A MITOSIS-SPECIFIC PHOSPHORYLATION OF THE GAP JUNCTION PROTEIN CONNEXIN43 IN HUMAN VASCULAR CELLS - BIOCHEMICAL-CHARACTERIZATION AND LOCALIZATION

Western blotting studies revealed that connexin43 (Cx43), one of the m ajor gap junction proteins in human vascular endothelial cells, is pos ttranslationally modified during mitosis. This mitosis-specific modifi cation results in a Cx43 species that migrates as a single protein ban d and was designated Cx43(m). Cx43(m) was shown to be the result of ad ditional Ser/Thr phosphorylation as indicated by: (a) the increased ge l mobility induced by both alkaline phosphatase and the Seri Thr-speci fic protein phosphatase-2A (PP2A) and (b) the removal of virtually all P-32(i) from CX43(m) by PP2A. Immunofluorescent confocal microscopy o f mitotic cells revealed that Cx43 is intracellularly located, while i n nonmitotic cells Cx43 is located at regions of cell-cell contact, Dy e coupling studies revealed that mitotic endothelial cells were uncoup led from each other and from nonmitotic cells. After cytokinesis, sist er cells resumed cell coupling independent of de novo protein synthesi s. The mitosis-specific phosphorylation of Cx43 correlates with the tr ansient loss of gap junction intercellular communication and redistrib ution of Cx43, suggesting that a protein kinase that regulates gap jun ctions is active in M-phase.