distinct pools of cdc25c are phosphorylated on specific tp sites and differentially localized in human mitotic cells不同的池cdc25c磷酸化在特定tp网站和不同局部在人类有丝分裂细胞.pdfVIP

Distinct Pools of cdc25C Are Phosphorylated on Specific TP Sites and Differentially Localized in Human Mitotic Cells Celine Franckhauser, Daria Mamaeva, Lisa Heron-Milhavet, Anne Fernandez, Ned J. C. Lamb* Cell Biology Unit, Institute de Genetique Humain, CNRS-UPR1142, Montpellier, France Abstract Background: The dual specificity phosphatase cdc25C was the first human cdc25 family member found to be essential in the activation of cdk1/cyclin B1 that takes place at the entry into mitosis. Human cdc25C is phosphorylated on Proline- dependent SP and TP sites when it becomes active at mitosis and the prevalent model is that this phosphorylation/ activation of cdc25C would be part of an amplification loop with cdk1/cyclin B1. Methodology/Principal Findings: Using highly specific antibodies directed against cdc25C phospho-epitopes, pT67 and pT130, we show here that these two phospho-forms of cdc25C represent distinct pools with differential localization during human mitosis. Phosphorylation on T67 occurs from prophase and the cdc25C-pT67 phospho-isoform closely localizes with condensed chromosomes throughout mitosis. The phospho-T130 form of cdc25C arises in late G2 and associates predominantly with centrosomes from prophase to anaphase B where it colocalizes with Plk1. As shown by immunoprecipitation of each isoform, these two phospho-forms are not simultaneously phosphorylated on the other mitotic TP sites or associated with one another. Phospho-T67 cdc25C co-precipitates with MPM2-reactive proteins while pT130-cdc25C is associated with Plk1. Interaction and colocalization of phosphoT130-cdc25C with Plk1 demonstrate in living cells, that the sequence around pT130 acts as a true Polo Box Domain (PBD) binding site as previously identified f