotemporal control of gene expression is conventionally attributed to distal enhancers. The extent to which different core promoters actively shape transcriptional output from enhancers remains unclear, especially in the context of genes that contain alternate promoters. We have explored these questions using the Drosophila grainy head (grh) locus, and tested the activities conferred by its alternate promoters in combination with a well characterised Notch responsive enhancer. We find that, of the alternate promoters tested, only the grh short promoter is competent to drive neuroblast-specific expression. Notably, when paired with identical enhancers, grh-short and grh-medium promoters drive distinct expression patterns. The activity of other core promoters, such as hsp70, is also selective. Together the results establish that the core promoter acts alongside enhancers to confer specificity and robustness on gene expression profiles.

developmental biology↗

bioRxiv2026

Gut bacterium of Leishmania donovani-non-transmitting sand fly induces IL-23~Th17 axis to protect hosts from drug-resistant visceral leishmaniasis

Background: Visceral leishmaniasis (VL) is caused by Leishmania donovani and differentiates under the influence of biotope-specific gut bacteria in the Leishmania donovani-transmitting sand fly vector, Phlebotomus argentipes, which coexists with the Leishmania donovani-non-transmitting Sergentomyia species in the Indian subcontinent. Hypothesis: Sergentomyia are primarily herpetophilic. Interestingly, we identified a specific population of anthropophilic Sergentomyia babu babu, a proven naturally non-competent vector for transmitting Leishmania donovani, with high abundance and human-bite incidences in VL-non-endemic regions, compared with VL-endemic regions of India. Thus, the possibility of egesting gut bacteria to humans from VL-non-transmitting Sergentomyia and inducing downstream immune cascades remains undeniable. Innovation: We characterized gut bacteria from anthropophilic Sergentomyia babu babu in both VL-endemic and VL-non-endemic regions of India and found that a heat-inactivated formulation of Heyndrickxia sporothermodurans [NCBI Sequence Read Archive (SRA): PRJNA1403236], isolated from anthropophilic Sergentomyia babu babu from a VL-non-endemic region, prevented and cured L. donovani infection by inducing IL-23~Th17. More importantly, it also prevented the initial parasite establishment by inducing phagocytosis, ROS, and TNF- in circulatory Ly6G-CD11b+Ly6Chi monocytes. To the best of our knowledge, this is the first report confirming that the gut bacteria of VL-non-transmitting sand flies are critical and need to be investigated in depth for developing therapeutics and/or prophylactics against visceral leishmaniasis (VL), and suggests that possible bite-transfer of gut bacteria from disease-non-transmitting vectors to hosts may restrict disease pathogenicity. Further clinical trials are needed to warrant (Indian patent: 202511041013, April 28, 2025).

immunology↗

bioRxiv2026

CD40 Signaling Restricts Retrograde Viral Spread and Provides Neuroprotection to Retinal Ganglion Cells in a Murine β-Coronavirus Model of Optic Neuritis

CD40, a co-stimulatory receptor of the tumor necrosis factor receptor superfamily expressed on microglia and macrophages, is an upstream regulator of innate antiviral defense in coronavirus-induced neuroinflammation, but its specific role in the visual system remains undefined. Here, we demonstrate that CD40 signaling is essential for restricting retrograde axonal transport of the murine {beta}-coronavirus RSA59 from the brain to the retina and for preventing chronic neurodegeneration in a model of viral optic neuritis. Wild-type and CD40-/- mice were intracranially inoculated with RSA59, and viral burden, neuroinflammation, and neurodegeneration were assessed at acute (day 5), bridging (day 7), and chronic (day 30) stages. CD40-/- mice exhibited significantly increased clinical severity and [~]30% mortality by day 12 post-infection, compared to 100% survival in WT mice. CD40 deficiency resulted in elevated viral loads in the optic nerve and enhanced retrograde viral dissemination across all retinal layers, whereas in WT mice, the virus was largely confined to the ganglion cell layer. CD40-/- mice exhibited impaired early microglial activation and compensatory astrogliosis during the acute and bridging phases. By day 30 p.i., although viral-N protein was undetectable by immunohistochemistry in both genotypes, CD40-/- optic nerves retained significantly higher persistent viral RNA and exhibited extensive demyelination, oligodendrocyte loss, axonal depletion, and upregulation of phagocytic markers. Critically, CD40-/- retinas showed persistent astrogliosis, accumulation of phagocytic microglia/macrophages, and a significant loss of Brn3a+ retinal ganglion cells. These findings establish CD40 as a critical molecular node governing coronavirus optic neuritis, linking early innate immune regulation to long-term neuronal survival.

