Evidence for a regulated interaction between heterotrimeric G proteins and caveolin.
Caveolae are flask-shaped plasma membrane specializations. A 22-kDa protein, caveolin, is a principal component of caveolar membranes in vivo. As recent evidence suggests that caveolae may participate in G protein-coupled signaling events, we have investigated the potential interaction of caveolin with heterotrimeric G proteins. Using cell fractionation techniques, we found that mutational or pharmacologic activation of Gs alpha prevents its cofractionation with caveolin. In a second independent approach, we directly examined the interaction of G proteins with caveolin. For this purpose, we recombinantly expressed caveolin as a glutathione S-transferase fusion protein. Using an in vitro binding assay, we found that caveolin interacts with G protein alpha subunits (Gs, Go, and Gi). Mutational or pharmacologic activation (with guanosine 5'-O-(thiotriphosphate)) of G alpha subunits prevents this interaction, indicating that the inactive GDP-bound form of G alpha subunits preferentially interacts with caveolin. This G protein binding activity is located within a 41-amino acid region of caveolin's cytoplasmic N-terminal domain (residues 61-101). Further functional analysis shows that a polypeptide derived from this region of caveolin (residues 82-101) effectively suppresses the basal activity of purified G proteins, apparently by inhibiting GDP/GTP exchange. This caveolin sequence is homologous to a region of the Rab GDP dissociation inhibitor, a known inhibitor of GDP/GTP exchange for Rab proteins. These data suggest that caveolin could function to negatively regulate the activation state of heterotrimeric G proteins.
Li
S
S
Whitehead Institute for Biomedical Research, Cambridge, Massachusetts 02142-1479, USA.
Okamoto
T
T
Chun
M
M
Sargiacomo
M
M
Casanova
J E
JE
Hansen
S H
SH
Nishimoto
I
I
Lisanti
M P
MP
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D051242:Caveolin 1; D022461:Caveolins; D002478:Cells, Cultured; D004285:Dogs; D020558:GTP Phosphohydrolases / Q000378:metabolism; D019204:GTP-Binding Proteins / Q000737:chemistry / Q000378:metabolism*; D006153:Guanosine Diphosphate / Q000378:metabolism; D006160:Guanosine Triphosphate / Q000378:metabolism; D008565:Membrane Proteins / Q000378:metabolism*; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000378:metabolism
D051242:Caveolin 1; D022461:Caveolins; D008565:Membrane Proteins; D010446:Peptide Fragments; D006153:Guanosine Diphosphate; D006160:Guanosine Triphosphate; D020558:GTP Phosphohydrolases; D019204:GTP-Binding Proteins