PHOSPHORYLATION BY P34(CDC2) PROTEIN-KINASE REGULATES BINDING OF THE KINESIN-RELATED MOTOR HSEG5 TO THE DYNACTIN SUBUNIT P150(GLUED)

The kinesin-related motor HsEg5 is essential for centrosome separation , and its association with centrosomes appears to be regulated by phos phorylation of tail residue threonine 927 by the p34(cdc2) protein kin ase. To identify proteins able to interact with the tail of HsEg5, we performed a yeast two-hybrid screen with a HsEg5 stalk-tail construct as bait. We isolated a cDNA coding for the central, alpha-helical regi on of human p150(Glued), a prominent component of the dynactin complex . The interaction between HsEg5 and p150(Glued) was enhanced upon acti vation of p34(CDC28), the budding yeast homolog of p34(cdc2), provided that HsEg5 had a phosphorylatable residue at position 927. Phosphoryl ation also enhanced the specific binding of p150(Glued) to the tail do main of HsEg5 in vitro, indicating that the two proteins are able to i nteract directly. Immunofluorescence microscopy revealed co-localizati on of HsEg5 and p150(Glued) during mitosis but not during interphase, consistent with a cell cycle-dependent association between the two pro teins. Taken together, these results suggest that HsEg5 and p150(Glued ) may interact in mammalian cells in vivo and that p34(cdc2) may regul ate this interaction. Furthermore, they imply that the dynactin comple x may functionally interact not only with dynein but also with kinesin -related motors.