Biology subjects

Sarkar, M. K.

Publications and source records attributed to Sarkar, M. K..

9 recordsLinked to original sources

bioRxiv2026

IFN-κ directs antiviral immunity in human skin

Inducible expression of type I IFNs is a well-established host innate defense mechanism to limit virus infection. Yet, many viruses have evolved strategies to suppress the induction of these cytokines to enhance replication, spread, and transmission between hosts. Whether additional antiviral mechanisms protect against infection when inducible responses are compromised is not well understood. Here, we demonstrate that human keratinocytes preemptively protect the skin against virus infection through constitutive production of IFN-kappa (IFN-{kappa}), a poorly studied type I IFN family member. We find that constitutive IFNK expression protects keratinocytes against skin tropic RNA and DNA viruses, including vesicular stomatitis virus (VSV) and herpes simplex virus-1 (HSV-1). Using a human skin organoid model, we further demonstrate that keratinocyte-derived IFN-{kappa} establishes an antiviral state in dermal fibroblasts. Genetic and chemical analysis of the type I IFN receptor (IFNAR) signaling pathway in monocultured keratinocytes and skin organoids revealed that distinct pathways control VSV and HSV-1 replication. While canonical JAK-STAT signaling provided protection against VSV, HSV-1 infection was controlled through a JAK-STAT-independent mechanism. Using transcriptomic analysis, we further identified an IFN-{kappa}-dependent gene signature in keratinocytes that is not similarly driven by other type I IFNs. Together, this work establishes constitutive IFN-{kappa} production by keratinocytes as a broadly antiviral tissue autonomous defense mechanism.

immunology↗

bioRxiv2026

Integrated in silico identification of fungal-derived dual-target inhibitors for anti-schistosomal drug discovery

Praziquantel, which has limited efficacy against young parasites and reduces susceptibility, is the principal therapy for schistosomiasis, a neglected tropical illness. This work identified fungal-derived compounds with potential dual inhibitory action against Schistosoma mansoni DHODH and cathepsin B1 using an integrated computational method. From 1,831 fungal metabolites, 120 were selected for molecular docking after structure standardisation, drug-likeness evaluation, ADMET, and toxicity screening. Ganoderlactone B (CID_122184973) and Spiroapplanatumine F (CID_132962217) strongly bound to DHODH and CB1, with docking affinities of -10.6 and -10.0 kcal/mol and -9.0 and -8.9, respectively. Docking validation showed good discrimination between active compounds and decoys with area-under-the-curve values of 0.98 for DHODH and 0.96 for CB1. The protein-ligand interaction study showed hydrogen bonding, hydrophobic interactions, and van der Waals contacts at the binding sites. In 100 ns molecular dynamics simulations, protein complexes exhibited modest backbone deviations and maintained compactness, although CID_132962217 showed sustained binding and a constrained conformation. MM-PBSA analysis identified CID_122184973 as the best common ligand for both targets. At the B3LYP/6-31G level, density functional theory calculations showed that CID_139584993 had the lowest HOMO-LUMO energy gap and hardness values of 3.552 and 1.776 eV, respectively, and the highest softness value of 0.282 eV-1, indicating greater electronic reactivity. Electrostatic potential mapping and QSAR predictions supported their interaction and antiparasitic potential. Overall, CID_122184973 and CID_132962217 are promising fungal scaffolds for dual-target anti-schistosomal drug development. Experimental enzyme inhibition, parasite viability, toxicity, and in vivo research are needed to confirm computational findings.

bioinformatics↗

bioRxiv2026

Hailey-Hailey disease models identify synergistic therapeutic effects of MEK and ROCK inhibition

Hailey-Hailey disease (HHD) is a genetic skin blistering disorder lacking approved treatments despite linkage to ATP2C1 variants 25 years ago. Since knockout mice did not replicate HHD, we ablated ATP2C1 in human keratinocytes or chemically inhibited its encoded Golgi calcium pump SPCA1. In organotypic epidermis, SPCA1 deficiency or inhibition reproduced HHD pathology, disrupting desmosomal cadherins and severing cell-cell junctions, termed acantholysis. RNA sequencing of heterozygous cells identified dysregulation of actin and Rho GTPases along with EGF receptor signaling as potential pathogenic drivers. Accordingly, SPCA1-depleted organotypic epidermis and HHD biopsies exhibited cortical actin disorganization and hyper-phosphorylation of the Rho kinase (ROCK) target, myosin light chain. Rho activation was sufficient to induce acantholysis, while ROCK inhibition partially restored heterozygous keratinocyte cohesion. A fluorescent biosensor demonstrated ERK hyper-activation in heterozygous cells along with desmosomal cadherin mis-localization. Importantly, treating SPCA1-deficient keratinocyte sheets with MEK and ROCK inhibitors together fully restored their integrity. Our results show HHD blistering is driven by desmosome and cortical actin dysfunction that was mitigated by targeting MEK and ROCK with repurposed drugs, offering a viable treatment strategy. Moreover, our model provides a blueprint for replicating genetic epidermal disorders to delineate pathogenic mechanisms and vet therapeutics for other orphan skin diseases. GRAPHICAL ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=199 SRC="FIGDIR/small/726679v1_ufig1.gif" ALT="Figure 1"> View larger version (57K): org.highwire.dtl.DTLVardef@b2de6aorg.highwire.dtl.DTLVardef@128411borg.highwire.dtl.DTLVardef@1ca760forg.highwire.dtl.DTLVardef@10cd6c7_HPS_FORMAT_FIGEXP M_FIG C_FIG

