Mutations in the large subunit of U2AF disrupt pre-mRNA splicing, cell cycle progression and nuclear structure.
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.
Beales
M
M
Department of Cellular and Molecular Pharmacology, Finch University of Health Sciences/The Chicago Medical School, 3333 Green Bay Road, North Chicago, IL 60064, USA.
Flay
N
N
McKinney
R
R
Habara
Y
Y
Ohshima
Y
Y
Tani
T
T
Potashkin
J
J
D000483:Alleles; D002453:Cell Cycle / Q000502:physiology; D002467:Cell Nucleus / Q000648:ultrastructure; D053487:DEAD-box RNA Helicases; D005656:Fungal Proteins / Q000235:genetics / Q000378:metabolism; D006918:Hydroxyurea / Q000494:pharmacology; D009154:Mutation; D009687:Nuclear Proteins; D012322:RNA Precursors / Q000378:metabolism*; D012326:RNA Splicing; D012331:RNA, Fungal / Q000235:genetics / Q000378:metabolism; D012261:Ribonucleoproteins / Q000235:genetics* / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins; D012568:Schizosaccharomyces / Q000166:cytology / Q000235:genetics* / Q000254:growth & development / Q000378:metabolism; D000072668:Splicing Factor U2AF; D013696:Temperature; D020798:Two-Hybrid System Techniques
D005656:Fungal Proteins; D009687:Nuclear Proteins; D012322:RNA Precursors; D012331:RNA, Fungal; D012261:Ribonucleoproteins; D029701:Saccharomyces cerevisiae Proteins; D000072668:Splicing Factor U2AF; C066490:PRP2 protein, S cerevisiae; D053487:DEAD-box RNA Helicases; D006918:Hydroxyurea