Prolonged exposure to agonist results in a reduction in the levels of the Gq/G11 alpha subunits in cultured vascular smooth muscle cells.
Recent studies have shown that G proteins are a potential regulatory site in the transmembrane signaling cascade. The aim of this study was to examine the effects of prolonged agonist exposure on expression of the Gq class of G protein alpha subunits (G alpha q/G alpha 11) in cultured rat vascular smooth muscle cells (VSMC). Treatment with 100 nM angiotensin II (Ang II) led to a substantial sustained down-regulation of cellular levels of immunologically detectable G alpha q/G alpha 11 by 50% within 6 hr. The effect of Ang II was dose dependent with an EC50 of 2 nM and was specifically blocked by the vascular type-1 Ang II receptor-specific antagonist losartan. The Ang II-induced reduction in cellular levels of G protein alpha subunits was specific for G alpha q/G alpha 11. The calcium ionophore ionomycin or activators of ubiquitous protein kinases (phorbol-12-myristate-13-acetate, forskolin, and 8-bromo-cGMP) did not mimic the effects of Ang II. However, [Arg8]vasopressin also induced a significant loss in cellular G alpha q/G alpha 11 levels. Ang II-induced G alpha q/G alpha 11 down-regulation was reversed by prevention of cellular receptor processing with phenylarsine oxide or chronic potassium depletion. The effects of Ang II on G alpha q/G alpha 11 levels were inhibited when protein kinase C activity was abolished. G alpha q mRNA levels were down-regulated by 30% after 4-hr incubation with Ang II, in part by transcriptional regulation. Although a short term vasopressin pretreatment had no effect on inositol-1,4,5-trisphosphate (IP3) generation in response to subsequent Ang II stimulation, a partial heterologous desensitization of the IP3 response was induced after a long term vasopressin pretreatment, which concurrently down-regulated cellular G alpha q/G alpha 11 levels. Homologous desensitization of IP3 generation on a second Ang II stimulation was observed after both a short and long term Ang II pretreatment. In conclusion, prolonged exposure to Ang II induces down-regulation of cellular G alpha q/G alpha 11 levels in intact VSMC. The effect of Ang II appears to be mediated by the signaling pathway sensitive to inhibition of receptor processing. The present study raises the possibility that agonist-induced G alpha q/G alpha 11 down-regulation participates in the mechanism of long term desensitization of the G alpha q/G alpha 11-mediated signaling system in VSMC.
Kai
H
H
Department of Medicine, Emory University School of Medicine, Atlanta, Georgia 30322, USA.
Fukui
T
T
Lassègue
B
B
Shah
A
A
Minieri
C A
CA
Griendling
K K
KK
D000804:Angiotensin II / Q000494:pharmacology*; D000818:Animals; D001483:Base Sequence; D002478:Cells, Cultured; D017931:DNA Primers; D004789:Enzyme Activation; D019204:GTP-Binding Proteins / Q000819:agonists* / Q000235:genetics / Q000378:metabolism; D007700:Kinetics; D008297:Male; D008969:Molecular Sequence Data; D009131:Muscle, Smooth, Vascular / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D011493:Protein Kinase C / Q000378:metabolism; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D015398:Signal Transduction; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology
D017931:DNA Primers; D012333:RNA, Messenger; D000804:Angiotensin II; D011493:Protein Kinase C; D019204:GTP-Binding Proteins; D013755:Tetradecanoylphorbol Acetate