Identification of the lesion in the stimulatory GTP‐binding protein of the uncoupled S49 lymphoma

Rall, T B; Harris, Bruce A.

doi:10.1016/0014-5793(87)80486-6

Cited by 62 publications

(23 citation statements)

References 24 publications

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“…The SCG1 Pr°a67 defects were somewhat more severe in a cells than in a cells (Table 2). This phenotype is similar to the effect of the analogous mutation in G~xs (Haga et al 1977;Rail and Harris 1987;Sullivan et al 1987), that is, both result in reduced activation of the respective pathways. The SCG1 e~°a6~ mutant showed similar effects as the SCG1Pr°a67 mutant in a cells {Table 2; Figs.…”

Section: Mutations Resulting M Reduced Mating Are Consistent With Defsupporting

confidence: 53%

“…In addition, binding of Gat to rhodopsin can be competed by peptides corresponding to regions near the carboxyl terminus {Hamm et al 1988). Rat Gas unc mutants are defective in the interaction with the B-adrenergic receptor {Haga et al 1977); this defect results from a single arginine to proline substitution 6 amino acids from the carboxyl terminus of Gas (Rail and Harris 1987;Sullivan et al 1987).…”

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confidence: 99%

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The carboxyl terminus of Scg1, the G alpha subunit involved in yeast mating, is implicated in interactions with the pheromone receptors.

Hirsch

Dietzel²,

Kurjan³

1991

Genes Dev.

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The carboxyl termini of ¢x subunits of mammalian G proteins have been implicated in receptor interactions. We have used a genetic analysis to test such a role for the carboxyl terminus of Scgl, the ot subunit involved in the yeast pheromone response pathway. A 22-amino-acid truncation (scgl A~t'4sl) resulted in defects in growth and cellular morphology. This phenotype is similar to the null phenotype and represents constitutive activation of the pheromone response pathway; it could result from various effects, including protein instability or constitutive guanine nucleotide exchange, as reported for some altered mammalian Gas constructs. A 5-amino-acid truncation (SCG1 °ch't68) resulted in pheromone response and mating defects in both a and a cells, which is consistent with defects in interactions with the pheromone receptors.Lysine-to-proline mutations near the carboxyl terminus (SCG1 pr°467 and SCG1 J'r°46s) resulted in pheromone response and mating defects, the severity of which differed in a and o~ cells. This differential effect in the two mating types suggests that the specificity for the interactions with the two pheromone receptors may involve different residues of the Scgl carboxyl terminus. Mutations leading to constitutive activation of the pathway were recessive, whereas mutations that result in decreased pheromone response and mating were partially dominant. These relationships are consistent with the model for the mechanism of action of the G protein subunits in the pheromone response pathway and indicate the importance of the stoichiometry of components of this system. Signal transduction mediated by regulatory G proteins in mammalian systems involves the interaction of a ligand-receptor complex with an intracellular G protein composed of o~, 13, and ~/subunits (for review, see Gilman 1987). Before activation, the a subunit has GDP bound and is complexed with 13~/. After receptor activation, GDP is replaced by GTP, and the a subunit dissociates from the 13~ complex. The intrinsic GTPase activity of the a subunit hydrolyzes the bound GTP, regenerating the GDP-bound form of the a subunit, which reassociates with 13~/to complete the cycle.A similar system in yeast is involved in the response of cells to peptide pheromones and is required for mating (for review, see Cross et al. 1988}. Cells of a and a mating type secrete a-factor and o~-factor, respectively, and respond to the pheromone secreted by the opposite mating type. Response to pheromone causes arrest of cells in the Present addresses: tRockefeller University,

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Section: Mutations Resulting M Reduced Mating Are Consistent With Defsupporting

confidence: 53%

mentioning

confidence: 99%

The carboxyl terminus of Scg1, the G alpha subunit involved in yeast mating, is implicated in interactions with the pheromone receptors.

Hirsch

Dietzel²,

Kurjan³

1991

Genes Dev.

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“…The inhibition seen to direct activation of Gs by fluoride was unexpected, suggesting antibody effects are more generalized than other 'uncoupling' lesions involving the extreme G~ C-terminus, such as the Arg 3s9 to Pro mutation in unc $49 Gsa [6,7] or ADP-ribosylation by pertussis toxin of ai and ao [1][2][3]. In the case of the latter modifications, AIF~ or GTP [7]S can still activate the G-protein involved as demonstrated by subunit dissociation, effector activation or stabilization to tryptic digestion.…”

Section: Discussionmentioning

confidence: 99%

“…Furthermore G-ct subunits which are functionall3/ uncoupled from receptors by pertussis toxin undergo a mono-ADP-ribosylation at a cysteine residue four amino acids from the carboxylterminus [1][2][3]. More recently, a point mutation was identified in the genome of the unc variant of $49 mouse lymphoma cells [6,7] which results in a Pro for Arg substitution six amino acids from the carboxyl-terminus of ces, and accounts for the uncoupling of receptor from Gs characteristic of this phenotype [8].…”

Section: Introductionmentioning

confidence: 99%

Receptor and effector interactions of G_s Functional studies with antibodies to the α_s carboxyl‐terminal decapeptide

et al. 1989

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Antibodies generated to a synthetic decapeptide, RMHLRQYELL, representing the carboxyl-terminus of Gs-ct have been characterized in immunoblots and functional studies. This antibody, designated RM, reacts exclusively with a doublet of proteins of 52 and 45 kDa in immunoblots of bovine brain and wild-type $49 murine lymphoma cell membranes. No such reactivity is seen in membranes from cyc-$49 cells, which lack Gs. RM blocks receptor-mediated activation of Gs and adenylyl cyclase in membranes from wild-type $49 cells. RM could also immunoprecipitate adenylyl cyclase activity in detergent extracts from GTP[),]S-or fluoride-preactivated bovine brain membranes; thus binding of cts to effector and carboxyl-terminal antibody was mutually compatible. Such experiments provide an approach for the elucidation of functionally relevant interactions of G-proteins with receptors and effectors in the membrane.

