Mutational activation of the STE5 gene product bypasses the requirement for G protein beta and gamma subunits in the yeast pheromone response pathway.

Abstract

The STES gene encodes an essential element of the pheromone response pathway which is known to act either after the G subunit encoded by the STE4 gene or at the same step. Mutations in STES, designated STESHYP, that partially activate the pathway in the absence of pheromone were isolated. One allele (STE5HrYP2) indicating that the STES product acts after the receptor (STE2 product) and after the G protein ,B and 'y subunits (STE4 and STE18 products, respectively). However, the phenotypes of the STESHYP mutations were less pronounced in ste4 and stel8 mutants, suggesting that the STE5HYP generated signal partially depends on the proposed G., complex. The STESHYP alleles did not suppress ste7, stell, stel2, orfi*s3 kssl null mutants, consistent with previous findings that the STES product acts before the protein kinases encoded by STE7, STE]], FUS3, and KSSI and the transcription factor encoded by STE12. The mating defects of the ste2 deletion mutant and the temperature-sensitive ste4-3 mutant were also suppressed by overexpression of wild-type STE5.The slow-growth phenotype manifested by cells carrying STESHYP alleles was enhanced by the sst2-1 mutation; this effect was eliminated in ste4 mutants. These results provide the first evidence that the STES gene product performs its function after the G protein subunits.The pheromone response in the yeast Saccharomyces cerevisiae provides a model system for studying general features of peptide-hormone action (see references 59 and 75 for reviews). In this yeast, conjugation of haploid a and a cells yields a/a diploid cells. Conjugation requires the action of peptide pheromones: a cells secrete a-factor pheromone and respond to a-factor, whereas a cells secrete a-factor pheromone and respond to a-factor. The binding of the pheromones to specific receptors on the target cell initiates the mating program. Specific cellular responses to pheromones include the arrest of cell division in G1 (34) and the production of cellular factors required for cell aggregation, cell fusion, karyogamy, and pheromone desensitization (17,18,54,60,72,79). These responses are mediated, at least in part, by changes in the transcription of cellular genes. Pheromones also provide spatial information which allows cells to locate and choose specific mating partners (42,43,51).Previous genetic analyses of the pheromone response pathway identified a number of components required for signal transduction. The STE2 and STE3 genes encode the a-factor and a-factor receptors, respectively (4, 35, 46, 57,

The MAP kinase Fus3 associates with and phosphorylates the upstream signaling component Ste5.

“…Thus, the primary role for Ste5 as a nucleation site for the assembly of protein kinases involved in signal transduction is consistent with the previous epistasis analyses (MacKay 1983;Blinder et al 1989;Elion et al 1991a, b;Cairns et al 1992;Leberer et al 1992Leberer et al , 1993Stevenson et al 1992;Hasson et al 1994;Ramer and Davis 1993), if we assume that the enzymatic reactions, but not necessarily the physical associations, are in a linear series. The fact that Ste5 is so limiting for Fus3 activation may suggest Ste5 is in low abundance, consistent with a critical role as a potent activator.…”

“…Ste5 is thought to act at a single step between the Ste20 and Stell protein kinases on the basis of genetic analysis with hyperactive forms of Ste5 and Stell (Cairns et al 1992;Leberer et al 1992;Stevenson et al 1992;Hasson et al 1994); however, several observations suggest a more complex picture. First, hyperactive forms of Stel 1 do not fully suppress ste5 mutations (Stevenson et al 1992) and hyperactive forms of SteS only partially suppress G-protein mutants (Hasson et al 1993).…”

“…This ubiquitous pattern of suppression suggests that SteS function cannot be defined in simple linear terms. Therefore, more complicated models for the role of SteS in signal transduction, such as an interdependence with other signal transduction components (e.g., the G-protein, Stell, or Fus3), cannot be excluded (Elion et al 1991a;Stevenson et al 1992;Hasson, Blinder, Thomer, Jenness, 1994).…”

RGS Proteins and Signaling by Heterotrimeric G Proteins

“…First, certain mutations in the G␣ gene could bypass the requirement for a fully active SST2 gene (30,31). Second, the recovery-promoting effect of G␣ overexpression was more efficient if residual SST2 function was present (32,33). Third, dominant gain-of-function mutations in SST2 confer resistance to receptor-mediated activation of the pathway but not to activation by mutations that permit constitutive release of G␤␥ (33).…”

A cell‐density sensing factor regulates the lifetime of a chemoattractant‐induced Gα‐GTP conformation

“…There are proteins called Regulators of G protein Signaling (RGS) which appear to regulate the activity of Ga subunits in yeast, worms and mammals [33]. Although genetic analysis has placed RGS proteins upstream of the G protein they regulate [34,35], it is not known if they activate the GTPase activity of Ga subunits. We have thus found a potentially general way for one signal transduction pathway to regulate a second G protein-mediated signal transduction pathway by regulating the lifetime of the Ga-GTP state.…”