cell biology↗

bioRxiv2025

Insulin-Regulated Actin Dynamics is Disrupted in a Human Keratinocyte Model of Hailey Hailey Disease

Hailey Hailey Disease (HHD) is an autosomal dominant cutaneous disorder caused by mutations in ATP2C1, the gene encoding the Golgi/secretory pathway Ca2+-ATPase SPCA1. Characterized by suprabasal acantholysis and intertriginous blistering of the skin, HHD treatment focuses on managing symptoms as there is no cure. Challenges to targeted therapy are due to the lack of facile and reliable models, both human and rodent, for mechanistic studies. Here we validate and characterize CRISPR/Cas9 mediated single and bi-allelic ATP2C1 knockouts in immortalized human hTERT keratinocytes. Whereas SPCA1 expression, Golgi morphology and Golgi Ca2+ accumulation were proportionately affected in heterozygous and homozygous ATP2C1 null mutants as expected, both single and double allelic mutants showed near complete loss of cadherins associated with desmosomal and adherens junctions. HHD is characterized by poor wound healing and impaired keratinocyte migration. We show that SPCA1 is required for dynamic reorganization of actin cytoskeleton in keratinocyte spreading. We identified an insulin activated PI3K-AKT-Rac1 signaling pathway required for lamellipodia formation and keratinocyte spreading, defective in SPCA1 mutants. Transgenic expression of hSPCA1 or treatment with CDN1163, a small molecule Ca2+-ATPase agonist, restored defective phenotypes in the HHD model, paving the way for future therapeutic approaches to treat this disorder.

cell biology↗

bioRxiv2025

The endoplasmic reticulum autophagy receptor TEX264 drives epidermal differentiation and is dysregulated in Darier disease

Differentiating keratinocytes break down their organelles and nuclei to become the compacted cornified layers of the epidermal barrier in a poorly understood catabolic process. Live confocal imaging of stratified human organotypic epidermis revealed endoplasmic reticulum (ER) fragmentation and lysosomal engulfment in the cornifying layers, where we found up-regulation of TEX264, a receptor that mediates selective autophagy of the ER (reticulophagy). TEX264 expression was increased by ER stress, which caused precocious cornification of organotypic epidermis. In undifferentiated keratinocytes, ectopic TEX264 was sufficient to fragment the ER, while in highly differentiated keratinocytes, it accelerated ER elimination and induced nuclear shrinkage; these effects were abolished by mutating the LC3 interacting region required for its autophagic function. Knockout of TEX264 or inhibiting its activation disrupted maturation of organotypic cultures, pointing to a critical role for reticulophagy in cornification. Finally, in patient biopsies and an organotypic model of Darier disease, a genetic cornification disorder linked to ER dysfunction, we found increased TEX264 in areas of premature cornification (dyskeratosis). Our results identified TEX264 as a key driver of epidermal differentiation and led us to propose a novel model of cornification in which keratinocytes activate selective autophagy receptors to orchestrate orderly organelle elimination during cutaneous barrier formation. GRAPHICAL ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=66 SRC="FIGDIR/small/668774v1_ufig1.gif" ALT="Figure 1"> View larger version (28K): org.highwire.dtl.DTLVardef@46f7a6org.highwire.dtl.DTLVardef@12cd441org.highwire.dtl.DTLVardef@23327aorg.highwire.dtl.DTLVardef@c6e9b5_HPS_FORMAT_FIGEXP M_FIG C_FIG

cell biology↗

bioRxiv2025

VGLL3-centered network connects placental, vascular, and immune defects in preeclampsia