immunology↗

bioRxiv2026

Pattern of Circulating Mesenchymal Stromal Cells and Hematopoietic Progenitor and Stem Cells in the Peripheral Blood of Trauma Patients with and without Hemorrhagic Shock

Circulating stem and progenitor cells (SPCs), including mesenchymal stromal cells (MSCs) and hematopoietic stem/progenitor cells (HSPCs), are mobilised after tissue injury but their temporal behaviour after hemorrhagic shock (HS) and relationship to cytokine milieus and outcome remain unclear. In a prospective observational cohort at JPN Apex Trauma Centre, AIIMS, New Delhi we studied 100 participants: 50 trauma patients with hemorrhagic shock and traumatic brain injury (HS index group), 25 trauma patients without HS, and 25 minor-injury controls. Peripheral blood was collected at admission (day 0) for all groups and additionally at days 3, 7 and 14 for the HS group. PBMCs were phenotyped by flow cytometry (HSPC markers: CD45, CD123, CD38, CD34; MSC markers: CD105, CD73, CD90) and serum SDF-1, VEGF-A, EGF, GRO- and GRO-{beta}, GM-CSF and G-CSF were measured by ELISA; group and time effects were evaluated with mixed-effects models and correlations by Spearman tests (two-tailed p<0.05). At admission, trauma patients without HS had significantly higher MSC and HSPC-like populations versus controls (p<0.0001). In the HS cohort SPC percentages rose modestly at day 0-3 then declined sharply by days 7-14 (time effect p<0.0001); non-survivors exhibited significantly higher early SPC and cytokine levels that persisted until death while survivors showed an early rise followed by decline (outcome and time interaction p<0.0001). All cytokines were up-regulated in trauma groups, peaked at day 0-3 in HS patients, and correlated positively with SPC counts (notably SDF-1, VEGF-A, G-CSF, Gro- and GM-CSF; Spearman p<0.05); higher early SPC and cytokine signatures associated with greater organ dysfunction (higher SOFA) and with timing of sepsis. These findings indicate that trauma provokes an early SPC and cytokine response that in HS is followed by later decline, and that persistent early elevation predicts worse outcomes, suggesting serial SPC and cytokine profiling may have prognostic value and identify an early therapeutic window for regenerative or immunomodulatory interventions.

physiology↗

bioRxiv2026

Prime editing of an evolutionarily informed PEPC residue enhances photosynthesis and grain nutrition in C₃ rice

C4 photosynthesis evolved through adaptive changes in key metabolic enzymes that enhance carbon fixation efficiency, yet transferring such traits into C3 crops has met limited success. Using precision prime editing, we introduced an evolutionarily conserved single amino acid substitution (Arg[->]Gly) into phosphoenolpyruvate carboxylase (PEPC) in rice. Edited plants exhibited enhanced PEPC activity in the presence of the feedback inhibitor malate, increased chlorophyll content, elevated photosynthetic rates under ambient CO2, and increased seed size and weight. Remarkably, seeds from the edited lines showed simultaneous enrichment of zinc (Zn), iron (Fe), and protein, hereafter referred to as ZiP rice. This work demonstrates that a single, evolutionarily guided residue change can reprogram core carbon metabolism in a C3 crop, enabling concurrent improvements in photosynthesis and grain nutritional quality.

plant biology↗

bioRxiv2025

CK2α phosphorylates CHIP for oncogenesis: A novel post-translational switch in cancer

