The prp2 gene of fission yeast has previously been shown to encode the large subunit of the splicing factor spU2AF. SpU2AF(59) is an evolutionarily conserved protein that has an arginine/serine-rich region and three RNA recognition motifs (RRMs). We have sequenced three temperature-sensitive alleles of prp2 and determined that the mutations result in single amino acid changes within one of the RRMs or between RRMs. All mutant alleles of prp2 have pre-mRNA splicing defects at the non-permissive temperature. Although the mutant strains are growth-arrested at 37 degrees C, they do not elongate like typical fission yeast cell cycle mutants. The DNA of the prp2(-) strains stains more intensely than a wild-type strain, suggesting that the chromatin may be condensed. Ultrastructural studies show differences in the mutant nuclei including a prominent distinction between the chromatin- and non-chromatin-enriched regions compared to the more homogenous wild-type nucleus. Two-hybrid assays indicate that some of the wild-type protein interactions are altered in the mutant strains. These results suggest that normal functioning of spU2AF(59) may be essential not only for pre-mRNA splicing but also for the maintenance of proper nuclear structure and normal cell cycle progression.