Biology subjects

Grover, M.

Publications and source records attributed to Grover, M..

8 recordsLinked to original sources

bioRxiv2026

Maladaptive Piezo1 Mechanotransduction Drives Smooth Muscle Aging in the Gut

Age-related gastrointestinal dysfunction is common, but the mechanisms of aging-associated smooth muscle failure remain unclear. We show that aging in mice slows whole gut and colonic transit, increases regional stiffness, and reduces smooth muscle contractility. Inducible smooth muscle cell (SMC) specific deletion of Piezo1 preserved youthful transit and force generation, whereas Piezo1 activation in young mice phenocopied aging-associated transit delay. Single cell transcriptomics, RNA velocity, stiffness-controlled cell and tissue cultures, and pharmacologic studies revealed that Piezo1 couples increased stiffness to Ca2+, calcineurin, NFAT signaling, loss of contractile gene programs, leading to age-related contractile loss and contractile to synthetic SMC remodeling and gut wall stiffening. Human intestinal SMCs supported conservation of this pathway, and PIEZO1 gain of function carriers showed a trend toward delayed colonic transit. Thus, maladaptive SMC Piezo1 mechanotransduction is a targetable mechanism of aging-associated gut dysmotility.

physiology↗

bioRxiv2026

Validation of a high throughput fluorescent Capillary Electrophoresis Sodium Dodecyl Sulfate method for monoclonal antibody size heterogeneity assessment

Fluorescent capillary gel electrophoresis (CGE) with sodium dodecyl sulfate (CE-SDS) provides a powerful, high-sensitivity alternative to ultraviolet (UV)-based detection for characterizing therapeutic monoclonal antibodies (mAb). Regulatory and standards organizations, such as the United States Pharmacopeia (USP), include only UV based CE-SDS methods, hindering adoption, of alternative detection methods. There is growing opportunity to expand beyond exclusively UV-based CE-SDS methods. In this study, we present a full analytical validation of a light-emitting diode (LED) fluorescence-based parallel CE-SDS method for both non-reduced and reduced analysis of therapeutic antibodies. Using the NISTmAb reference material as a model system, size heterogeneity critical quality attributes (CQAs) including monomeric purity, percent glycosylation, and percent thioether were assessed. The fluorescence method demonstrated high specificity and precision with relative standard deviation (RSD) values <1% for monomeric purity and glycosylation, and <3% for thioether), as well as robust performance across variations in injection voltage, electrophoresis voltage, labeling temperature, and Labeling Buffer concentration. Ruggedness testing across users and reagent lots confirmed reproducibility, and accuracy assessments showed strong agreement with reported values from the National Institute of Standards (NIST) and traditional UV detection measurements. Linearity studies yielded coefficient of determination (R2) values >0.995 for both non-reduced and reduced analyses. These results highlight the high sensitivity, stable baseline performance, and suitability of LED fluorescence-based parallel CE-SDS as a validated, higher-throughput alternative to traditional UV-based methods for mAb quality control (QC).

biochemistry↗

bioRxiv2026

C1qa⁺ muscularis macrophages maintain enteric synaptic homeostasis to regulate gastrointestinal motility

The enteric nervous system (ENS) is a complex peripheral neural network that coordinates gastrointestinal motility through highly organized synaptic communication. Although tissue-resident muscularis macrophages (MMs) closely associate with enteric neurons, whether they regulate enteric synaptic organization remains unknown. In the central nervous system (CNS), microglia sculpt neural circuits through complement-dependent synaptic remodeling, raising the possibility that analogous neuroimmune mechanisms operate in the gut. Here, we identify a previously unrecognized role for C1qa{square} MMs in regulating enteric synaptic homeostasis and gastrointestinal motility. Using macrophage-specific constitutive and inducible C1qa deletion models, single-cell RNA sequencing, enteric synaptosome proteomics, physiology, and advanced imaging, we demonstrate that loss of MMs-derived C1qa increases enteric synaptic density without altering neuronal numbers. C1qa deficiency induced broad transcriptional changes in enteric neurons and macrophages, including altered synapse-associated, lysosomal, and endocytic programs. Proteomic analysis revealed that enteric synapses share a conserved molecular architecture with brain synapses while exhibiting distinct gastrointestinal-specific complement-associated synaptic networks enriched for structural and receptor-localization pathways. Functionally, macrophage-specific C1qa deletion altered excitatory and inhibitory enteric neurotransmission, enhanced cholinergic signaling, reduced nitrergic responses, and accelerated gastrointestinal transit, while smooth muscle responsiveness remained preserved. C1qa{square} MMs displayed transcriptional and functional features consistent with a phagocytic synapse-remodeling phenotype, including enrichment of complement, lysosomal, and engulfment pathways. Loss of C1qa impaired macrophage phagocytic activity both in vitro and in vivo and was associated with synapse accumulation and altered macrophage morphology. Importantly, inducible deletion of C1qa in adulthood recapitulated the synaptic and motility phenotypes, demonstrating that C1qa{square} MMs continuously regulate enteric synaptic organization beyond development. Together, these findings identify a complement-dependent neuroimmune mechanism that regulates enteric circuit organization and gut motility, establishing MMs as active modulators of adult ENS synaptic homeostasis. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=117 SRC="FIGDIR/small/729640v1_ufig1.gif" ALT="Figure 1"> View larger version (52K): org.highwire.dtl.DTLVardef@16aadd8org.highwire.dtl.DTLVardef@bb74f3org.highwire.dtl.DTLVardef@fb040corg.highwire.dtl.DTLVardef@1c6e4a5_HPS_FORMAT_FIGEXP M_FIG C_FIG

