title: Binding of 14-3-3beta but not 14-3-3sigma controls the cytoplasmic localization of CDC25B: binding site preferences of 14-3-3 subtypes and the subcellular localization of CDC25B.
abstract: The dual specificity phosphatase CDC25B positively controls the G2-M transition by activating CDK1/cyclin B. The binding of 14-3-3 to CDC25B has been shown to regulate the subcellular redistribution of CDC25B from the nucleus to the cytoplasm and may be correlated with the G2 checkpoint. We used a FLAG-tagged version of CDC25B to study the differences among the binding sites for the 14-3-3 subtypes, 14-3-3beta, 14-3-3epsilon and 14-3-3sigma, and the relationship between subtype binding and the subcellular localization of CDC25B. All three subtypes were found to bind to CDC25B. Site-directed mutagenesis studies revealed that 14-3-3beta bound exclusively near serine-309 of CDC25B1, which is within a potential consensus motif for 14-3-3 binding. By contrast, 14-3-3sigma bound preferentially to a site around serine-216, and the presence of serine-137 and -309 enhanced the binding. In addition to these binding-site differences, we found that the binding of 14-3-3beta drove CDC25B to the cytoplasm and that mutation of serine-309 to alanine completely abolished the cytoplasmic localization of CDC25B. However, co-expression of 14-3-3sigma and CDC25B did not affect the subcellular localization of CDC25B. Furthermore, serine-309 of CDC25B was sufficient to produce its cytoplasmic distribution with co-expression of 14-3-3beta, even when other putative 14-3-3 binding sites were mutated. 14-3-3epsilon resembled 14-3-3beta with regard to its binding to CDC25B and the control of CDC25B subcellular localization. The results of the present study indicate that two 14-3-3 subtypes can control the subcellular localization of CDC25B by binding to a specific site and that 14-3-3sigma has effects on CDC25B other than the control of its subcellular localization.
authors.affiliation: Division of Life Science, Graduate School of Natural Science and Technology, Kanazawa University, Kakuma-machi, Kanazawa, 920-1192, Japan.
authors.forename: Sanae
authors.initials: S
authors.lastname: Uchida
authors.forename: Akiko
authors.initials: A
authors.lastname: Kuma
authors.forename: Motoaki
authors.initials: M
authors.lastname: Ohtsubo
authors.forename: Mari
authors.initials: M
authors.lastname: Shimura
authors.forename: Masato
authors.initials: M
authors.lastname: Hirata
authors.forename: Hitoshi
authors.initials: H
authors.lastname: Nakagama
authors.forename: Tsukasa
authors.initials: T
authors.lastname: Matsunaga
authors.forename: Yukihito
authors.initials: Y
authors.lastname: Ishizaka
authors.forename: Katsumi
authors.initials: K
authors.lastname: Yamashita
mesh_terms: D048948:14-3-3 Proteins / Q000235:genetics / Q000378:metabolism*; D001665:Binding Sites; D014408:Biomarkers, Tumor / Q000235:genetics / Q000378:metabolism*; D018797:Cell Cycle Proteins / Q000378:metabolism*; D002467:Cell Nucleus / Q000378:metabolism; D002478:Cells, Cultured; D005092:Exonucleases / Q000235:genetics / Q000378:metabolism*; D005095:Exoribonucleases; D006801:Humans; D016297:Mutagenesis, Site-Directed; D009363:Neoplasm Proteins / Q000235:genetics / Q000378:metabolism*; D011485:Protein Binding; D021381:Protein Transport; D020687:cdc25 Phosphatases / Q000378:metabolism*
chemical_list: D048948:14-3-3 Proteins; D014408:Biomarkers, Tumor; D018797:Cell Cycle Proteins; D009363:Neoplasm Proteins; C482982:YWHAB protein, human; C482983:YWHAE protein, human; D005092:Exonucleases; D005095:Exoribonucleases; C413442:SFN protein, human; C517008:CDC25B protein, human; D020687:cdc25 Phosphatases