Functional analysis of the P box, a domain in cyclin B required for the activation of Cdc25.
Cyclin B-cdc2 complexes are kept inactive by inhibitory phosphorylations on Thr-14 and Tyr-15 of cdc2 until they are dephosphorylated at the end of G2 by the phosphatase cdc25. Recent work has suggested that a small region of cyclin B, which we call the P box, may contribute part of a phosphatase-activating domain to cdc25. Individual conservative substitutions at three invariant residues within the P box yield mutant cyclin B proteins that bind cdc2 in vitro and then show the predicted cell cycle arrest, with cdc25 remaining in the low activity interphase form and cyclin B-cdc2 complexes remaining phosphorylated and inactive. While the low activity interphase form of cdc25 cannot act on cdc2 complexed with a mutant P box cyclin, the high activity M phase form of cdc25 can. These results demonstrate that the P box domain of cyclin B is required for cdc25 activation and support a two-step mechanism for the cdc25-dependent activation of cyclin B-cdc2.
Zheng
X F
XF
Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, Massachusetts 02115.
Ruderman
J V
JV
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D003001:Cloning, Molecular; D003062:Codon; D016213:Cyclins / Q000235:genetics / Q000378:metabolism*; D004926:Escherichia coli / Q000235:genetics; D005260:Female; D006801:Humans; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009838:Oligodeoxyribonucleotides; D009865:Oocytes / Q000378:metabolism; D010766:Phosphorylation; D011506:Proteins / Q000378:metabolism*; D011994:Recombinant Proteins / Q000378:metabolism; D017384:Sequence Deletion; D017386:Sequence Homology, Amino Acid; D013912:Threonine; D014443:Tyrosine; D014981:Xenopus; D020687:cdc25 Phosphatases
D003062:Codon; D016213:Cyclins; D009838:Oligodeoxyribonucleotides; D011506:Proteins; D011994:Recombinant Proteins; D013912:Threonine; D014443:Tyrosine; D020687:cdc25 Phosphatases