Mutations at phosphorylation sites of Xenopus microtubule-associated protein 4 affect its microtubule-binding ability and chromosome movement during mitosis

Microtubule-associated proteins (MAPs) bind to and stabilize microtubules ( MTs) both in vitro and in vivo and are thought to regulate MT dynamics duri ng the cell cycle. It is known that p220, a major MAP of Xenopus, is phosph orylated by p34(cdc2) kinase as well as MAP kinase in mitotic cells, and th at the phosphorylated p220 loses its MT-binding and -stabilizing abilities in vitro. We cloned a full-length cDNA encoding p220, which identified p220 as a Xenopus homologue of MAP4 (XMAP4). To examine the physiological relev ance of XMAP4 phosphorylation in vivo, Xenopus A6 cells were transfected wi th cDNAs encoding wild-type or various XMAP4 mutants fused with a green flu orescent protein. Mutations of serine and threonine residues at p34cdc2 kin ase-specific phosphorylation sites to alanine interfered with mitosis-assoc iated reduction in MT affinity of XMAP4, and their overexpression affected chromosome movement during anaphase A. These findings indicated that phosph orylation of XMAP4 (probably by p34(cdc2) kinase) is responsible for the de crease in its MT-binding and -stabilizing abilities during mitosis, which a re important for chromosome movement during anaphase A.