This revised competitive renewal will elucidate the physiological role of the store-operated channel (SOC) and its complex regulation by iPLA2 and PKC in vascular constriction, activation of transcriptional factors and SMC proliferation. The central hypothesis of this proposal is that complex molecular regulation of SOC and their interplay with other channels and signaling cascades determines major SMC functions.
Three specific aims are designed to test the following specific hypotheses: 1) iPLA2-dependent activation of SOC channels provide not only a path for Ca2+ entry, but also a crucial trigger for activation of Ca2+ L channels, Ca2+ influx and constriction in cerebral arteries, 2) complex PKCe-dependent regulation of iPLA2 may provide a fine-tuning mechanism for SOC and capacitative Ca2+ entry during SMC proliferation, 3) activation of transcription factors and SMC proliferation depends on iPLA2 and SOC channels. An integrative approach that involves advanced electrophysiological, molecular, biochemical and imaging techniques will be used to test these hypotheses on the level of iPLA2, PKCepsilon, P-CREB and NFAT4, their translocation and cross-talk in freshly isolated and cultured SMC, and in intact blood vessels (fresh or organ culture). All these methods are established and used in PI's lab, and the feasibility of the proposed studies is strongly supported by preliminary data and recent publications.
Specific aims are:
Aim 1. To establish a novel role for iPLA2 and SOC channels as major determinants of Ca2+ influx and constriction in cerebral arteries. We will assess a complex role of SOC and its cross-talk with Ca2+ L channels during agonist-induced constriction and myogenic tone;
Aim 2. To define a complex role of PKCe in iPLA2-dependent regulation of SOC channels and CCE during SMC proliferation;
Aim 3. To establish the role of SOC and its iPLA2, PKC and lysophospholipid-dependent regulation in activation of CREB and NFAT, SMC proliferation and differentiation. .
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