Functional molecular complexes of junctophilin‐2 and caveolin‐1 provide a structural/functional basis for Ca2+‐microdomain formation between BKCa channels and RyRs in vascular smooth muscle cells

Abstract

Functional coupling between large-conductance Ca2+-activated K+ (BKCa) channels in plasma membrane (PM) and ryanodine receptors (RyRs) in sarcoplasmic reticulum (SR) is an essential mechanism for mechanical force regulation in smooth muscle cells (SMCs). Spontaneous Ca2+ release through RyRs (knows as Ca2+ sparks) and subsequent BKCa channels activation detected as spontaneous transient outward currents (STOCs) occur within PM-SR junctional sites, often called Ca2+ microdomains. We have previously reported that Caveolae, W-shaped invaginations of PM, accumulate BKCa channels within the structure and enhance coupling efficiency between Ca2+ sparks and BKCa channels/STOC activity in mouse mesenteric artery SMCs (mMASMCs). However, the molecular basis between caveolae and RyRs in SR membrane is unclear. In the present study, we demonstrated that both caveolin-1 (Cav1), a caveola forming protein, and junctophilin-2 (JP2), a bridging protein between PM and SR, reciprocally contribute to Ca2+microdomain formation. Co-immunoprecipitation using rat mesenteric arterial tissues and double-immunocytochemical staining analyses by total internal reflection fluorescent microscopy revealed the direct interaction between JP2 and Cav1. Mouse mesenteric arterial tissues were organ-cultured and concomitantly treated with siRNA using reversible permeabilization procedures. Knockdown of JP2 in mMASMCs significantly decreased the colocalization of Cav1 and RyRs and the amplitude of STOCs. The contraction induced by the inhibition of BKCa channels/STOC activity and subsequent membrane depolarization in siJP2-treated mMASMCs was significantly smaller than that in siControl. These results suggested that the novel interaction of JP2 with Cav1 seems to provide a structural/functional basis for Ca2+-mediated crosstalk between RyRs and BKCa channels, and likely positions the PM-SR junctional sites to convert Ca2+ spark signals into hyperpolarizing signals in SMCs lacking transverse tubular system. This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.