Several fission yeast temperature-sensitive mutants defective in pre-mRNA processing (prp^- mutants) at the nonpermissive temperature have been identified. Here, the prp2^+ gene has been cloned by its ability to complement the temperature-sensitive growth defect of a prp2^- mutant. The gene also corrects the pre-mRNA splicing defect of prp2^- mutants and encodes a 59-kilodalton polypeptide (PRP2). A molecular characterization indicates that PRP2 is a previously uncharacterized yeast splicing factor with extensive similarity to the mammalian splicing factor U2AF65. Thus, this study provides evidence that a U2AF homolog participates in RNA processing in vivo.