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“…1) (22, 44, 46). Most G ␣ mutations that result in receptor coupling defects cluster in G ␣ C termini (19,20,42,43,45,49,55).In the budding yeast Saccharomyces cerevisiae, a G proteinreceptor system is involved in the response of haploid cells to mating pheromones (reviewed in reference 30). The two haploid cell types, MATa and MAT␣, secrete the peptide pheromones a-factor and ␣-factor, respectively, which bind to mating type-specific receptors; the a-factor receptor Ste3p is expressed by MAT␣ cells, and the ␣-factor receptor Ste2p is expressed by MATa cells.…”

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confidence: 99%

Analysis of the Receptor Binding Domain of Gpa1p, the G_α Subunit Involved in the Yeast Pheromone Response Pathway

Kallal

Kurjan

1997

Molecular and Cellular Biology

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The Saccharomyces cerevisiae G protein ␣ subunit Gpa1p is involved in the response of both MATa and MAT␣ cells to pheromone. We mutagenized the GPA1 C terminus to characterize the receptor-interacting domain and to investigate the specificity of the interactions with the a-and ␣-factor receptors. The results are discussed with respect to a structural model of the Gpa1p C terminus that was based on the crystal structure of bovine transducin. Some mutants showed phenotypes different than the pheromone response and mating defects expected for mutations that affect receptor interactions, and therefore the mutations may affect other aspects of Gpa1p function. Most of the mutations that resulted in pheromone response and mating defects had similar effects in MATa and MAT␣ cells, suggesting that they affect the interactions with both receptors. Overexpression of the pheromone receptors increased the mating of some of the mutants tested but not the wild-type strain, consistent with defects in mutant Gpa1p-receptor interactions. The regions identified by the matingdefective mutants correlated well with the regions of mammalian G ␣ subunits implicated in receptor interactions. The strongest mating type-specific effects were seen for mutations to proline and a mutation of a glycine residue predicted to form a C-terminal ␤ turn. The analogous ␤ turn in mammalian G ␣ subunits undergoes a conformational change upon receptor interaction. We propose that the conformation of this region of Gpa1p differs during the interactions with the a-and ␣-factor receptors and that these mating type-specific mutations preclude the orientation necessary for interaction with one of the two receptors.G protein-mediated signaling involves heterotrimeric proteins that consist of ␣, ␤, and ␥ subunits. G proteins couple to membrane receptors of the rhodopsin/␤-adrenergic family of seven transmembrane-helix receptors (reviewed in reference 9). In the absence of a stimulus, the G protein ␣ subunit is bound to GDP and is complexed with the ␤␥ dimer. When a G protein-linked receptor is stimulated, the G ␣ subunit exchanges GTP for GDP and dissociates from G ␤␥ . After dissociation, G ␣ and/or G ␤␥ interact with targets that are effectors of cellular response pathways.Biochemical and genetic studies in mammalian systems have indicated that G ␣ C termini are involved in receptor interactions. Pertussis toxin modification of a cysteine residue 4 amino acids from the C terminus of G ␣t (transducin) results in reduced affinity for rhodopsin (58). Antibodies against C-terminal peptides of G ␣t and G ␣i2 block the interactions with rhodopsin and the ␣2-adrenergic receptor, respectively (6, 23, 51), and peptides corresponding to the C termini of G ␣t and G ␣s compete for binding to rhodopsin and the ␤-adrenergic receptor, respectively ( Fig. 1) (22, 44, 46). Most G ␣ mutations that result in receptor coupling defects cluster in G ␣ C termini (19,20,42,43,45,49,55).In the budding yeast Saccharomyces cerevisiae, a G proteinreceptor system is involved in the resp...

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Identification of the lesion in the stimulatory GTP‐binding protein of the uncoupled S49 lymphoma

Cited by 62 publications

References 24 publications

The carboxyl terminus of Scg1, the G alpha subunit involved in yeast mating, is implicated in interactions with the pheromone receptors.

The carboxyl terminus of Scg1, the G alpha subunit involved in yeast mating, is implicated in interactions with the pheromone receptors.

Receptor and effector interactions of G_s Functional studies with antibodies to the α_s carboxyl‐terminal decapeptide

Analysis of the Receptor Binding Domain of Gpa1p, the G_α Subunit Involved in the Yeast Pheromone Response Pathway

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Identification of the lesion in the stimulatory GTP‐binding protein of the uncoupled S49 lymphoma

Cited by 62 publications

References 24 publications

The carboxyl terminus of Scg1, the G alpha subunit involved in yeast mating, is implicated in interactions with the pheromone receptors.

The carboxyl terminus of Scg1, the G alpha subunit involved in yeast mating, is implicated in interactions with the pheromone receptors.

Receptor and effector interactions of Gs Functional studies with antibodies to the αs carboxyl‐terminal decapeptide

Analysis of the Receptor Binding Domain of Gpa1p, the Gα Subunit Involved in the Yeast Pheromone Response Pathway

Contact Info

Product

Resources

About

Receptor and effector interactions of G_s Functional studies with antibodies to the α_s carboxyl‐terminal decapeptide

Analysis of the Receptor Binding Domain of Gpa1p, the G_α Subunit Involved in the Yeast Pheromone Response Pathway