Yeast Lrg1p acts as a specialized RhoGAP regulating 1,3-beta-glucan synthesis

Selection of an extragenic suppressor of fks1-1154 Deltafks2, mutations in the catalytic subunits of yeast 1,3-beta-glucan synthase (GS) conferring temperature-sensitivity, led to the LRG1 gene, which was originally identified as a LIM-RhoGAP homologous gene. Mutations in the LRG1 gene restore impaired 1,3-beta-glucan synthesis in the fks1-1154 Deltafks2 mutant as well as that in rho1-2, a temperature-sensitive mutant of Rho-type GTPase that functions as a regulatory subunit of GS. Two-hybrid analyses of Lrg1p, which contains a sequence conserved among Rho GTPase-activating proteins (GAPs), revealed its specific interactions with the active form of Rho1p. Among eight potential yeast RhoGAPs, Lrg1p is the only member that negatively regulates GS activity: mutations in the rest of GAPs, including bem2, Deltabem3, Deltasac7, Deltabag7, Deltarga1, Deltarga2 and Deltargd1, do not suppress impairment of 1,3-beta-glucan synthesis. Analyses of Mpk1p phosphorylation revealed the inability of Lrg1p to regulate the Pkc1p-MAP kinase cascade, a distinct Rho1p-regulating signalling pathway known to be affected by the GAPs, Bem2p and Sac7p. Thus, different groups of Rho1p GAPs control the activity of different Rho1p-effector proteins.