The post-translational regulation of tumor suppressors by oncogenic kinases remains a critical yet underexplored determinant of proteostasis reprogramming in cancer. Here, we identified the E3 ubiquitin ligase CHIP (C-terminus of Hsc70-Interacting Protein) as a direct and functionally relevant substrate of the serine/threonine kinase CK2, which is frequently overexpressed in solid tumors. Using LC/MS analysis, in silico kinase prediction, and molecular interaction mapping, we demonstrated that CK2 phosphorylates CHIP at serine 19 (represented as S19), a conserved residue within its TPR domain, thereby promoting ubiquitination and subsequent proteasomal degradation of CHIP. This phosphorylation-dependent destabilization of CHIP impairs its ability to target oncogenic substrates, such as AKT, resulting in sustained AKT phosphorylation and activation, required for oncogenesis. Clinically, we observed a robust inverse correlation between CK2 and CHIP expressions across colorectal and breast cancer patient datasets, which was validated by immunofluorescence (IF) analyses in tumor samples. Multiple functional assays revealed that CK2 suppression, either by genetic ablation or pharmacological inhibition by TBCA, restores CHIP stability, reactivates apoptotic signalling cascades, and attenuates tumor cell proliferation, migration, and 3D spheroid integrity. Moreover, CK2 blockade in syngeneic murine models diminishes primary tumor burden and metastatic dissemination, concomitant with increased CHIP accumulation and reduced AKT signalling. Mechanistically, a phosphorylation-resistant mutant of CHIP at S19 (CHIP-S19A), is refractory to CK2- mediated degradation and preserves anti-tumor functions, delineating a phosphorylation-dependent proteolytic switch as a central node in the CK2-CHIP-AKT regulatory axis. These findings establish a novel paradigm wherein an oncogenic kinase hijacks protein quality control pathway to suppress tumor suppressive activity and facilitate malignancy. Our study positions CK2 as a druggable modulator of CHIP turnover, offering a translational framework for restoring proteostatic checkpoints and constraining oncogenic signalling in aggressive cancers. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=112 SRC="FIGDIR/small/682604v1_ufig1.gif" ALT="Figure 1"> View larger version (37K): org.highwire.dtl.DTLVardef@8fad58org.highwire.dtl.DTLVardef@bd73dforg.highwire.dtl.DTLVardef@2746dorg.highwire.dtl.DTLVardef@129b6ad_HPS_FORMAT_FIGEXP M_FIG C_FIG

cancer biology↗

bioRxiv2025

NANOME: A Nextflow pipeline for haplotype-aware allele-specific consensus DNA methylation detection by nanopore long-read sequencing

Nanopore long-read sequencing has expanded the capacity of long-range, single-base, and single-molecule DNA-methylation (DNAme) detection and haplotype-aware allele-specific epigenetic phasing. Previously, we benchmarked and ranked the robustness of seven computational tools for DNAme detection using nanopore sequencing. The top performers were Megalodon, Nanopolish, DeepSignal and Guppy. However, these algorithms exhibit lower performance at regions with discordant non-singleton DNAme patterns compared to genome-wide regions. Furthermore, long-read sequencing analysis of mammalian genomes requires higher computational resources than next-generation sequencing. To address these issues, we developed a NANOpore Methylation (NANOME) a consensus DNAme predictive model using XGBoost, which integrates the output of Megalodon, Nanopolish, and Deepsignal for analyzing data obtained using Oxford Nanopore Technologies (ONT). NANOME enhanced DNAme detection precision (mean square error) at single-base resolution by 11% and improved accuracy (F1-score) at single-molecule resolution by 2.4% for human B-lymphocyte European cell lines (NA12878). The consensus model also detected [~]200,000 more CpGs than all three tools. Combing variant calling and long-read phasing, NANOME can detect haplotype-aware allele-specific DNAme in known imprinting controls in resolved and previously unresolved regions. We conducted haplotype-aware methylation detection on the T2T genome for dataset NA12878, revealing significant variations in differentially methylated region (DMR) density between gap and non-gap regions. Overall, NANOME represents a significant step forward in DNAme detection and long-range epigenetic phasing, offering a robust and accessible tool for researchers studying the epigenome.

