Probes derived from cDNAs encoding isozymes of rat protein kinase C (PKC) were used to screen the genome of the budding yeast Saccharomyces cerevisiae. We reported previously the isolation of the yeast PKC1 gene, a homolog of the alpha, beta, and gamma subspecies of mammalian PKC. Here we report the isolation and genetic characterization of a pair of previously described genes (YPK1 and YPK2) which are predicted to encode protein kinases that share 90% amino acid identity with each other and 44-46% identity with various isozymes of PKC throughout their putative catalytic domains. Deletion of YPK2 resulted in no apparent phenotypic defect, but loss of YPK1 resulted in slow growth. Cells deleted for both YPK1 and YPK2 were defective in vegetative growth, indicating that the protein kinases predicted to be encoded by these genes are functionally overlapping and play an essential role in the proliferation of yeast cells. The YPK1 gene was mapped to the left arm of chromosome XI and YPK2 was mapped to the right arm of chromosome XIII.