The Cdc42p GTPase is involved in many aspects of growth and cell-cycle regulation, including actin cytoskeletal rearrangements and activation of signal transduction pathways. To further investigate these functions, genetic interactions were examined between Schizosaccharomyces pombe Cdc42p, its effectors Pak1p and Pak2p, and the Mkh1p-Pek1p-Spm1p signal transduction pathway, which functions in cytokinesis and cell division. Expression of a truncated version of Pak2p lacking its N-terminal autoinhibitory domain led to a growth defect that was suppressed by deltamkh1 and deltaspm1 null mutations and an elongated cell phenotype indicative of a cell division defect that was suppressed by the deltamkh1 mutation. In addition, expression of the constitutively activated cdc42G12V mutant allele led to a growth defect that was rescued by the deltapak2 and deltamkh1 mutations. The deltapak2 mutation did not suppress the growth defect conferred by plasmid expression of Mkh1p, suggesting that Pak2p functions upstream of Mkh1p in this pathway. A two-hybrid protein interaction was observed between Pak2p and Mkh1p, but not between Pak1p and Mkh1p. These results are consistent with Cdc42p interacting with Pak2p to signal through the Mkh1p-Pek1p-Spm1p pathway.