Expression of the BAR1 gene in Saccharomyces cerevisiae: Induction by the α mating pheromone of an activity associated with a secreted protein

Abstract

We have demonstrated and partially characterized the genetic control and pheromonal regulation of a soluble activity, produced only by mating-type a cells, that inhibits the action of the α mating pheromone, α-factor, on mating-type a cells. This activity was found to be associated with a heat-stable protein and to be secreted by MATaBAR1, matα2 BAR1, and matα1 matα2 BAR1 strains, but not by MATα BAR1, MAta/MATα BAR1, matα1 BAR1, or MATa bar1 strains, demonstrating that it is under the control of both the MATα2 and the BAR1 genes. Secretion of this activity was also found to be stimulates to as much as five times the basal level by exposure of the cells to α-factor. This stimulation was maximal after 6 h at a pheromone concentration of approximately 2 U/ml. An assay for this activity was developed by using a refined, quantitative assay for α-factor. The pheromone activity of samples added to wells in an agar plate was related to the size of the halo of growth inhibition produced in a lawn of mutant cells that are abnormally sensitive. The α-factor-inhibiting activity was related to a reduction of the halo size when active samples were added to the lawn. Although the assay for α-factor was found to be relatively insensitive to pH over a range of several units, the α-factor-inhibiting activity displayed a sharp pH optimum at approximately 6.5. The properties of this activity have important implications concerning the role of the BAR1 gene product in recovery of mating-type a cells from cell division arrest by α-factor.