{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,10,16]],"date-time":"2026-10-16T06:21:12Z","timestamp":1792131672859,"version":"4.4.0"},"reference-count":76,"publisher":"Wiley","issue":"15","license":[{"start":{"date-parts":[[2024,11,4]],"date-time":"2024-11-04T00:00:00Z","timestamp":1730678400000},"content-version":"vor","delay-in-days":0,"URL":"http:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"content-domain":{"domain":["physoc.onlinelibrary.wiley.com"],"crossmark-restriction":true},"short-container-title":["The Journal of Physiology"],"published-print":{"date-parts":[[2025,8]]},"abstract":"<jats:sec><jats:title>Abstract<\/jats:title><jats:p>Volume\u2010regulated anion channels (VRACs) formed by leucin\u2010rich repeat containing 8 (LRRC8) proteins play a pivotal role in regulatory volume decrease by mediating the release of chloride and organic osmolytes. Apart from the regulation of cell volume, LRRC8\/VRAC function underlies numerous physiological processes in vertebrate cells including membrane potential regulation, glutamate release and apoptosis. LRRC8\/VRACs are also permeable to antibiotics and anti\u2010cancer drugs, representing therefore important therapeutic targets. The activation mechanisms for LRRC8\/VRACs are still unclear. Besides through osmotic cell swelling, LRRC8\/VRACs can be activated by various stimuli under isovolumetric conditions. Sphingosine\u20101\u2010phosphate (S1P), an important signalling lipid, which signals through a family of G protein\u2010coupled receptors (GPCRs), has been reported to activate LRRC8\/VRACs in several cell lines. Here, we measured inter\u2010subunit F\u00f6rster resonance energy transfer (FRET) and used whole\u2010cell patch clamp electrophysiology to investigate S1P\u2010induced LRRC8\/VRAC activation. We systematically assessed the involvement of GPCRs and G protein\u2010mediated signal transduction in channel activation. We found that S1P\u2010induced channel activation is mediated by S1PR1 in HeLa cells. Following the downstream signalling pathway of S1PR1 and using toxin\u2010mediated inhibition of the associated G proteins, we showed that G\u03b2\u03b3 dimers rather than G\u03b1i or G\u03b1q play a critical role in S1P\u2010induced VRAC activation. We could also show that S1P causes protein kinase D (PKD) phosphorylation, suggesting that G\u03b2\u03b3 recruits phospholipase C\u03b2 (PLC\u03b2) with the consequent PKD activation by diacylglycerol. Notably, S1P did not activate LRRC8\/VRAC in HEK293 cells, but overexpression of G\u03b2\u03b3\u2010responsive PLC\u03b2 isoform could facilitate S1P\u2010induced LRRC8\/VRAC currents. We thus identified S1PR1\u2010mediated G\u03b2\u03b3\u2010PLC\u03b2 signalling as a key mechanism underlying isosmotic LRRC8\/VRAC activation.\n<jats:boxed-text content-type=\"graphic\" position=\"anchor\"><jats:graphic xmlns:xlink=\"http:\/\/www.w3.org\/1999\/xlink\" mimetype=\"image\/png\" position=\"anchor\" specific-use=\"enlarged-web-image\" xlink:href=\"graphic\/tjp16402-gra-0001-m.png\"><jats:alt-text>image<\/jats:alt-text><\/jats:graphic><\/jats:boxed-text><\/jats:p><\/jats:sec><jats:sec><jats:title>Key points<\/jats:title><jats:p><jats:list list-type=\"bullet\">\n<jats:list-item><jats:p>Leucin\u2010rich repeat containing 8 (LRRC8) anion\/osmolyte channels are involved in multiple physiological processes where they can be activated as volume\u2010regulated anion channels (VRACs) by osmotic cell swelling or isovolumetric stimuli such as sphingosine\u20101\u2010phosphate (S1P).<\/jats:p><\/jats:list-item>\n<jats:list-item><jats:p>In the present study, using pharmacological modulation and gene\u2010depleted cells in patch clamp recording and optical monitoring of LRRC8 activity, we find that LRRC8\/VRAC activation by S1P is mediated by the G protein\u2010coupled receptor S1PR1 coupled to G proteins of the Gi family.<\/jats:p><\/jats:list-item>\n<jats:list-item><jats:p>The signal transduction to LRRC8\/VRAC activation specifically involves phospholipase C\u03b2 activation by \u03b2\u03b3 subunits of pertussis toxin\u2010insensitive heteromeric Gi proteins.<\/jats:p><\/jats:list-item>\n<jats:list-item><jats:p>S1P\u2010mediated and hypotonicity\u2010induced LRRC8\/VRAC activation pathways converge in protein kinase D activation.<\/jats:p><\/jats:list-item>\n<\/jats:list><\/jats:p><\/jats:sec>","DOI":"10.1113\/jp286665","type":"journal-article","created":{"date-parts":[[2024,11,4]],"date-time":"2024-11-04T21:11:24Z","timestamp":1730754684000},"page":"4329-4344","update-policy":"https:\/\/doi.org\/10.1002\/crossmark_policy","source":"Crossref","is-referenced-by-count":6,"title":["Sphingosine\u20101\u2010phosphate activates LRRC8 volume\u2010regulated anion channels through G\u03b2\u03b3 signalling"],"prefix":"10.1113","volume":"603","author":[{"ORCID":"https:\/\/orcid.org\/0000-0001-9933-2918","authenticated-orcid":false,"given":"Yulia","family":"Kostritskaia","sequence":"first","affiliation":[{"name":"Institute for Molecular Medicine MSH Medical School Hamburg  Hamburg 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