Antibodies to the alpha q subfamily of guanine nucleotide-binding regulatory protein alpha subunits attenuate activation of phosphatidylinositol 4,5-bisphosphate hydrolysis by hormones.
The subfamily of guanine nucleotide-binding regulatory (G proteins) designated Gq has been shown to regulate the activity of phospholipase C by reconstitution. However, the role of these proteins in hormonal regulation of this activity has not been demonstrated. Two antisera were used in attempts to interrupt this pathway. Antiserum W082, developed against a peptide representing an internal sequence in alpha q, was specific for alpha q by immunoblots but did not recognize the native protein. Antiserum X384 was developed against a peptide representing the 12 amino acids of the common carboxyl termini of alpha q and alpha 11. It had a broader specificity for this subfamily of G protein alpha subunits and recognized the native proteins. Antiserum X384 specifically immunoprecipitated alpha q and its homologs from purified preparations and detergent extracts of membranes. Affinity-purified antibodies attenuated stimulation of phosphatidylinositide 4,5-bisphosphate hydrolysis by bradykinin, angiotensin, and histamine in membranes derived from NG108-15 cells, rat liver, and 1321N1 cells, respectively. Activation of the phospholipase C activity by guanosine 5'-3-O-(thio)triphosphate alone was also inhibited. Inclusion of the peptide to which the antisera were raised blocked the effect of the antibody. In contrast, affinity-purified W082, which did not recognize native proteins, did not alter regulation of phospholipase C. This indicates that the Gq family of signaling proteins can couple to several receptors and is responsible for the hormonal regulation of phospholipase C in these diverse systems. The further generality of this regulatory pathway remains to be established.
Gutowski
S
S
Department of Pharmacology, University of Texas Southwestern Medical Center, Dallas 75235.
Smrcka
A
A
Nowak
L
L
Wu
D G
DG
Simon
M
M
Sternweis
P C
PC
D000804:Angiotensin II / Q000494:pharmacology; D000818:Animals; D015153:Blotting, Western; D001920:Bradykinin / Q000494:pharmacology; D002460:Cell Line; D004591:Electrophoresis, Polyacrylamide Gel; D019204:GTP-Binding Proteins / Q000276:immunology*; D006728:Hormones / Q000378:metabolism*; D006868:Hydrolysis; D007106:Immune Sera; D019269:Phosphatidylinositol 4,5-Diphosphate; D043265:Phosphatidylinositol Diacylglycerol-Lyase; D018129:Phosphatidylinositol Phosphates / Q000378:metabolism*; D010727:Phosphoric Diester Hydrolases / Q000378:metabolism; D011233:Precipitin Tests; D051381:Rats
D006728:Hormones; D007106:Immune Sera; D019269:Phosphatidylinositol 4,5-Diphosphate; D018129:Phosphatidylinositol Phosphates; D000804:Angiotensin II; D010727:Phosphoric Diester Hydrolases; D019204:GTP-Binding Proteins; D043265:Phosphatidylinositol Diacylglycerol-Lyase; D001920:Bradykinin