Slo1 Caveolin-binding Motif, a Mechanism of Caveolin-1-Slo1 Interaction Regulating Slo1 Surface Expression*
(2007)
Journal - Journal of Biological Chemistry
Abstract :
The large conductance, voltage- and Ca2+-activated potassium(MaxiK, BK) channel and caveolin-1 play important roles in regulatingvascular contractility. Here, we hypothesized that the MaxiK-subunit (Slo1) and caveolin-1 may interact with each other.Slo1 and caveolin-1 physiological association in native vasculartissue is strongly supported by (i) detergent-free purificationof caveolin-1-rich domains demonstrating a pool of aortic Slo1co-migrating with caveolin-1 to light density sucrose fractions,(ii) reverse co-immunoprecipitation, and (iii) double immunolabelingof freshly isolated myocytes revealing caveolin-1 and Slo1 proximityat the plasmalemma. In HEK293T cells, Slo1-caveolin-1 associationwas unaffected by the smooth muscle MaxiK ß1-subunit. Sequenceanalysis revealed two potential caveolin-binding motifs alongthe Slo1 C terminus, one equivalent, 1007YNMLCFGIY1015, andanother mirror image, 537YTEYLSSAF545, to the consensus sequence,XXXXXX. Deletion of 1007YNMLCFGIY1015 caused 80% loss of Slo1-caveolin-1association while preserving channel normal folding and overallSlo1 and caveolin-1 intracellular distribution patterns. 537YTEYLSSAF545deletion had an insignificant dissociative effect. Interestingly,caveolin-1 coexpression reduced Slo1 surface and functionalexpression near 70% without affecting channel voltage sensitivity,and deletion of 1007YNMLCFGIY1015 motif obliterated channelsurface expression. The results suggest 1007YNMLCFGIY1015 possibleparticipation in Slo1 plasmalemmal targeting and demonstrateits role as a main mechanism for caveolin-1 association withSlo1 potentially serving a dual role: (i) maintaining channelsin intracellular compartments downsizing their surface expressionand/or (ii) serving as anchor of plasma membrane resident channelsto caveolin-1-rich membranes. Because the caveolin-1 scaffoldingdomain is juxtamembrane, it is tempting to suggest that Slo1-caveolin-1interaction facilitates the tethering of the Slo1 C-terminalend to the membrane.* This work was supported by National Institutes of Health GrantsHL54970 and HL77705 (to L. T.), HD046510 (to E. S.), and P01-HL080111(to E. S. and L. T.) and by American Heart Association NationalCenter Grants 0435084N (to A. A.) and 0435116N (to M. E.). Thecosts of publication of this article were defrayed in part bythe payment of page charges. This article must therefore behereby marked "advertisement" in accordance with 18 U.S.C. Section1734 solely to indicate this fact.1 Both authors contributed equally to this work.3 Co-senior authors.