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Alternative splicing of Cav1.2 channel exons in smooth muscle cells of resistance-size arteries generates currents with unique electrophysiological properties.

ABSTRACT: Voltage-dependent calcium (Ca(2+), Ca(V)1.2) channels are the primary Ca(2+) entry pathway in smooth muscle cells of resistance-size (myogenic) arteries, but their molecular identity remains unclear. Here we identified and quantified Ca(V)1.2 alpha(1)-subunit splice variation in myocytes of rat resistance-size (100-200 microm diameter) cerebral arteries. Full-length clones containing either exon 1b or the recently identified exon 1c exhibited additional primary splice variation at exons 9*, 21/22, 31/32, and +/- 33. Real-time PCR confirmed the findings from full-length clones and indicated that the major Ca(V)1.2 variant contained exons 1c, 8, 21, and 32+33, with approximately 57% containing 9*. Exon 9* was more prevalent in clones containing 1c (72%) than in those containing 1b (33%), suggesting exon-selective combinatorial splicing. To examine the functional significance of this splicing profile, membrane currents produced by each of the four exon 1b/c/ +/- 9* variants were characterized following transfection in HEK293 cells. Exon 1c and 9* caused similar hyperpolarizing shifts in both current-voltage relationships and voltage-dependent activation of currents. Furthermore, exon 9* induced a hyperpolarizing shift only in the voltage-dependent activation of channels containing exon 1b, but not in those containing exon 1c. In contrast, exon 1b, 1c, or +9* did not alter voltage-dependent inactivation. In summary, we have identified the Ca(V)1.2 alpha(1)-subunit splice variant population that is expressed in myocytes of resistance-size arteries and the unique electrophysiological properties of recombinant channels formed by exon 1 and 9* variation. The predominance of exon 1c and 9* in smooth muscle cell Ca(V)1.2 channels causes a hyperpolarizing shift in the voltage sensitivity of currents toward the physiological arterial voltage range.

SUBMITTER: Cheng X

PROVIDER: S-EPMC2724194 | biostudies-literature | 2009 Aug

REPOSITORIES: biostudies-literature

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Alternative splicing of Cav1.2 channel exons in smooth muscle cells of resistance-size arteries generates currents with unique electrophysiological properties.

Cheng Xiaoyang X Pachuau Judith J Blaskova Eva E Asuncion-Chin Maria M Liu Jianxi J Dopico Alejandro M AM Jaggar Jonathan H JH

American journal of physiology. Heart and circulatory physiology 20090605 2

Voltage-dependent calcium (Ca(2+), Ca(V)1.2) channels are the primary Ca(2+) entry pathway in smooth muscle cells of resistance-size (myogenic) arteries, but their molecular identity remains unclear. Here we identified and quantified Ca(V)1.2 alpha(1)-subunit splice variation in myocytes of rat resistance-size (100-200 microm diameter) cerebral arteries. Full-length clones containing either exon 1b or the recently identified exon 1c exhibited additional primary splice variation at exons 9*, 21/2 ...[more]

PMID: 19502562

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Project description:Physiological functions of mitochondria in contractile arterial myocytes are poorly understood. Mitochondria can uptake calcium (Ca(2+)), but intracellular Ca(2+) signals that regulate mitochondrial Ca(2+) concentration ([Ca(2+)](mito)) and physiological functions of changes in [Ca(2+)](mito) in arterial myocytes are unclear.To identify Ca(2+) signals that regulate [Ca(2+)](mito), examine the significance of changes in [Ca(2+)](mito), and test the hypothesis that [Ca(2+)](mito) controls functional ion channel transcription in myocytes of resistance-size cerebral arteries.Endothelin (ET)-1 activated Ca(2+) waves and elevated global Ca(2+) concentration ([Ca(2+)](i)) via inositol 1,4,5-trisphosphate receptor (IP(3)R) activation. IP(3)R-mediated sarcoplasmic reticulum (SR) Ca(2+) release increased [Ca(2+)](mito) and induced mitochondrial depolarization, which stimulated mitochondrial reactive oxygen species (mitoROS) generation that elevated cytosolic ROS. In contrast, a global [Ca(2+)](i) elevation did not alter [Ca(2+)](mito), mitochondrial potential, or mitoROS generation. ET-1 stimulated nuclear translocation of nuclear factor (NF)-kappaB p50 subunit and ET-1-induced IP(3)R-mediated mitoROS elevated NF-kappaB-dependent transcriptional activity. ET-1 elevated voltage-dependent Ca(2+) (Ca(V)1.2) channel expression, leading to an increase in both pressure (myogenic tone)- and depolarization-induced vasoconstriction. Baseline Ca(V)1.2 expression and the ET-1-induced elevation in Ca(V)1.2 expression were both reduced by IP(3)R inhibition, mitochondrial electron transport chain block, antioxidant treatment, and NF-kappaB subunit knockdown, leading to vasodilation.IP(3)R-mediated SR Ca(2+) release elevates [Ca(2+)](mito), which induces mitoROS generation. MitoROS activate NF-kappaB, which stimulates Ca(V)1.2 channel transcription. Thus, mitochondria sense IP(3)R-mediated SR Ca(2+) release to control NF-kappaB-dependent Ca(V)1.2 channel expression in arterial myocytes, thereby modulating arterial contractility.

Dataset Information

Alternative splicing of Cav1.2 channel exons in smooth muscle cells of resistance-size arteries generates currents with unique electrophysiological properties.

Publications

Alternative splicing of Cav1.2 channel exons in smooth muscle cells of resistance-size arteries generates currents with unique electrophysiological properties.

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