Preeclampsia affects approximately 1 in 10 pregnancies, leading to severe complications and long-term health risks for both mother and offspring. While the etiology remains unclear, preeclampsia has been linked to both autoimmunity and the timing of menarche. Through human single-cell and spatial analyses, coupled with in vitro, in vivo, and ex vivo models, we demonstrate that VGLL3, a transcription co-regulator in the Hippo pathway, is upregulated in preeclamptic placentas. VGLL3 promotes immune activation, impairs trophoblast differentiation, and induces endothelial dysfunction, all of which contribute to pregnancy-related hypertension, fetal growth restriction, and offspring mortality. Our data reveal that VGLL3 acts upstream of preeclampsia-associated processes, including the production of sFLT1, a key biomarker of the disease. Notably, targeting VGLL3--either by genetic deletion in mouse placentas or through therapeutic inhibition in human placentas--protects against preeclampsia and alleviates disease pathology. These findings position VGLL3 as a promising novel therapeutic target for preeclampsia.

immunology↗

bioRxiv2025

A Transcriptomic Atlas of Healthy Human Skin Links Regional Identity to Inflammatory Disease.

Human skin is not a uniform organ but a mosaic of anatomically distinct niches, with each site finely tuned to unique environmental demands and immune pressures. Yet, the molecular determinants that define these regional identities and their relationship to site-specific vulnerability to inflammatory disease remain poorly understood. Here, we generate a high-resolution single-cell atlas of human skin, profiling 274,834 cells from 96 healthy samples across 7 anatomically distinct sites (acral, arm, axilla, back, face, leg and scalp). Our analysis reveals striking region-specific transcriptional and cellular networks, uncovering how local immune-stromal crosstalk governs tissue homeostasis and underpins anatomical susceptibility to distinct inflammatory diseases such as such as systemic lupus erythematosus (SLE), atopic dermatitis (AD), and psoriasis. These findings illuminate the tissue-intrinsic foundations of regional immune identity and provide a blueprint/resource for the development of precision therapies tailored to the distinct immunological microenvironments of specific anatomical skin sites.

systems biology↗

bioRxiv2024

Keratin 16 spatially inhibits type I interferon responses in stressed skin

The stress-induced keratin 16 is broadly used as a biomarker in inflammatory skin disorders while pathogenic variants in KRT16 cause pachyonychia congenita (PC), a condition in which differentiation and homeostasis are disrupted in palmoplantar epidermis and epithelial appendages. How K16 impacts these disorders at a molecular level is poorly understood. Here we report that K16 spatially restricts type I interferon (IFN) signaling and innate immunity in palmoplantar keratoderma (PPK) lesions in PC patients, imiquimod- and phorbol ester-induced models of sterile inflammation in mouse skin, and poly(I:C)-treated human keratinocytes ex vivo. Mechanistically, K16 interacts with effectors of the RIG-I-like receptor (RLR) pathway, including 14-3-3{varepsilon}, and inhibits the 14-3-3{varepsilon}:RIG-I interaction upstream of IFN activation. Topical application of the JAK inhibitor Ruxolitinib reduces the severity of PC-PPK-like lesions in Krt16 null mice. These findings uncover a new paradigm for keratin-dependent regulation of innate immunity and suggest a new approach to PC treatment. One sentence summaryKRT16 negatively regulates type I interferon signaling and innate immune responses in the skin, offering insight into the pathophysiology of inflammatory skin diseases including pachyonychia congenita, psoriasis and others.

immunology↗

bioRxiv2023

Targeting SERCA2 in organotypic epidermis reveals MEK inhibition as a therapeutic strategy for Darier disease.

Mutation of the ATP2A2 gene encoding sarco-endoplasmic reticulum calcium ATPase 2 (SERCA2) was linked to Darier disease more than two decades ago; however, there remain no targeted therapies for this disorder causing recurrent skin blistering and infections. Since Atp2a2 knockout mice do not phenocopy its pathology, we established a human tissue model of Darier disease to elucidate its pathogenesis and identify potential therapies. Leveraging CRISPR/Cas9, we generated human keratinocytes lacking SERCA2, which replicated features of Darier disease, including weakened intercellular adhesion and defective differentiation in organotypic epidermis. To identify pathogenic drivers downstream of SERCA2 depletion, we performed RNA sequencing and proteomic analysis. SERCA2-deficient keratinocytes lacked desmosomal and cytoskeletal proteins required for epidermal integrity and exhibited excess MAP kinase signaling, which modulates keratinocyte adhesion and differentiation. Immunostaining patient biopsies substantiated these findings with lesions showing keratin deficiency, cadherin mis-localization, and ERK hyper-phosphorylation. Dampening ERK activity with MEK inhibitors rescued adhesive protein expression and restored keratinocyte sheet integrity despite SERCA2 depletion or chemical inhibition. In sum, coupling multi-omic analysis with human organotypic epidermis as a pre-clinical model, we found that SERCA2 haploinsufficiency disrupts critical adhesive components in keratinocytes via ERK signaling and identified MEK inhibition as a treatment strategy for Darier disease.

cell biology↗