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Molecular and Cellular Biology, February 2002, p. 801-815, Vol. 22, No. 3
0270-7306/01/$04.00+0     DOI: 10.1128/MCB.22.3.801-815.2002
Copyright © 2002, American Society for Microbiology. All Rights Reserved.

Removal of a Single -Tubulin Gene Intron Suppresses Cell Cycle Arrest Phenotypes of Splicing Factor Mutations in Saccharomyces cerevisiae

C. Geoffrey Burns,^1,2, Ryoma Ohi,^1,2, Sapna Mehta,^1,2 Eileen T. O’Toole,³ Mark Winey,⁴ Tyson A. Clark,⁵ Charles W. Sugnet,⁵ Manuel Ares, Jr.,⁵ and Kathleen L. Gould^1,2*

Howard Hughes Medical Institute,¹ Department of Cell Biology, Vanderbilt University School of Medicine, Nashville, Tennessee 37232,² Boulder Laboratory for 3D Fine Structure,³ Department of Molecular, Cellular, and Developmental Biology, University of Colorado—Boulder, Boulder, Colorado 80309-0347,⁴ Center for Molecular Biology of RNA, University of California, Santa Cruz, Santa Cruz, California 95064⁵

Received 18 September 2001/ Accepted 1 November 2001

Genetic and biochemical studies of Schizosaccharomyces pombe and Saccharomyces cerevisiae have identified gene products that play essential functions in both pre-mRNA splicing and cell cycle control. Among these are the conserved, Myb-related CDC5 (also known as Cef1p in S. cerevisiae) proteins. The mechanism by which loss of CDC5/Cef1p function causes both splicing and cell cycle defects has been unclear. Here we provide evidence that cell cycle arrest in a new temperature-sensitive CEF1 mutant, cef1-13, is an indirect consequence of defects in pre-mRNA splicing. Although cef1-13 cells harbor global defects in pre-mRNA splicing discovered through intron microarray analysis, inefficient splicing of the -tubulin-encoding TUB1 mRNA was considered as a potential cause of the cef1-13 cell cycle arrest because cef1-13 cells arrest uniformly at G₂/M with many hallmarks of a defective microtubule cytoskeleton. Consistent with this possibility, cef1-13 cells possess reduced levels of total TUB1 mRNA and -tubulin protein. Removing the intron from TUB1 in cef1-13 cells boosts TUB1 mRNA and -tubulin expression to near wild-type levels and restores microtubule stability in the cef1-13 mutant. As a result, cef1-13 tub1i cells progress through mitosis and their cell cycle arrest phenotype is alleviated. Removing the TUB1 intron from two other splicing mutants that arrest at G₂/M, prp17 and prp22-1 strains, permits nuclear division, but suppression of the cell cycle block is less efficient. Our data raise the possibility that although cell cycle arrest phenotypes in prp mutants can be explained by defects in pre-mRNA splicing, the transcript(s) whose inefficient splicing contributes to cell cycle arrest is likely to be prp mutant dependent.

Present address: Cardiovascular Research Center, Massachusetts General Hospital, Charlestown, MA 02129.

Present address: Department of Cell Biology, Harvard Medical School, Boston, MA 02115.

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