bioinformatics↗

bioRxiv2025

LuxS/AI-2 signaling in Salmonella Typhimurium regulates phoP/phoQ to protect against acidic pH for enhanced survival in macrophages

The intestinal milieu is largely characterized by the complex array of chemical compounds produced through the metabolic activity of resident microbiota. Enteric pathogens like Salmonella, which have evolved refined mechanisms to persist within this environment, utilize these microbial metabolites and self-produce quorum molecules as molecular cues to identify ecological niches and modulate their survival and virulence strategies. Salmonella quorum sensing involves producing and detecting Autoinducer-2 (AI-2) signaling molecules. Our research reveals that Salmonella Typhimurium enhances AI-2 biosynthesis and transport under acidic conditions, aiding environmental adaptation and facilitating pathogenesis in macrophages. AI-2 signaling regulates the pH-sensing two-component system genes, phoP/phoQ, ensuring cytosolic pH homeostasis, survival, and acid tolerance. It also involves regulating the lysine/cadaverine-mediated acid tolerance response and maintaining bacterial cytosolic pH. Furthermore, we show that the repressor LsrR protein, apart from the lsr promoter, binds to only one strand of the phoP promoter via its Y25 and R43 amino acids to negatively modulate phoP expression. Additionally this signaling ameliorates the intracellular survival by modulating Salmonella Pathogenicity Island-2 (SPI-2) regulators (ssrA/ssrB) and SPI-2 effector expression via PhoP. Mouse models demonstrate that AI-2 signaling is essential for colonizing the primary and secondary infection sites, showcasing its critical role in pathogenesis and low pH survival mechanisms in Salmonella.

microbiology↗

bioRxiv2025

Antagonizing the serotonin receptor HTR2B drives antigen-specific cytotoxic T-cell responses and controls colorectal cancer growth.

Neuroimmune communication is known to control various regulatory systems in the body. Serotonin (also known as 5-hydroxytryptamine, HT) is one of the neurotransmitters produced predominantly by enterochromaffin cells in the gut and the neurons of the central nervous system. Immune and cancer cells express several HT receptors (HTRs), and how HTRs modulate various immune pathways and contribute to cancer growth and metastasis is unclear. RNA-seq data analysis shows that human colon adenocarcinoma tissues with high HTR2B expression are associated with poor anti-tumor immune response and poor patient survival. Using an orthotopic mouse model of tumors, we show that HTR2B and other serotonergic molecule expressions are positively associated with colon tumor progression. Antagonizing the HTR2B using specific chemical antagonists significantly suppresses the progression of colon, breast, and melanoma tumors and metastasis in mice by promoting antigen-specific cytotoxic CD8 T cell response and downregulating the PD-L1 expression on tumor cells. Further, treatment with a combination of suboptimal doses of HTR2B antagonists and suboptimal doses of immunotherapy (anti-PD1, anti-PD-L1, or anti-CTLA4 mAb) or chemotherapeutic drugs (5-fluorouracil, oxaliplatin, or irinotecan) showed a robust anti-tumor response and inhibited colon tumor growth. Our finding showed that antagonizing the HTR2B holds a potent therapeutic advantage as a combinatorial regimen of either immunotherapy or chemotherapy to treat colon cancer. One sentence summaryHTR2B expression is associated with colon tumor progression, and antagonizing the HTR2B inhibits tumor growth and metastasis.

immunology↗

bioRxiv2025

Development of Bioisosteric Iboga-alkaloids as Antinociceptive and Anxiolytic Agents with Neuroprotective Effects

