MAMMALIAN P55CDC MEDIATES ASSOCIATION OF THE SPINDLE CHECKPOINT PROTEIN MAD2 WITH THE CYCLOSOME ANAPHASE-PROMOTING COMPLEX, AND IS INVOLVEDIN REGULATING ANAPHASE ONSET AND LATE MITOTIC EVENTS/

We have investigated the function of p55CDC, a mammalian protein relat ed to Cdc20 and Hct1/Cdh1 in Saccharomyces cerevisiae, and Fizzy and F izzy-related in Drosophila. Immunofluorescence studies and expression of a p55CDC-GFP chimera demonstrate that p55CDC is concentrated at the kinetochores in M phase cells from late prophase to telophase. Some p 55CDC is also associated with the spindle microtubules and spindle pol es, and some is diffuse in the cytoplasm, At anaphase, the concentrati on of p55CDC at the kinetochores gradually diminishes, and is gone by late telophase. In extracts prepared from M phase, but not from interp hase HeLa cells, p55CDC coimmunoprecipitates with three important elem ents of the M phase checkpoint machinery: Cdc27, Cdc16, and Mad2. p55C DC is required for binding Mad2 with the Cdc27 and Cdc16. Thus, it is likely that p55CDC mediates the association of Mad2with the cyclosome/ anaphase-promoting complex. Microinjection of anti-p55CDC antibody int o mitotic mammalian cells induces arrest or de lay at metaphase, and i mpairs progression of late mitotic events. These studies suggest that mammalian p55CDC may be part of a regulatory and targeting complex for the anaphase-promoting complex.