neuroscience↗

bioRxiv2025

Intestinal Stem Cells Retain an Epigenetic Memory of Prior Inflammation

Intestinal epithelial damage and impaired repair are hallmarks of ulcerative colitis (UC), even after inflammation resolves. Intestinal stem cells (ISCs) can retain stable epigenetic changes after inflammation, highlighting the potential for long-lived epithelial memory in the gut. Inflammatory injury in barrier tissues induces epigenetic memory in epithelial stem cells, and the tendency of UC to relapse at previously inflamed sites led us to hypothesize that ISCs from IBD patients acquire lasting memory of prior inflammation. To test this, we derived colonic organoids from inflamed and uninflamed regions of the same UC patients and propagated in long-term culture. Chromatin profiling revealed 2,252 accessible regions unique to prior-inflamed (PI) organoids, associated with stress response, repair, and inflammatory genes. Although these regions remained accessible, [~]95% of associated genes were not upregulated in PI organoids, indicating a primed state. Upon inflammatory or injury re-challenge, PI organoids exhibited heightened transcriptional responses and accelerated wound closure, despite reduced clonogenicity and impaired barrier function, indicating a retained inflammatory memory program. Our findings demonstrate that human ISCs retain a chromatin-based memory of inflammation that persists in the absence of immune cues and shapes future responses to injury. While this may support epithelial adaptation to secondary insults, it may predispose tissue to relapse in patients with UC.

molecular biology↗

bioRxiv2024

Paired C-type lectin receptors mediate specific recognition of divergent oomycete pathogens in C. elegans.

Innate immune responses can be initiated through the detection of pathogen or damage-associated molecular patterns by host receptors that are often present on the surface of immune cells. While certain invertebrates like Caenorhabditis elegans lack professional immune cells, they still respond to infection in a pathogen-specific manner. It has been debated for years whether homologues of the canonical pathogen recognition receptors are also functioning in the nematode. Here we show that C-type lectin receptors mediate species-specific recognition of divergent oomycetes in C. elegans. A CLEC-27/CLEC-35 pair is essential for recognition of the oomycete Myzocytiopsis humicola, while a CLEC-26/CLEC-36 pair is required for detection of Haptoglossa zoospora. Both clec pairs are transcriptionally regulated through a shared promoter by the conserved PRD-like homeodomain transcription factor CEH-37/OTX2 and act in sensory neurons and the anterior intestine to trigger a protective immune response in the epidermis. This system enables redundant tissue sensing of oomycete threats through canonical CLEC receptors and host defense via cross-tissue communication. HighlightsO_LIA CLEC-27/CLEC-35 pair is required for recognition of the oomycete Myzocytiopsis humicola C_LIO_LIA CLEC-26/CLEC-36 pair is required for recognition of the oomycete Haptoglossa zoospora C_LIO_LIBoth CLEC pairs are co-regulated by the homeodomain transcription factor CEH-37/OTX2 C_LIO_LIBoth CLEC pairs function redundantly in sensory neurons and the intestine for host defense C_LI

immunology↗

bioRxiv2023

Impairment of the SKN-1A/NRF1 proteasome surveillance pathway triggers tissue-specific protective immune responses against distinct natural pathogens in C. elegans.

