ACTIVE MAP KINASE IN MITOSIS - LOCALIZATION AT KINETOCHORES AND ASSOCIATION WITH THE MOTOR PROTEIN CENP-E

To investigate possible involvement of the mitogen-activated protein ( MAP) kinases ERK1 and ERK2 (extracellular signal-regulated kinases) in somatic cell mitosis, we have used indirect immunofluorescence with a highly specific phospho-MAP kinase antibody and found that a portion of the active MAP kinase is localized at kinetochores, asters, and the midbody during mitosis. Although the aster labeling was constant from the time of nuclear envelope breakdown, the kinetochore labeling firs t appeared at early prometaphase, started to fade during chromosome co ngression, and then disappeared at midanaphase, At telophase, active M AP kinase localized at the midbody, Based on colocalization and the pr esence of a MAP kinase consensus phosphorylation site, we identified t he kinetochore motor protein CENP-E as a candidate mitotic substrate f or MAP kinase, CENP-E was phosphorylated in vitro by MAP kinase on sit es that are known to regulate its interactions with microtubules and w as found to associate in vivo preferentially with the active MAP kinas e during mitosis. Therefore, the presence of active MAP kinase at spec ific mitotic structures and its interaction with CENP-E suggest that M AP kinase could play a role in mitosis at least in part by altering th e ability of CENP-E to mediate interactions between chromosomes and mi crotubules.