Minimal ectopic expression of the p58GTA protein kinase results in a provocative phenotype involving cell cycle delay, mitotic catastrophe, and decreased cell viability. In addition, this kinase is well conserved evolutionarily, ubiquitously expressed, and its genes map to a position on human chromosome 1 frequently deleted in the late stages of tumorigenesis. Here we report that the p58GTA protein kinase is a member of a larger subfamily of proteins. The mRNAs encoding these proteins are generated by alternative splicing from multiple duplicated genes. These isoforms range in size from 50 to 110 kDa. Divergence between the alternatively spliced isoforms is localized to the amino-terminal region of the molecule. The entire p58GTA open reading frame is conserved in most of these p58GTA isoforms. The predicted sequences of the larger isoforms encode bipartite nuclear localization signal sequences and extensive polyglutamic acid domains. Antibodies to the p58GTA isoform were used to confirm the presence of the alternatively spliced isoforms in different cell types as well as identify two additional isoforms that appear to arise from a separate gene(s). Cellular fractionation studies indicate that one of the isoforms is found only in the nucleus, and the remainder are found in both the cytoplasm and the nucleus. Expression and localization of some p58GTA isoforms suggest that they may have specialized cellular functions. Because of the large number of isoforms generated from multiple genes we propose naming these kinases PITSLRE alpha 1, alpha 2-1, alpha 2-2, alpha 2-3 alpha 2-4, beta 1, beta 2-1, and beta 2-2 based on the conserved sequence of the PSTAIRE box unique to p34cdc2 kinases and the gene from which they are transcribed.