Loss of gap junction plaques and inhibition of intercellular communication in ilimaquinone-treated BICR-M1Rk and NRK cells.
To examine the mechanism(s) and pathways of gap junction formation and removal a novel and reversible inhibitor of protein secretion, ilimaquinone (IQ), was employed. IQ has been reported to cause the vesiculation of Golgi membranes, block protein transport at the cis-Golgi and depolymerize cytoplasmic microtubules. Connexin43 (Cx43) immunolabeling and dye microinjection experiments revealed that gap junction plaques were lost and intercellular communication was inhibited following IQ treatment for 1 hr in BICR-M1Rk rat mammary tumor cells and for 2 hr in normal rat kidney (NRK) cells. Gap junction plaques and intercellular communication recovered within 2 hr when IQ was removed. IQ, however, did not affect the distribution of zonula occludens-1, a protein associated with tight junctions. Western blot analysis revealed that the IQ-induced loss of gap junction plaques was accompanied by a limited reduction in the highly phosphorylated form of Cx43, previously shown to be correlated with gap junction plaques. The presence of IQ inhibited the formation of new gap junction plaques in BICR-M1Rk cells under conditions where preexisting gap junctions were downregulated by brefeldin A treatment. Treatment of BICR-M1Rk and NRK cells with other microtubule depolymerization agents did not inhibit plaque formation or promote rapid gap junction removal. These findings suggest that IQ disrupts intercellular communication by inhibiting the events that are involved in plaque formation and/or retention at the cell surface independent of its effects on microtubules. Our results also suggest that additional factors other than phosphorylation are necessary for Cx43 assembly into gap junction plaques.
Feldman
P A
PA
Department of Anatomy and Cell Biology, McGill University, 3640 University St., Montreal, Quebec, Canada, H3A 2B2.
Kim
J
J
Laird
D W
DW
D000818:Animals; D015153:Blotting, Western; D002450:Cell Communication / Q000187:drug effects*; D002478:Cells, Cultured; D018031:Connexin 43 / Q000096:biosynthesis / Q000187:drug effects; D017629:Gap Junctions / Q000187:drug effects*; D007668:Kidney / Q000166:cytology; D015674:Mammary Neoplasms, Animal; D018613:Microscopy, Confocal; D011809:Quinones / Q000494:pharmacology*; D051381:Rats
D018031:Connexin 43; D011809:Quinones; C067346:illimaquinone