Loss of RanGEF/Pim1 activity abolishes the orchestration of Ran-mediated mitotic cellular events in S. pombe.
RCC1, a conserved chromosomal protein with a seven-bladed propeller is a GDP/GTP nucleotide exchange factor for RanGTPase that mediates various cellular events. We isolated 16 temperature-sensitive (ts) mutants of S. pombeRCC1-homolog, pim1+, by error-prone PCR. Five pim1(ts) mutants had a single mutation. The obtained pim1(ts) mutations and previously reported mutations were localized on similar sites in seven RCC1 repeats. Those mutations resulted in a reduced binding of Pim1 with Spi1. All pim1(ts) mutants showed a defect in nucleocytoplasmic protein transports, whereas the majority of them showed a normal mRNA export. In all pim1(ts) examined, chromosomal DNA replication was completed. However, mitotic spindle formation was abrogated, the septum was formed being uncoupled with nuclear division and abnormally widened, thus resulting in chromosomal DNA mis-segregation and the accumulation of enucleated cells. As a result, a defect of RanGEF/Pim1 abolished the orchestration of sequential mitotic events, spindle formation, septation and cytokinesis that are essential to produce two identical daughter cells.
Hirose
Eiji
E
Department of Molecular Biology, Graduate School of Medical Science, Kyushu University, 3-1-1, Maidashi, Fukuoka 812-8582, Japan.
Mukai
Mari
M
Shimada
Atsushi
A
Nishitani
Hideo
H
Shibata
Yosaburo
Y
Nishimoto
Takeharu
T
D021581:Active Transport, Cell Nucleus; D000595:Amino Acid Sequence; D000818:Animals; D020662:Guanine Nucleotide Exchange Factors / Q000235:genetics* / Q000378:metabolism; D006358:Hot Temperature; D006801:Humans; D008938:Mitosis / Q000502:physiology*; D008969:Molecular Sequence Data; D009154:Mutation; D021381:Protein Transport; D012333:RNA, Messenger / Q000096:biosynthesis; D012568:Schizosaccharomyces / Q000166:cytology* / Q000235:genetics* / Q000378:metabolism; D029702:Schizosaccharomyces pombe Proteins / Q000235:genetics* / Q000378:metabolism; D016415:Sequence Alignment; D020931:ran GTP-Binding Protein / Q000378:metabolism*
D020662:Guanine Nucleotide Exchange Factors; D012333:RNA, Messenger; D029702:Schizosaccharomyces pombe Proteins; C490057:pim1 protein, S pombe; D020931:ran GTP-Binding Protein