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

• Published in: Journal of cell science Vol. 117; no. Pt 14; p. 3011
• Main Authors: Uchida, Sanae, Kuma, Akiko, Ohtsubo, Motoaki, Shimura, Mari, Hirata, Masato, Nakagama, Hitoshi, Matsunaga, Tsukasa, Ishizaka, Yukihito, Yamashita, Katsumi
• Format: Journal Article
• Language: English
• Published: England 15.06.2004
• Subjects: 14-3-3 Proteins - genetics; 14-3-3 Proteins - metabolism; Binding Sites; Biomarkers, Tumor - genetics; Biomarkers, Tumor - metabolism; cdc25 Phosphatases - metabolism; Cell Cycle Proteins - metabolism; Cell Nucleus - metabolism; Cells, Cultured; Exonucleases - genetics; Exonucleases - metabolism; Exoribonucleases; Humans; Mutagenesis, Site-Directed; Neoplasm Proteins - genetics; Neoplasm Proteins - metabolism; Protein Binding; Protein Transport
• ISSN: 0021-9533

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.