Protein quality control pathways play important roles in resistance against pathogen infection. For example, the conserved transcription factor SKN-1/NRF upregulates proteostasis capacity after blockade of the proteasome, and also promotes resistance against bacterial infection in the nematode C. elegans. SKN-1/NRF has three isoforms, and the SKN-1A/NRF1 isoform in particular regulates proteasomal gene expression upon proteasome dysfunction as part of a conserved bounce-back response. We report here that, in contrast to the previously reported role of SKN-1 in promoting resistance against bacterial infection, loss-of-function mutants in skn-1a and its activating enzymes ddi-1 and png-1, show constitutive expression of immune response programmes against natural eukaryotic pathogens of C. elegans. These programmes are the Oomycete Recognition Response (ORR), which promotes resistance against oomycetes that infect through the epidermis, and the Intracellular Pathogen Response (IPR), which promotes resistance against intestine-infecting microsporidia. Consequently, skn-1a mutants show increased resistance to both oomycete and microsporidia infections. We also report that almost all ORR/IPR genes induced in common between these programmes are regulated by the proteasome and interestingly, specific ORR/IPR genes can be induced in distinct tissues depending on the exact trigger. Furthermore, we show that increasing proteasome function significantly reduces oomycete-mediated induction of multiple ORR markers. Altogether, our findings demonstrate that proteasome regulation keeps innate immune responses in check in a tissue-specific manner, against natural eukaryotic pathogens of the C. elegans epidermis and intestine.

immunology↗

bioRxiv2022

A receptor tyrosine kinase regulated by the transcription factor VAB-3/PAX6 pairs with a pseudokinase to trigger immune signalling upon oomycete recognition in C. elegans

Oomycetes were recently discovered as natural pathogens of Caenorhabditis elegans and pathogen recognition alone was shown to be sufficient to activate a protective transcriptional program in the host characterised by the expression of multiple chitinase-like (chil) genes. However, the molecular mechanisms underlying oomycete recognition in animals remain fully unknown. We performed here a forward genetic screen to uncover regulators of chil gene induction and found several independent loss-of-function alleles of old-1 and flor-1, which encode receptor tyrosine kinases belonging to the C. elegans-specific KIN-16 family. We present evidence that OLD-1 is an active kinase mounting the immune response, and FLOR-1 a pseudokinase that is also required for the response and regulates the distribution of OLD-1 at the epidermal membrane. Interestingly, the old-1 locus is adjacent to the chil genes in the nematode genome, thereby revealing a genetic cluster important for oomycete-resistance. Furthermore, we identify the VAB-3/PAX-6 transcription factor known for its role in visual system development to regulate old-1 expression, and consequently the spatial pattern of the response to oomycete recognition. Taken together, our study reveals both conserved and species-specific factors shaping the response to oomycete recognition.

immunology↗

bioRxiv2022

A pals-25 gain-of-function allele triggers systemic resistance against natural pathogens of C. elegans

Regulation of immunity throughout an organism is critical for host defense. Previous studies in the nematode Caenorhabditis elegans have described an "ON/OFF" immune switch comprised of the antagonistic paralogs PALS-25 and PALS-22, which regulate resistance against intestinal and epidermal pathogens. Here, we identify and characterize a PALS-25 gain-of-function mutant protein with a premature stop (Q293*), which we find is freed from physical repression by its negative regulator, the PALS-22 protein. PALS-25(Q293*) activates two related gene expression programs, the Oomycete Recognition Response (ORR) against natural pathogens of the epidermis, and the Intracellular Pathogen Response (IPR) against natural intracellular pathogens of the intestine. A subset of ORR/IPR genes is upregulated in pals-25(Q293*) mutants, and they are resistant to oomycete infection in the epidermis, and microsporidia and virus infection in the intestine, but without compromising growth. Surprisingly, we find that activation of PALS-25 seems to primarily stimulate the downstream bZIP transcription factor ZIP-1 in the epidermis, which leads to upregulation of gene expression in both the epidermis and in the intestine. Interestingly, we find that this epidermal-to-intestinal signaling promotes resistance to the N. parisii intestinal pathogen, demonstrating cross-tissue protective immune induction from one epithelial tissue to another in C. elegans. Author summaryMulticellular organisms need to monitor the health and function of multiple tissues simultaneously to respond appropriately to pathogen infection. Here, we study an ON/OFF switch in the roundworm C. elegans that controls immune responses to diverse natural pathogens of the skin and gut. We show a physical association between the ON switch protein PALS-25 and the OFF switch protein PALS-22, and that this association is disrupted in a mutant, activated form of PALS-25. When either PALS-22 is lost, or PALS-25 is activated, a downstream immune regulator ZIP-1 is activated specifically in the skin but not the gut. Excitingly, our findings show that skin-specific loss of PALS-22 or skin-specific activation of PALS-25 can induce immune responses in the worm gut. These findings highlight the coordination of immune responses across different tissues that are commonly infected by microbial pathogens.

immunology↗