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Protein kinase C (PKC) activity regulates functional effects of Kvß1.3 subunit on Kv1.5 channels Identification of a cardiac Kv1.5 channelsome

AuthorsDavid, Miren CSIC; Macías, Álvaro CSIC ORCID; Moreno, Cristina CSIC; Prieto, Ángela CSIC; González, Teresa CSIC ORCID; Felipe, Antonio; Tamkun, Michael M.; Valenzuela, Carmen CSIC ORCID CVN
Issue Date2012
PublisherAmerican Society for Biochemistry and Molecular Biology
CitationJournal of Biological Chemistry 287(25): 21416-21428 (2012)
AbstractKv1.5 channels are the primary channels contributing to the ultrarapid outward potassium current (IKur). The regulatory Kvβ1.3 subunit converts Kv1.5 channels from delayed rectifiers with a modest degree of slow inactivation to channels with both fast and slow inactivation components. Previous studies have shown that inhibition of PKC with calphostin C abolishes the fast inactivation induced by Kvβ1.3. In this study, we investigated the mechanisms underlying this phenomenon using electrophysiological, biochemical, and confocal microscopy approaches. To achieve this, we used HEK293 cells (which lack Kvβ subunits) transiently cotransfected with Kv1.5+Kvβ1.3 and also rat ventricular and atrial tissue to study native α-β subunit interactions. Immunocytochemistry assays demonstrated that these channel subunits colocalize in control conditions and after calphostin C treatment. Moreover, coimmunoprecipitation studies showed that Kv1.5 and Kvβ1.3 remain associated after PKC inhibition. After knocking down all PKC isoforms by siRNA or inhibiting PKC with calphostin C, Kvβ1.3-induced fast inactivation at +60 mV was abolished. However, depolarization to +100 mV revealed Kvβ1.3-induced inactivation, indicating that PKC inhibition causes a dramatic positive shift of the inactivation curve. Our results demonstrate that calphostin C-mediated abolishment of fast inactivation is not due to the dissociation of Kv1.5 and Kvβ1.3. Finally, immunoprecipitation and immunocytochemistry experiments revealed an association between Kv1.5, Kvβ1.3, the receptor for activated C kinase (RACK1), PKCβI, PKCβII, and PKCθ in HEK293 cells. A very similar Kv1.5 channelosome was found in rat ventricular tissue but not in atrial tissue.
Publisher version (URL)
Identifiersdoi: 10.1074/jbc.M111.328278
issn: 0021-9258
e-issn: 1083-351X
Appears in Collections:(IIBM) Artículos

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