Essential role for STIM1/Orai1-mediated calcium influx in PDGF-induced smooth muscle migration.
We recently demonstrated that thapsigargin-induced passive store depletion activates Ca(2+) entry in vascular smooth muscle cells (VSMC) through stromal interaction molecule 1 (STIM1)/Orai1, independently of transient receptor potential canonical (TRPC) channels. However, under physiological stimulations, despite the ubiquitous depletion of inositol 1,4,5-trisphosphate-sensitive stores, many VSMC PLC-coupled agonists (e.g., vasopressin and endothelin) activate various store-independent Ca(2+) entry channels. Platelet-derived growth factor (PDGF) is an important VSMC promigratory agonist with an established role in vascular disease. Nevertheless, the molecular identity of the Ca(2+) channels activated by PDGF in VSMC remains unknown. Here we show that inhibitors of store-operated Ca(2+) entry (Gd(3+) and 2-aminoethoxydiphenyl borate at concentrations as low as 5 microM) prevent PDGF-mediated Ca(2+) entry in cultured rat aortic VSMC. Protein knockdown of STIM1, Orai1, and PDGF receptor-beta (PDGFRbeta) impaired PDGF-mediated Ca(2+) influx, whereas Orai2, Orai3, TRPC1, TRPC4, and TRPC6 knockdown had no effect. Scratch wound assay showed that knockdown of STIM1, Orai1, or PDGFRbeta inhibited PDGF-mediated VSMC migration, but knockdown of STIM2, Orai2, and Orai3 was without effect. STIM1, Orai1, and PDGFRbeta mRNA levels were upregulated in vivo in VSMC from balloon-injured rat carotid arteries compared with noninjured control vessels. Protein levels of STIM1 and Orai1 were also upregulated in medial and neointimal VSMC from injured carotid arteries compared with noninjured vessels, as assessed by immunofluorescence microscopy. These results establish that STIM1 and Orai1 are important components for PDGF-mediated Ca(2+) entry and migration in VSMC and are upregulated in vivo during vascular injury and provide insights linking PDGF to STIM1/Orai1 during neointima formation.
Bisaillon
Jonathan M
JM
Center for Cardiovascular Sciences, Albany Medical College, NY 12208, USA. trebakm@mail.amc.edu
Motiani
Rajender K
RK
Gonzalez-Cobos
José C
JC
Potier
Marie
M
Halligan
Katharine E
KE
Alzawahra
Wael F
WF
Barroso
Margarida
M
Singer
Harold A
HA
Jourd'heuil
David
D
Trebak
Mohamed
M
D000199:Actins; D000818:Animals; D002118:Calcium / Q000378:metabolism*; D015220:Calcium Channels / Q000235:genetics / Q000378:metabolism*; D002465:Cell Movement / Q000187:drug effects; D002478:Cells, Cultured; D005786:Gene Expression Regulation / Q000502:physiology; D008297:Male; D008562:Membrane Glycoproteins / Q000235:genetics / Q000378:metabolism*; D009131:Muscle, Smooth, Vascular / Q000166:cytology; D032389:Myocytes, Smooth Muscle / Q000187:drug effects* / Q000502:physiology; D000071740:ORAI1 Protein; D010982:Platelet-Derived Growth Factor / Q000494:pharmacology*; D051381:Rats; D015398:Signal Transduction; D000071737:Stromal Interaction Molecule 1
D000199:Actins; D015220:Calcium Channels; D008562:Membrane Glycoproteins; D000071740:ORAI1 Protein; C532493:Orai1 protein, rat; D010982:Platelet-Derived Growth Factor; C518857:Stim1 protein, rat; D000071737:Stromal Interaction Molecule 1; D002118:Calcium