CDC25B phosphorylation by p38 and MK-2.

CDC25B is one of the three human phosphatases that are involved in the control of the activation of cyclin-dependent kinases. CDC25B participates in regulating entry into mitosis and appears to play a key role in the checkpoint response to DNA injury. CDC25B has been reported to be regulated by a number of kinases and controversial evidence suggests that it is phosphorylated by p38SAPK and/or MAPKAP kinase-2. In this report, we clarify this issue using an approach combining mass spectrometry and the use of specific antibodies against phosphorylated CDC25B residues. We report that MAPKAP kinase-2 phosphorylates CDC25B on multiple sites including S169, S323, S353 and S375, while p38SAPK phosphorylates CDC25B on S249. We show that the S323-phosphorylated form of CDC25B is detected at the centrosome during a normal cell cycle. Since most of these sites are also phosphorylated by several other kinases, our observations highlight the difficulty in characterizing and understanding in vivo phosphorylation patterns.