The clinical importance of iboga alkaloids lies in their efficacy in reversing drug addiction and modulating drug tolerance. However, due to safety concerns, their use is restricted to appropriate medical supervision. These alkaloids often cause severe hallucinogenic effects due to differential binding to various brain receptors and cardiotoxicity by blocking the human ether-a-go-go-related gene (hERG) potassium channel. To create safer analogs, our group previously synthesized various benzofuran-containing iboga analogs with good opioid binding selectivity and excellent antinociceptive property. However, the present manuscript disclosed a step-economical and cost-effective synthesis of modified ibogaine/ibogamine analogs (C1, C2, C3 & C4) with bio-isosteric replacement of the indole scaffold with a benzofuran moiety, and comparing their antinociceptive/anxiolytic activity with their natural counterparts. Among the synthesized iboga analogs, the Endo-iboga analogs (C2 & C4, epimers of C1 and C3, respectively) not only exhibited superior anti-inflammatory and oxidative stress-relieving activity, but also effectively improved restricted locomotor activity in a formalin-induced acute pain model in mice. These Endo-iboga analogs significantly elevated the levels of inhibitory neurotransmitters (GABA and dopamine) and brain-derived neurotrophic factor (BDNF) compared to their Exo-counterparts or previously published benzofuran-containing iboga analogs lacking the tetrahydroazepine ring. Amongst, C2 and C4, the latter exhibited superior cardiac safety profile in C2C12 cells (IC50 = 235 {micro}M) and showed no adverse effects on rat hearts during in vivo ECG tests, indicated by no significant QTc prolongation. Overall, the development of bioisosteric iboga analogs, particularly C4, demonstrated significant potential for acute pain management without notable cardiotoxicity, representing a breakthrough in pain therapy innovation.

physiology↗

bioRxiv2024

Antimicrobial resistance, pathogenicity and virulence patterns of Escherichia coli isolated from cockroaches (Blattella germanica) across diverse environments: a public health concern

This investigation explores the prevalence and antimicrobial resistance (AMR) of Escherichia coli isolated from Blattella germanica (cockroaches) from diverse locations, including homes, kitchens, and laboratories, over the course of six months, from August 2022 to January 2023. A total of 125 cockroaches were analyzed, yielding 67 (53%) of positive E. coli isolates, with kitchen environments having the highest incidence (57.77%). The existence of virulence genes (stx1, stx2, and rfbO157) was confirmed by pathogenicity assessments carried out on mouse model, which led to a considerable increase in morbidity and mortality. 82.08% of isolates showed evidence of resistance to at least one antibiotic, according to the antimicrobial susceptibility test with {beta}-lactams displaying the highest rates of resistance. Remarkably, complex resistance patterns were observed in 77.61% of the isolates, which were categorized as multidrug-resistant. Multiple antibiotic resistance genes (ARGs) were found by molecular analysis, particularly blaTEM and tetA as well as virulence-associated genes (VAGs) linked to extraintestinal pathogenic E. coli. Phylogenetic grouping indicated that 90.38% of the MDR isolates belonged to virulent groups B2 and D. These findings highlight the role of cockroaches as potential reservoirs for pathogenic E. coli, raising significant public health concerns regarding AMR. The study underscores the imperative need for thorough investigation and feasible control strategies to ease the dissemination of AMR in diverse contexts.

microbiology↗

bioRxiv2024

A Novel Cisplatin-Based Prodrug Inhibits Lysine Deacetylases, Suppresses Nucleotide Excision Repair, and Overcomes Resistance

