Altered G protein activity in a desensitization-resistant mutant of the Y1 adrenocortical tumor cell line.
Mutant isolates [designated desensitization resistant (DR)] from the Y1 mouse adrenocortical tumor cell line resist agonist-induced desensitization of adenylyl cyclase by preventing the uncoupling of receptors from their guanyl nucleotide-binding regulatory G proteins. In this study, we tested the hypothesis that an underlying G protein defect is associated with the DR phenotype. We found that the G protein reagent guanyl-5'-yl imidodiphosphate [Gpp(NH)p] shifted beta2-adrenergic receptors from a high affinity state to a low affinity state 4-fold more effectively in mutant DR cells than in parent Y1 cells. In the DR mutant, Gpp(NH)p was able to shift receptors to a low affinity state in the absence of NaCl, whereas the effect of Gpp(NH)p in parent Y1 cells was dependent upon the presence of NaCl. Moreover, these differences in sensitivity to Gpp(NH)p and NaCl were transferred to Gs alpha-deficient S49(CYC-) lymphoma cell membranes in G protein reconstitution assays. These observations suggested that the DR mutation was associated with altered activity of the stimulatory G protein, Gs. Cloning and sequence analysis demonstrated that Gs alpha transcripts in the DR mutant were normal, suggesting that another factor involved in guanyl nucleotide exchange is responsible for the altered G protein activity in DR mutant cells.
Colantonio
C M
CM
Banting and Best Department of Medical Research, University of Toronto, Ontario, Canada.
Kwan
W K
WK
Czerwinski
W
W
Mitchell
J
J
Schimmer
B P
BP
D000222:Adaptation, Physiological / Q000502:physiology*; D000306:Adrenal Cortex Neoplasms / Q000235:genetics* / Q000378:metabolism* / Q000473:pathology; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D018076:DNA, Complementary / Q000235:genetics; D019204:GTP-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D006165:Guanylyl Imidodiphosphate / Q000494:pharmacology; D008223:Lymphoma / Q000235:genetics / Q000473:pathology; D051379:Mice; D008969:Molecular Sequence Data; D009154:Mutation; D010641:Phenotype; D011943:Receptors, Adrenergic, beta / Q000187:drug effects / Q000378:metabolism; D012965:Sodium Chloride / Q000494:pharmacology; D014407:Tumor Cells, Cultured
D018076:DNA, Complementary; D011943:Receptors, Adrenergic, beta; D006165:Guanylyl Imidodiphosphate; D012965:Sodium Chloride; D019204:GTP-Binding Proteins