Involvement of STIM1 in the proteinase-activated receptor 1-mediated Ca2+ influx in vascular endothelial cells.
Thrombin increases the cytosolic Ca(2+) concentrations and induces NO production by activating proteinase-activated receptor 1 (PAR(1)) in vascular endothelial cells. The store-operated Ca(2+) influx is a major Ca(2+) influx pathway in non-excitable cells including endothelial cells and it has been reported to play a role in the thrombin-induced Ca(2+) signaling in endothelial cells. Recent studies have identified stromal interaction molecule 1 (STIM1) to function as a sensor of the store site Ca(2+) content, thereby regulating the store-operated Ca(2+) influx. However, the functional role of STIM1 in the thrombin-induced Ca(2+) influx and NO production in endothelial cells still remains to be elucidated. Fura-2 and diaminorhodamine-4M fluorometry was utilized to evaluate the thrombin-induced changes in cytosolic Ca(2+) concentrations and NO production, respectively, in porcine aortic endothelial cells transfected with small interfering RNA (siRNA) targeted to STIM1. STIM1-targeted siRNA suppressed the STIM1 expression and the thapsigargin-induced Ca(2+) influx. The degree of suppression of the STIM1 expression correlated well to the degree of suppression of the Ca(2+) influx. The knockdown of STIM1 was associated with a substantial inhibition of the Ca(2+) influx and a partial reduction of the NO production induced by thrombin. The thrombin-induced Ca(2+) influx exhibited the similar sensitivity toward the Ca(2+) influx inhibitors to that seen with the thapsigargin-induced Ca(2+) influx. The present study provides the first evidence that STIM1 plays a critical role in the PAR(1)-mediated Ca(2+) influx and Ca(2+)-dependent component of the NO production in endothelial cells.
Hirano
Katsuya
K
Division of Molecular Cardiology, Research Institute of Angiocardiology, Graduate School of Medical Sciences, Kyushu University, Higashi-ku, Fukuoka, Japan. khirano@molcar.med.kyushu-u.c.jp
Hirano
Mayumi
M
Hanada
Akiko
A
D000818:Animals; D001011:Aorta / Q000166:cytology; D001483:Base Sequence; D002121:Calcium Channel Blockers / Q000378:metabolism; D020013:Calcium Signaling; D015815:Cell Adhesion Molecules / Q000378:metabolism; D002460:Cell Line; D002478:Cells, Cultured; D042783:Endothelial Cells; D004730:Endothelium, Vascular / Q000166:cytology / Q000378:metabolism*; D055785:Gene Knockdown Techniques; D006367:HeLa Cells; D006801:Humans; D008565:Membrane Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D009363:Neoplasm Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D009569:Nitric Oxide / Q000096:biosynthesis*; D010446:Peptide Fragments / Q000378:metabolism; D034741:RNA, Small Interfering; D044463:Receptor, PAR-1 / Q000737:chemistry / Q000378:metabolism*; D053498:Sarcoplasmic Reticulum Calcium-Transporting ATPases / Q000037:antagonists & inhibitors; D016415:Sequence Alignment; D000071737:Stromal Interaction Molecule 1; D000071738:Stromal Interaction Molecule 2; D013552:Swine; D013917:Thrombin / Q000378:metabolism; D013997:Time Factors
D002121:Calcium Channel Blockers; D015815:Cell Adhesion Molecules; D008565:Membrane Proteins; D009363:Neoplasm Proteins; D010446:Peptide Fragments; D034741:RNA, Small Interfering; D044463:Receptor, PAR-1; C103530:STIM1 protein, human; C513279:STIM2 protein, human; D000071737:Stromal Interaction Molecule 1; D000071738:Stromal Interaction Molecule 2; D009569:Nitric Oxide; D013917:Thrombin; D053498:Sarcoplasmic Reticulum Calcium-Transporting ATPases