Cisplatin [cis-diamminedichloroplatinum(II)] is a widely used chemotherapeutic agent that induces cytotoxicity primarily through DNA damage, but drug resistance severely limits its efficacy and use. Cisplatin resistance is complex and multifactorial, involving DNA repair via nucleotide excision repair (NER), and overexpression of lysine deacetylases (KDACs), which reduce chromatin accessibility and alter transcription regulation. The combination of cisplatin and KDAC inhibitors has shown promise in improving treatment efficacy. This improved efficacy has been attributed to increased drug sensitivity due to higher chromatin accessibility, however, this hypothesis has not been validated. In this study, we synthesized a novel Pt(IV) derivative, cct-[Pt(NH3)2Cl2(VPA)(PhB)] (cPVP), which combines cisplatin and two KDAC inhibitors, phenyl butyrate and valproic acid. This triple-action prodrug enabled the simultaneous targeting of multiple cancer-related pathways. Compared to cisplatin, cPVP exhibited significantly enhanced damage formation and cytotoxicity. High-resolution mapping of cisplatin damage and repair, however, does not attribute the enhanced damage sensitivity to chromatin accessibility, but rather to increased drug uptake and inhibition of nucleotide excision repair. Moreover, cPVP treatment increased survival in a mouse mesothelioma model, and prevented the development of resistance to both cisplatin and itself in cancer cells. Our findings shed new light on the effect of KDAC inhibition on cisplatin treatment, and suggest that cPVP could serve as a promising alternative to cisplatin in the clinic.

molecular biology↗

bioRxiv2024

Functional mapping of epigenetic regulators uncovers coordinated tumor suppression by the HBO1 and MLL1 complexes

Epigenetic dysregulation is widespread in cancer. However, the specific epigenetic regulators and the processes they control to drive cancer phenotypes are poorly understood. Here, we employed a novel, scalable and high-throughput in vivo method to perform iterative functional screens of over 250 epigenetic regulatory genes within autochthonous oncogenic KRAS-driven lung tumors. We identified multiple novel epigenetic tumor suppressor and tumor dependency genes. We show that a specific HBO1 complex and the MLL1 complex are among the most impactful tumor suppressive epigenetic regulators in lung. The histone modifications generated by the HBO1 complex are frequently absent or reduced in human lung adenocarcinomas. The HBO1 and MLL1 complexes regulate chromatin accessibility of shared genomic regions, lineage fidelity and the expression of canonical tumor suppressor genes. The HBO1 and MLL1 complexes are epistatic during lung tumorigenesis, and their functional correlation is conserved in human cancer cell lines. Together, these results demonstrate the value of quantitative methods to generate a phenotypic roadmap of epigenetic regulatory genes in tumorigenesis in vivo.

cancer biology↗

bioRxiv2024

Aging represses lung tumorigenesis and alters tumor suppression

Most cancers are diagnosed in persons over the age of sixty, but little is known about how age impacts tumorigenesis. While aging is accompanied by mutation accumulation -- widely understood to contribute to cancer risk -- it is also associated with numerous other cellular and molecular changes likely to impact tumorigenesis. Moreover, cancer incidence decreases in the oldest part of the population, suggesting that very old age may reduce carcinogenesis. Here we show that aging represses tumor initiation and growth in genetically engineered mouse models of human lung cancer. Moreover, aging dampens the impact of inactivating many, but not all, tumor suppressor genes with the impact of inactivating PTEN, a negative regulator of the PI3K/AKT pathway, weakened to a disproportionate extent. Single-cell transcriptomic analysis revealed that neoplastic cells from tumors in old mice retain many age-related transcriptomic changes, showing that age has an enduring impact that persists through oncogenic transformation. Furthermore, the consequences of PTEN inactivation were strikingly age-dependent, with PTEN deficiency reducing signatures of aging in cancer cells and the tumor microenvironment. Our findings suggest that the relationship between age and lung cancer incidence may reflect an integration of the competing effects of driver mutation accumulation and tumor suppressive effects of aging.

cancer biology↗

bioRxiv2024

Modeling the genomic complexity of human cancer using Cas12a mice

Somatic genome editing in mouse models has increased our understanding of the in vivo effects of genetic alterations in areas ranging from neuroscience to cancer biology and beyond. However, existing models are limited in their ability to create multiple targeted edits. Thus, our understanding of the complex genetic interactions that underlie development, homeostasis, and disease remains incomplete. Cas12a is an RNA-guided endonuclease with unique attributes that enable simple targeting of multiple genes with crRNA arrays containing tandem guides. To accelerate and expand the generation of complex genotypes in somatic cells, we generated transgenic mice with Cre-regulated and constitutive expression of enhanced Acidaminococcus sp. Cas12a (enAsCas12a). In these mice, enAsCas12a-mediated somatic genome editing robustly generated compound genotypes, as exemplified by the initiation of diverse cancer types driven by homozygous inactivation of trios of tumor suppressor genes. We further integrated these modular crRNA arrays with clonal barcoding to quantify the size and number of tumors with each array, as well as the efficiency of each crRNA. These Cas12a alleles will enable the rapid generation of disease models and broadly facilitate the high-throughput investigation of coincident genomic alterations in somatic cells in vivo.

