Abstract: Genes CDC24, CDC42, and CDC43 are required for the establishment of cell polarity and the localization of secretion in Saccharomyces cerevisiae; mutants defective in these genes fail to form buds and display isotropic expansion of the cell surface. To identify other genes that may be involved in these processes, we screened yeast genomic DNA libraries for heterologous genes that, when overexpressed from a plasmid, can suppressa temperature-sensitive cdc24 mutation. We identified four such genes. One of these proved to be CDC42, which has previously been shown to be a member of the rho (ras-homologous) family of genes, and a second is a newly identified ras-related gene that we named RSR1. RSR1 maps between CDC62 and ADE3 on the right arm of chromosome VII; its predicted product is approximately 50% identical to other proteins in the ras family. Deletion of RSR1 is nonlethal but disrupts the normal pattern of bud site selection. Although both CDC42 and RSR1 can suppress cdc24 and both appear to encode GTP-binding proteins, these genes do not themselves appear to be functionally interchangeable. However, one of the other genes that was isolated by virtue of its ability to suppress cdc24 can also suppress cdc42. This gene, named MSB1, maps between ADE9 and HIS3 on the right arm of chromosome XV.