A C-terminal mutant of the G protein beta subunit deficient in the activation of phospholipase C-beta.
The molecular mechanism by which the G protein betagamma complex modulates multiple mammalian effector pathways is unknown. Homolog-scanning mutagenesis of the G protein beta subunit was employed to identify residues critical for the activation of phospholipase C-beta2 (PLC-beta2). A series of chimeras was made by introducing small segments of the Dictyostelium beta subunit into a background of mammalian beta1 and tested in COS cell cotransfection assays for their ability to activate PLC-beta2 and assemble with mammalian gamma2. A chimera that contained four Dictyostelium beta substitutions within the C-terminal 14 residues was unable to activate PLC-beta2 when cotransfected with gamma, despite its demonstrable expression in a gamma-dependent manner. Cotransfection of the mutant blocked m2 muscarinic receptor activation of PLC by a pertussis toxin-sensitive pathway. This C-terminal mutant retained the ability, however, to stimulate the mitogen-activated protein kinase pathway. These results imply that activation of different betagamma-responsive effectors is mediated by distinct domains.
Zhang
S
S
Metabolic Diseases Branch, NIDDK, National Institutes of Health, Bethesda, Maryland 20892, USA.
Coso
O A
OA
Collins
R
R
Gutkind
J S
JS
Simonds
W F
WF
D000595:Amino Acid Sequence; D000818:Animals; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000378:metabolism; D002417:Cattle; D002478:Cells, Cultured; D002522:Chlorocebus aethiops; D004023:Dictyostelium / Q000737:chemistry; D004789:Enzyme Activation; D019204:GTP-Binding Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D006801:Humans; D007527:Isoenzymes / Q000378:metabolism*; D019950:Mitogen-Activated Protein Kinase 1; D008969:Molecular Sequence Data; D054799:Phospholipase C beta; D011993:Recombinant Fusion Proteins; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D015398:Signal Transduction; D013329:Structure-Activity Relationship; D014162:Transfection; D010738:Type C Phospholipases / Q000378:metabolism*
D007527:Isoenzymes; D011993:Recombinant Fusion Proteins; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D019950:Mitogen-Activated Protein Kinase 1; D010738:Type C Phospholipases; C518033:PLCB1 protein, human; C518036:PLCB2 protein, human; D054799:Phospholipase C beta; D019204:GTP-Binding Proteins