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Accessory β 3 subunits promote the functional expression of voltage‐gated calcium channels in vascular smooth muscle cells
Author(s) -
Sonkusare Swapnil,
Rhee Sung W,
Thakali Keshari,
Fletcher Terry W,
Stimers Joseph R,
Telemaque Sabine,
Rusch Nancy J
Publication year - 2008
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.22.1_supplement.912.35
Subject(s) - vascular smooth muscle , protein subunit , western blot , microbiology and biotechnology , myocyte , voltage dependent calcium channel , biology , chemistry , medicine , endocrinology , calcium , smooth muscle , biochemistry , gene
Four distinct β subunits (β 1–4 ) promote tissue‐specific expression of the voltage‐dependent Ca 2+ channel pore‐forming α 1C subunits. Our goal was to identify β subunits promoting the functional expression of α 1C in vascular smooth muscle cells (VSMCs). Expression of all four β subunits was detected at the transcript level in rat small renal, mesenteric and femoral arteries. Western blot analysis suggested that β 3 and to a lesser extent, β 2 subunits were expressed at the protein level in these resistance arteries, which led us to postulate that β 3 promotes α 1C expression. To evaluate this hypothesis, β 3 siRNA was delivered into A10 and A7r5 (embryonic rat thoracic aorta) smooth muscle cell lines by electroporation. RT‐PCR showed nearly 75% reduction in β 3 transcript 24 hrs after siRNA delivery, but no change in α 1C transcript. At the protein level, β 3 subunit expression was reduced 48 hrs after siRNA delivery, and a concurrent loss of α 1C expression was observed. Expression levels of GAPDH and α‐actin (internal standards) were unchanged. Patch‐clamp studies showed a reduced voltage‐dependent Ca 2+ current density in both cell lines 48 hrs after siRNA delivery (A7r5 cells: −4.4±1.6 pA/pF Vs −11.1±2.4 pA/pF; A10 cells: −2.8±0.4 pA/pF Vs −4.8±0.5 pA/pF, n= 3–4). Thus, β 3 subunits may promote the expression of α 1C to enhance voltage‐dependent Ca 2+ influx at the surface membrane of VSMCs. Supported by R01 HL064806‐07(NJR).