Local Ca²⁺ Signals within Caveolae Cause Nuclear Translocation of CaMK1α in Mouse Vascular Smooth Muscle Cells

Abstract

An increase in intracellular Ca 2 concentration ([Ca 2 ] i ) activates Ca 2 -sensitive enzymes such as Ca 2 / calmodulin-dependent kinases (CaMK) and induces gene transcription in various types of cells. This signaling pathway is called excitation-transcription (E-T) coupling. Recently, we have revealed that a L-type Ca 2 channel/CaMK kinase (CaMKK) 2/CaMK1α complex located within caveolae in vascular smooth muscle cells (SMCs) can convert [Ca 2 ] i changes to gene transcription profiles that are related to chemotaxis. Although CaMK1α is expected to be the key molecular identity that can transport Ca 2 signals originated within caveolae to the nucleus, data sets directly proving this scheme are lacking. In this study, multicolor fluorescence imaging methods were utilized to address this question. Live cell imaging using mouse primary aortic SMCs revealed that CaMK1α can translocate from the cytosol to the nucleus; and that this movement was blocked by nifedipine or a CaMKK inhibitor, STO609. Experiments using two types of Ca 2 chelators, ethylene glycol-bis(2-aminoethylether)-N,N,N′,N′-tetraacetic acid (EGTA) and 1,2-bis(2-aminophenoxy)ethane-N,N,N′,N′-tetraacetic acid (BAPTA), combined with caveolin-1 knockout (cav1-KO) mice showed that local Ca 2 events within caveolae are required to trigger this CaMK1α nuclear translocation. Importantly, overexpression of cav1 in isolated cav1-KO myocytes recovered the CaMK1α translocation. In SMCs freshly isolated from mesenteric arteries, CaMK1α was localized mainly within caveolae in the resting state. Membrane depolarization induced both nuclear translocation and phosphorylation of CaMK1α. These responses were inhibited by nifedipine, STO609, cav1-KO, or BAPTA. These new findings strongly suggest that CaMK1α can transduce Ca 2 signaling generated within or very near caveolae to the nucleus and thus, promote E-T coupling.

Physiological and Pathological Significance of Chloride Channels

“…10) Caveolin 1 is a scaffold protein that assembles several molecules as a complex in caveolae, which are raft structures on the surface of plasma membrane. 30) Caveolin 1 interacts with VDCC, 31) large-conductance Ca 2+ -activated K + (BK Ca ) channels, 31,32) and Ca 2+ /calmodulin-dependent kinases 33,34) and their complexes accumulate in the caveolae of VSMC. These functional couplings may be recognized as a specific domain of efficient and effective Ca 2+ signaling in VSMC.…”