G-proteins of fat-cells. Role in hormonal regulation of intracellular inositol 1,4,5-trisphosphate.
Pertussis toxin abolishes hormonal inhibition of adenylate cyclase, hormonal stimulation of inositol 1,4,5-trisphosphate accumulation in rat fat-cells, and catalyses the ADP-ribosylation of two peptides, of Mr 39,000 and 41,000 [Malbon, Rapiejko & Mangano (1985) J. Biol. Chem. 260, 2558-2564]. The 41,000-Mr peptide is the alpha-subunit of the G-protein, referred to as Gi, that is believed to mediate inhibitory control of adenylate cyclase by hormones. The nature of the 39,000-Mr substrate for pertussis toxin was investigated. The fat-cell 39,000-Mr peptide was compared structurally and immunologically with the alpha-subunits of two other G-proteins, Gt isolated from the rod outer segments of bovine retina and Go isolated from bovine brain. After radiolabelling in the presence of pertussis toxin and [32P]NAD+, the electrophoretic mobilities of the fat-cell 39,000-Mr peptide and the alpha-subunits of Go and Gt were nearly identical. Partial proteolysis of these ADP-ribosylated proteins generates peptide patterns that suggest the existence of a high degree of homology between the fat-cell 39,000-Mr peptide and the alpha-subunit of Go. Antisera raised against purified G-proteins and their subunits were used to probe immunoblots of purified Gt, Gi, Go, and fat-cell membrane proteins. Although recognizing the 36,000-Mr beta-subunit band of Gt, Gi, Go and a 36,000-Mr fat-cell peptide, antisera raised against Gt failed to recognize either the 39,000- or the 41,000-Mr peptides of fat-cells or the alpha-subunits of Go and Gi. Antisera raised against the alpha-subunit of Go, in contrast, recognized the 39,000-Mr peptide of rat fat-cells, but not the alpha-subunit of either Gi or Gt. These data establish the identity of Go, in addition to Gi, in fat-cell membranes and suggest the possibility that either Go or Gi alone, or both, may mediate hormonal regulation of adenylate cyclase and phospholipase C.
D037361:Adenylate Cyclase Toxin; D000273:Adipose Tissue / Q000187:drug effects / Q000378:metabolism*; D000818:Animals; D001921:Brain / Q000378:metabolism; D002417:Cattle; D002462:Cell Membrane / Q000187:drug effects / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D004837:Epinephrine / Q000494:pharmacology; D005260:Female; D019204:GTP-Binding Proteins / Q000378:metabolism*; D015544:Inositol 1,4,5-Trisphosphate; D007295:Inositol Phosphates / Q000378:metabolism*; D037342:Pertussis Toxin; D051381:Rats; D011919:Rats, Inbred Strains; D012374:Rod Cell Outer Segment / Q000378:metabolism; D013403:Sugar Phosphates / Q000378:metabolism*; D010566:Virulence Factors, Bordetella / Q000494:pharmacology
D037361:Adenylate Cyclase Toxin; D007295:Inositol Phosphates; D013403:Sugar Phosphates; D010566:Virulence Factors, Bordetella; D015544:Inositol 1,4,5-Trisphosphate; D037342:Pertussis Toxin; D019204:GTP-Binding Proteins; D004837:Epinephrine