Luteinizing hormone/choriogonadotropin-dependent, cholera toxin-catalyzed adenosine 5'-diphosphate (ADP)-ribosylation of the long and short forms of Gs alpha and pertussis toxin-catalyzed ADP-ribosylation of Gi alpha*.
Although it is well established that activated LH/human (h) CG receptor stimulates adenylyl cyclase activity (via the heterotrimeric stimulatory guanine nucleotide-binding protein, Gs) and in some cells stimulates phospholipase C activity, there is no evidence for a direct physical interaction between the LH/CG receptor and Gs or any other G protein(s). We conducted studies using cholera toxin (CTX) and pertussis toxin (PTX) to determine which G alpha proteins were associated with the LH/CG receptor in ovarian follicular membranes. Since hormone-dependent, CTX-catalyzed ADP ribosylation (AR) constitutes evidence that a G alpha protein is specifically associated with a receptor, CTX-catalyzed AR of membrane proteins was examined both in the presence and absence of guanine nucleotides to determine which G proteins exhibit hCG-dependent labeling by [32P]NAD. Results demonstrated the time- and hCG-dependent AR of both a 45-kDa protein and a 48/50-kDa doublet as well as a 40-kDa protein that was also sensitive to AR by PTX in a time- and hCG-dependent manner. Using anti-G protein antisera to specifically immunoprecipitate photoaffinity-labeled G proteins, we were able to identify the 45- and 48/50 kDa proteins as the short and long forms of Gs alpha and the 40-kDa protein as Gi alpha. A monoclonal anti-hCG antibody immunoprecipitated the activated LH/CG receptor along with the long and short forms of Gs alpha and Gi. These results suggest that a portion of Gi along with the long and short forms of Gs alpha are associated physically with the LH/CG receptor in ovarian follicular membranes.
Rajagopalan-Gupta
R M
RM
Department of Cell and Molecular Biology, Northwestern University Medical School, Chicago, Illinois 60611, USA.
Rasenick
M M
MM
Hunzicker-Dunn
M
M
D000244:Adenosine Diphosphate / Q000378:metabolism*; D037361:Adenylate Cyclase Toxin; D000818:Animals; D002478:Cells, Cultured; D002772:Cholera Toxin / Q000494:pharmacology; D005260:Female; D019204:GTP-Binding Proteins / Q000378:metabolism*; D006801:Humans; D007986:Luteinizing Hormone / Q000378:metabolism*; D010053:Ovary / Q000378:metabolism*; D037342:Pertussis Toxin; D011974:Receptors, LH / Q000819:agonists / Q000378:metabolism*; D015398:Signal Transduction / Q000187:drug effects; D013552:Swine; D010566:Virulence Factors, Bordetella / Q000494:pharmacology
D037361:Adenylate Cyclase Toxin; D011974:Receptors, LH; D010566:Virulence Factors, Bordetella; D000244:Adenosine Diphosphate; D007986:Luteinizing Hormone; D002772:Cholera Toxin; D037342:Pertussis Toxin; D019204:GTP-Binding Proteins