In yeastSaccharomyces cerevisiae, the immunosuppressant rapamycin inhibits the TORC1 kinase causing rapid alteration in gene expression and leading to G1arrest. We recently reported the isolation and characterization from the histone mutant collection of a histone H2B R95A mutant that displays resistance to rapamycin. This mutant is defective in the expression of several genes belonging to the pheromone response pathway includingSTE5encoding a scaffold protein that promotes the activation of downstream MAP kinases. Cells lacking Ste5 cannot arrest the cell cycle in response to rapamycin and as a consequence exhibit similar resistance to rapamycin as the H2B R95A mutant. Herein, we show that the H2B R95A mutation weakens the association of H2B with Spt16 a component of the FACT complex (FAcilitatesChromatinTranscription), and an essential factor that interacts with the histone H2A-H2B dimer to promote transcription and preserve chromatin integrity. From a collection ofspt16mutants,spt16 E857Kandspt16-11showed striking sensitivity to rapamycin as compared to the parent strain.spt16 E857Kandspt16-11expressed distinct forms of Ste5, while a suppressor mutation H2B A84D of thespt16-11mutant prevents the expression of Ste5 and confers marked resistance to rapamycin. We interpret these findings to suggest that the Arg95 residue of histone H2B is required to recruit Spt16 to maintain the expression ofSTE5, which performs a role to arrest cells in the G1phase in response to rapamycin.