Fission yeast APC/cyclosome subunits, Cut20/Apc4 and Cut23/Apc8, in regulating metaphase-anaphase progression and cellular stress responses.
BACKGROUND
The 20S cyclosome/APC complex promotes metaphase-anaphase transition by ubiquitinating its specific substrates such as mitotic cyclins and anaphase inhibitor Cut2/Pds1/securin. The complex has been shown to contain more than 10 proteins in budding yeast and frog. In fission yeast, however, only five (Cut4, Cut9, Nuc2, Apc10, Hcn1) have been identified.


RESULTS
More than five hundred temperature-sensitive mutants were screened for identifying those defective in mitotic anaphase. Fifty-five showed the cut (cell untimely torn) phenotype or metaphase-arrest phenotypes, 27 of them locating at new loci. Their extracts were run in sucrose gradient centrifugation, and four showed alterations in the sedimentation profiles. The gene products of cut20+ and cut23+ were thus identified. Phenotypes of cut20-100 mutant highly resemble cut4-533 in many ways: they are hypersensitive to canavanine and CdCl2, and suppressed by PKA-inactivating regulators, cAMP-dependent phosphodiesterase and PKA regulatory subunits. Cut20 interacts closely with Cut4 in the assembly process of cyclosome. But cut20 mutant differs from cut4, as a novel gene stw1+ suppresses cut20 mutant but not cut4. cut23-194 mutant cells are sterile and blocked at metaphase, but does not show sensitivity to the stress and cAMP. TPR repeat-containing Cut23 may not be the stable component of APC/cyclosome, and its level significantly fluctuates during cell cycle. Cut23 may be ubiquitinated and degraded in a cell cycle dependent fashion.


CONCLUSIONS
We identified two new subunits of fission yeast cyclosome/APC complex. Our observations indicate that cyclosome components are divided into several subgroups with distinctly different roles.
Yamashita
Y M
YM
CREST Research Project, Department of Biophysics, Graduate School of Science, Japan.
Nakaseko
Y
Y
Kumada
K
K
Nakagawa
T
T
Yanagida
M
M
D025601:Adenomatous Polyposis Coli Protein; D000595:Amino Acid Sequence; D000705:Anaphase; D064173:Anaphase-Promoting Complex-Cyclosome; D002104:Cadmium / Q000494:pharmacology; D002172:Canavanine / Q000494:pharmacology; D002453:Cell Cycle; D018797:Cell Cycle Proteins / Q000235:genetics; D017124:Conserved Sequence; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism; D003598:Cytoskeletal Proteins / Q000235:genetics*; D005656:Fungal Proteins / Q000235:genetics; D008025:Ligases / Q000235:genetics*; D008677:Metaphase; D008957:Models, Genetic; D008969:Molecular Sequence Data; D010641:Phenotype; D010957:Plasmids; D029701:Saccharomyces cerevisiae Proteins; D012568:Schizosaccharomyces / Q000235:genetics*; D013489:Suppression, Genetic; D013696:Temperature; D013997:Time Factors; D043743:Ubiquitin-Protein Ligase Complexes; D044767:Ubiquitin-Protein Ligases; D014452:Ubiquitins / Q000378:metabolism
D025601:Adenomatous Polyposis Coli Protein; D018797:Cell Cycle Proteins; D003598:Cytoskeletal Proteins; D005656:Fungal Proteins; D029701:Saccharomyces cerevisiae Proteins; D014452:Ubiquitins; D002104:Cadmium; D002172:Canavanine; D043743:Ubiquitin-Protein Ligase Complexes; D064173:Anaphase-Promoting Complex-Cyclosome; D044767:Ubiquitin-Protein Ligases; D017868:Cyclic AMP-Dependent Protein Kinases; D008025:Ligases