Premium
Fission yeast Cut2 required for anaphase has two destruction boxes
Author(s) -
Funabiki Hironori,
Yamano Hiroyuki,
Nagao Koji,
Tanaka Hirofumi,
Yasuda Hideyo,
Hunt Tim,
Yanagida Mitsuhiro
Publication year - 1997
Publication title -
the embo journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 7.484
H-Index - 392
eISSN - 1460-2075
pISSN - 0261-4189
DOI - 10.1093/emboj/16.19.5977
Subject(s) - biology , anaphase , microbiology and biotechnology , metaphase , mitosis , schizosaccharomyces pombe , cyclin b , cdc20 , anaphase promoting complex , proteolysis , schizosaccharomyces , cyclin b1 , cell cycle , genetics , biochemistry , cyclin , yeast , saccharomyces cerevisiae , cyclin dependent kinase 1 , gene , chromosome , enzyme
The fission yeast Schizosaccharomyces pombe cut2 + gene is essential for sister chromatid separation. Cut2 protein, which locates in the interphase nucleus and along the metaphase spindle, disappears in anaphase with the same timing as mitotic cyclin destruction. This proteolysis depends on the APC (Anaphase‐Promoting Complex)–cyclosome which contains ubiquitin ligase activity. The N‐terminus of Cut2 contains two stretches similar to the mitotic cyclin destruction box. We show that both sequences (33RAPLGSTKQ and 52RTVLGGKST) serve as destruction boxes and are required for in vitro polyubiquitination and proteolysis. Cut2 with doubly mutated destruction boxes inhibits anaphase, whereas Cut2 with singly mutated boxes can suppress cut2 mutations. Strong expression of the N‐terminal 73 residues containing the destruction boxes leads to the accumulation of endogenous cyclin and Cut2, and arrests cells in metaphase, whereas the same fragment with the mutated boxes does not. Cut2 proteolysis occurs in vitro using Xenopus mitotic extracts in the presence of functional destruction boxes. Furthermore, Cut2 is polyubiquitinated in an in vitro system using HeLa extracts, and this polyubiquitination requires the destruction boxes.