Oxidation driven reversal of PIP2-dependent gating in GIRK2 channels

Oxidation driven reversal of PIP2-dependent gating in GIRK2 channels | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Article  Oxidation driven reversal of PIP 2 -dependent gating in GIRK2 channels Sun-Joo Lee, Colin Nichols This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-1792750/v1 This work is licensed under a CC BY 4.0 License Status: Posted Version 1 posted You are reading this latest preprint version Abstract Physiological activity of G protein gated inward rectifier K+ (GIRK, Kir3) channel activity is dynamically regulated by three key ligands, phosphoinositol-4,5-bisphosphate (PIP2), Gβγ, and Na+, which underlies cellular electrical response to multiple hormones and neurotransmitters in myocytes and neurons. In a reducing environment, matching that inside cells, purified GIRK2 (Kir3.2) channels demonstrated low basal activity, and appropriate sensitivity to the above ligands. However, under oxidizing conditions, anomalous behavior emerged, including rapid loss of agonist dependent activation, and high basal activity in the absence of any agonists that was paradoxically inhibited by PIP2. Mutagenesis identified two cysteine residues (C65 and C190) as being responsible for the loss of agonist dependent activation and the elevated basal activity, respectively. The results explain anomalous findings from earlier studies and illustrate the potential pathophysiologic consequences of oxidation on GIRK channel function, as well as providing insight to reversed ligand-dependence of gating in related Kir and KirBac channels. GIRK2 Kir3 PIP2 oxidation oxidative stress reversed gating Full Text Additional Declarations There is NO Competing Interest. Supplementary Files supplementary.pdf Cite Share Download PDF Status: Posted Version 1 posted You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. As a division of Research Square Company, we’re committed to making research communication faster, fairer, and more useful. We do this by developing innovative software and high quality services for the global research community. Our growing team is made up of researchers and industry professionals working together to solve the most critical problems facing scientific publishing. Also discoverable on Platform About Our Team In Review Editorial Policies Advisory Board Help Center Resources Author Services Accessibility API Access RSS feed Manage Cookie Preferences © Research Square 2026 | ISSN 2693-5015 (online) Privacy Policy Terms of Service Do Not Sell My Personal Information {"props":{"pageProps":{"initialData":{"identity":"rs-1792750","acceptedTermsAndConditions":true,"allowDirectSubmit":true,"archivedVersions":[],"articleType":"Article","associatedPublications":[],"authors":[{"id":119372999,"identity":"7585d552-7bda-4d4a-bcd3-e9ff17d02fb1","order_by":0,"name":"Sun-Joo Lee","email":"data:image/png;base64,iVBORw0KGgoAAAANSUhEUgAAAZAAAAAyAQMAAABI0h/eAAAABlBMVEX///8AAABVwtN+AAAACXBIWXMAAA7EAAAOxAGVKw4bAAAA5ElEQVRIiWNgGAWjYJACAyCWYWBgPsAAYYAFCGvhYWBgSwCxeYjSwgDRwmNAnBb5GbkHCj5UMPDws/d8k+b584eHgb15mwReR93ISzCccYaBR7Ln7DZp3jagLTzHyvBrkcgxMOZtA7rqRu6227wNQC0SOWZ4tcjPgGqxv//m2W2eP0At8m/wa2G4AbNFgoftNg8byBYe/FoMzrwxAPpFgkfiTJr5z7ltxjxsPGnFFngd1p5jZvChwkaOv/3wY4M3f+Tk+NkPb7yB12HAKARGA5JL2AgoBwHmB0QoGgWjYBSMgpEMAGWBOzycB9UjAAAAAElFTkSuQmCC","orcid":"https://orcid.org/0000-0002-6302-7986","institution":"Washington University in St. Louis","correspondingAuthor":true,"submittingAuthor":false,"prefix":"","firstName":"Sun-Joo","middleName":"","lastName":"Lee","suffix":""},{"id":119373000,"identity":"b0c12ab9-0428-46f3-a001-678b8d00fed0","order_by":1,"name":"Colin Nichols","email":"","orcid":"https://orcid.org/0000-0002-4929-2134","institution":"Washington University in St. Louis","correspondingAuthor":false,"submittingAuthor":false,"prefix":"","firstName":"Colin","middleName":"","lastName":"Nichols","suffix":""}],"badges":[],"createdAt":"2022-06-24 17:35:32","currentVersionCode":1,"declarations":"","doi":"10.21203/rs.3.rs-1792750/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-1792750/v1","draftVersion":[],"editorialEvents":[],"editorialNote":"","failedWorkflow":false,"files":[{"id":23602545,"identity":"b324c336-40c1-47a6-9dd2-28613faa3bbc","added_by":"auto","created_at":"2022-07-07 19:03:31","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":2769122,"visible":true,"origin":"","legend":"","description":"","filename":"Article.pdf","url":"https://assets-eu.researchsquare.com/files/rs-1792750/v1_covered.pdf"},{"id":23602544,"identity":"3698603e-7f98-4e23-acda-3c3257d60b74","added_by":"auto","created_at":"2022-07-07 19:03:14","extension":"pdf","order_by":2,"title":"","display":"","copyAsset":false,"role":"supplement","size":1445906,"visible":true,"origin":"","legend":"","description":"","filename":"supplementary.pdf","url":"https://assets-eu.researchsquare.com/files/rs-1792750/v1/da30b75933fb554cd4c6329d.pdf"}],"financialInterests":"There is \u003cb\u003eNO\u003c/b\u003e Competing Interest.","formattedTitle":"\u003cp\u003e\u003csub\u003e\u003cspan class=\"ql-cursor\"\u003e\u003c/span\u003e\u003c/sub\u003eOxidation driven reversal of PIP\u003csub\u003e2\u003c/sub\u003e-dependent gating in GIRK2 channels\u003c/p\u003e","fulltext":[{"header":"Full Text","content":"This preprint is available for \u003ca href='/article/rs-1792750/latest.pdf' target='_blank'\u003edownload as a PDF\u003c/a\u003e."}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":false,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":true,"hideJournal":true,"highlight":"","institution":"","isAcceptedByJournal":false,"isAuthorSuppliedPdf":true,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true},"keywords":"GIRK2, Kir3, PIP2, oxidation, oxidative stress, reversed gating","lastPublishedDoi":"10.21203/rs.3.rs-1792750/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-1792750/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"Physiological activity of G protein gated inward rectifier K+ (GIRK, Kir3) channel activity is dynamically regulated by three key ligands, phosphoinositol-4,5-bisphosphate (PIP2), Gβγ, and Na+, which underlies cellular electrical response to multiple hormones and neurotransmitters in myocytes and neurons. In a reducing environment, matching that inside cells, purified GIRK2 (Kir3.2) channels demonstrated low basal activity, and appropriate sensitivity to the above ligands. However, under oxidizing conditions, anomalous behavior emerged, including rapid loss of agonist dependent activation, and high basal activity in the absence of any agonists that was paradoxically inhibited by PIP2. Mutagenesis identified two cysteine residues (C65 and C190) as being responsible for the loss of agonist dependent activation and the elevated basal activity, respectively. 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