cancer biology↗

bioRxiv2024

Curtailed CaSR expression in peripheral blood mononuclear cells is associated with hypertension

The calcium-sensing receptor (CaSR) is a GPCR family member. CaSR is abundantly expressed in the parathyroid gland and controls calcium homeostasis. There are no particular discoveries involving CaSR expression in hypertensive peripheral blood mononuclear cells. We investigated lower CaSR expression in hypertensive human and animal models. Nitric oxide (NO) has an important role in vascular function. Impaired NO production disrupts CaSR-mediated fine adjustment of the intra- and extracellular [Ca2+] gradients. According to the law Ca2+ transportation through Ca channels maintains its connection from the circulatory micro-vascular bed to interstitial space, influenced by capillary and interstitial fluid hydrostatic pressure, as well as oncotic pressure. In that situation, the alterations in Ca2+ microenvironment and dynamics remain unknown. CaSR on circulatory mononuclear cells may detect changes in the Ca2+ microenvironment in the ISF of endothelial cells-VSMC and endothelium lining in hypertension. In this experiment, the decreased expression of CaSR revealed a molecular sensor in the presence of elevated blood pressure.

biochemistry↗

bioRxiv2023

Human coronavirus OC43 infection remodels Connexin 43 mediated gap junction intercellular communication in vitro

{beta}-coronaviruses cause acute infection in the upper respiratory tract, resulting in various symptoms and clinical manifestations. OC43 is a human {beta}-coronavirus that induces mild clinical symptoms and can be safely studied in the BSL2 laboratory. Due to its low risk, OC43 can be a valuable and accessible model for understanding {beta}-coronavirus pathogenesis. One potential target for limiting virus infectivity could be gap junction-mediated communication. This study aims to unveil the status of cell-to-cell communications through gap junctions in human {beta}-coronavirus infection. Infection with OC43 leads to reduced expression of Cx43 in A549, a lung epithelial carcinoma cell line. Infection with this virus also showed a significant ER and oxidative stress increase. Internal localization of Cx43 is observed post OC43 infection in the ERGIC region, which impairs the gap junction communication between two adjacent cells, confirmed by Lucifer yellow dye transfer assay. It also affects hemichannel formation, as depicted by the EtBr uptake assay. Altogether, these results suggest that several physiological changes accompany OC43 infection in A549 cells and can be considered an appropriate model system for understanding the differences in gap junction communication post-viral infections. This model system can provide valuable insights for developing therapies against human {beta}-coronavirus infections. ImportanceThe enduring impact of the recent SARS-CoV-2 pandemic underscores the importance of studying human {beta}-coronaviruses, advancing our preparedness for future coronavirus infections. Due to SARS-CoV-2 being highly infectious, another human {beta}-coronavirus OC43 can be considered as an experimental model. One of the crucial pathways that can be considered is gap junction communication, as it is vital for cellular homeostasis. Our study seeks to understand the change in Cx43-mediated cell-to-cell communication during human {beta}-coronavirus OC43 infection. In vitro studies showed the downregulation of the gap junction protein Cx43 and the upregulation of endoplasmic reticulum and oxidative stress markers post-OC43 infection. Furthermore, OC43 infection causes impairment of functional hemichannel and gap junction formation. Overall, this current study infers that OC43 infection reshapes intercellular communication, suggesting that this pathway may be a promising target for designing highly effective therapeutics against human coronaviruses by regulating Cx43 expression.

pathology↗