Sala-Rabanal, M. | bioRxiv Science

bioRxiv2020

Goblet cell LRRC26 regulates BK channel activation and protects against colitis in mice

Goblet cells (GCs) are specialized cells of the intestinal epithelium contributing critically to mucosal homeostasis. One of the functions of GCs is to produce and secrete MUC2, the mucin that forms the scaffold of the intestinal mucus layer coating the epithelium and separates the luminal pathogens and commensal microbiota from the host tissues. Although a variety of ion channels and transporters are thought to impact on MUC2 secretion, the specific cellular mechanisms that regulate GC function remain incompletely understood. Previously, we demonstrated that leucine-rich-repeat-containing protein 26 (LRRC26), a known regulatory subunit of the Ca2+-and voltage-activated K+ channel (BK channel), localizes specifically to secretory cells within the intestinal tract. Here, utilizing a mouse model in which MUC2 is fluorescently tagged allowing visualization of single GCs in intact colonic crypts, we show that murine colonic GCs have functional LRRC26-associated BK channels. In the absence of LRRC26, BK channels are present in GCs, but are not activated at physiological conditions. In contrast, all tested MUC2-negative cells completely lacked BK channels. Moreover, LRRC26-associated BK channels underlie the BK channel contribution to the resting transepithelial current across mouse distal colonic mucosa. Genetic ablation of either LRRC26 or BK-pore forming -subunit in mice results in a dramatically enhanced susceptibility to colitis induced by dextran sodium sulfate (DSS). These results demonstrate that normal potassium flux through LRRC26-associated BK channels in GCs has protective effects against colitis in mice. SignificanceA primary function of goblet cells (GCs) of the intestinal epithelium is to generate a protective mucus layer lining the intestinal lumen. GC dysfunction is linked to Inflammatory Bowel Disease (IBD). GC mucus secretion is thought to be dependent on contributions of an ensemble of anion and cation fluxes, although understanding remains limited. Here, it is shown in mouse colon that the Ca2+- and voltage-dependent BK-type K+ channel, specifically in association with the LRRC26 regulatory subunit, plays a critical role in normal GC function, protecting mice against chemically-induced colitis. The results demonstrate that normal K+ fluxes mediated by LRRC26-containing BK channels are required for normal GC function, potentially providing insights into the potential role of BK channels in IBD.

physiology↗

bioRxiv2017

Cantu syndrome-associated SUR2 (ABCC9) mutations in distinct structural domains result in KATP channel gain-of-function by differential mechanisms

The complex cardiovascular disorder Cantu Syndrome arises from gain-of-function mutations in either KCNJ8 or ABCC9, the genes encoding the Kir6.1 and SUR2 subunits of ATP-sensitive potassium (KATP) channels. Recent reports indicate that such mutations can increase channel activity by multiple molecular mechanisms. In this study, we determine the mechanism by which KATP function is altered by several mutations in distinct structural domains of SUR2: D207E in the intracellular L0-linker and Y985S, G989E, M1060I, and R1154Q/W in TMD2. Mutations were engineered at their equivalent position in rat SUR2A (D207E, Y981S, G985E, M1056I and R1150Q/W) and functional effects were investigated using macroscopic rubidium (86Rb+) efflux assays and patch clamp electrophysiology. The results show that D207E increases KATP activity by increasing intrinsic stability of the open state, whilst the cluster of Y981S/G985E/M1056I mutations, as well as R1150Q/W, augment Mg-nucleotide activation. The response of mutant channels to inhibition by the sulfonylurea drug glibenclamide, a potential pharmacotherapy for CS, was also tested. There was no major effect on glibenclamide sensitivity for the D207E, Y981S, G985E or M1056I mutations, but glutamine and tryptophan substitution at R1150 resulted in significant decreases in potency.

biophysics↗