Direct inhibition of the interaction between alpha-interaction domain and beta-interaction domain of voltage-dependent Ca2+ channels by Gem.
The Ras-related small G-protein Gem regulates voltage-dependent Ca2+ channels (VDCCs) through interaction with the beta-subunit of the VDCC. This action of Gem is mediated by regulated alpha1-subunit expression at the plasma membrane. In the present study, we examined the mechanism of the inhibition of VDCC activity by Gem. The beta-interaction domain (BID) of the beta-subunit, which specifically interacts with the alpha-interaction domain (AID) of the alpha1-subunit, is shown to be essential for the interaction between Gem and beta-subunits. In addition, the AID peptide inhibited interaction between Gem and beta-subunits in a dose-dependent manner. GemS88N mutant, which has low binding affinity for guanine nucleotide, did not interact with beta-subunits, allowing alpha1-subunit expression at the plasma membrane. This inhibitory effect of wild-type Gem on VDCC activity was reduced in cells expressing GemS88N. The overexpression of wild-type Gem in pancreatic beta-cell line MIN6 cells suppressed Ca2+-triggered secretion, whereas overexpression of GemS88N induced Ca2+-triggered secretion to control level. These results suggest that GTPase activity of Gem is required for the binding of Gem to BID that regulates VDCC activity through interaction with AID.
Sasaki
Takehide
T
Division of Cellular and Molecular Medicine, Kobe University Graduate School of Medicine, Kobe 650-0017, Japan.
Shibasaki
Tadao
T
Béguin
Pascal
P
Nagashima
Kazuaki
K
Miyazaki
Masaru
M
Seino
Susumu
S
D000818:Animals; D015220:Calcium Channels / Q000737:chemistry / Q000502:physiology*; D002460:Cell Line; D019382:Human Growth Hormone / Q000378:metabolism; D017874:Immediate-Early Proteins / Q000378:metabolism*; D051379:Mice; D020559:Monomeric GTP-Binding Proteins / Q000378:metabolism*; D016716:PC12 Cells; D010673:Pheochromocytoma; D011485:Protein Binding; D021122:Protein Subunits / Q000378:metabolism; D011817:Rabbits; D051381:Rats; D011993:Recombinant Fusion Proteins / Q000378:metabolism
D015220:Calcium Channels; D017874:Immediate-Early Proteins; D021122:Protein Subunits; D011993:Recombinant Fusion Proteins; D019382:Human Growth Hormone; C088119:Gem protein, mouse; D020559:Monomeric GTP-Binding Proteins