Quiescent mammary epithelial cells have reduced connexin43 but maintain a high level of gap junction intercellular communication

• Published in: Developmental genetics 1999, Vol.24 (1/2), p.111-122
• Main Authors: Sia, M.A, Woodward, T.L, Turner, J.D, Laird, D.W
• Format: Article
• Language: English
• Subjects: animal proteins; Animals; Blotting, Western; Brefeldin A - pharmacology; Cattle; Cell Communication; Cell Division; cell lines; Cell Membrane - metabolism; Connexin 43 - analysis; Connexin 43 - metabolism; connexin43; Contact Inhibition; Cytoplasm - chemistry; Epithelial Cells - cytology; Epithelial Cells - metabolism; epithelium; Female; Fluorescent Antibody Technique; gap junctions; Gap Junctions - chemistry; Gap Junctions - physiology; immunocytochemistry; intercellular communication; mammary glands; Mammary Glands, Animal - cytology; Mammary Glands, Animal - metabolism; Phosphorylation; protein content; quiescence; α1 connexin
• ISSN: 0192-253X; 1520-6408
• DOI: 10.1002/(SICI)1520-6408(1999)24:1/2<111::AID-DVG11>3.0.CO;2-P

Gap junctions have been implicated in growth control, but it remains unclear whether cells that enter a quiescent state continue to express connexins and maintain a high level of gap junction intercellular communication (GJIC). To this end, MAC-T cells, a bovine mammary epithelial cell line, were serum starved for 48 h to induce a quiescent (G0) state. In quiescent cells, [3H]thymidine incorporation was reduced by 97.3% from serum-fed controls. Western blotting in conjunction with Phosphorlmager analysis revealed up to a 20-fold decrease in the expression of the gap junction protein connexin43 (Cx43 or alpha 1) and a shift toward the unphosphorylated form in quiescent cells. However, cell-to-cell transfer of the gap junction-permeable fluorescent tracer, Lucifer yellow, was only moderately reduced in quiescent cells. In control cells, Cx43 was predominantly perinucleor, although it was also present at sites of cell-cell apposition. In quiescent cells, intracellular labeling for Cx43 decreased without a corresponding reduction at areas of cell-cell contact. Recovery from serum deprivation resulted in increased thymidine incorporation that corresponded with an elevation in Cx43 protein expression and phosphorylation. In parallel studies, MAC-T cells were also induced to enter a quiescent state through contact inhibition. Despite a 20-fold reduction in 5-bromo-2'-deoxyuridine and a substantial reduction in intracellular Cx43, contact inhibited MAC-T cells also maintained gap junctions and GJIC. These experiments demonstrate that the maintenance of dye coupling in quiescent mammary cells is correlated with a redistribution of intracellular stores of Cx43.