Bone morphogenetic protein-2 (BMP-2) and transforming growth factor-b1 (TGF-b1) alter connexin 43 phosphorylation in MC3T3-E1 Cells

Bone morphogenetic proteins (BMPs) and transforming growth factor-bs (TGF-bs) are important regulators of bone repair and regeneration. BMP-2 and TGF-b1 have been shown to inhibit gap junctional intercellular communication (GJIC) in MC3T3-E1 cells. Connexin 43 (Cx43) has been shown to mediate GJIC in osteoblasts and it is the predominant gap junctional protein expressed in these murine osteoblast-like cells. We examined the expression, phosphorylation, and subcellular localization of Cx43 after treatment with BMP-2 or TGF-b1 to investigate a possible mechanism for the inhibition of GJIC. Northern blot analysis revealed no detectable change in the expression of Cx43 mRNA. Western blot analysis demonstrated no significant change in the expression of total Cx43 protein. However, significantly higher ratios of unphosphorylated vs. phosphorylated forms of Cx43 were detected after BMP-2 or TGF-b1 treatment. Immunofluorescence and cell protein fractionation revealed no detectable change in the localization of Cx43 between the cytosol and plasma membrane. BMP-2 and TGF-b1 do not alter expression of Cx43 at the mRNA or protein level. BMP-2 and TGF-b1 may inhibit GJIC by decreasing the phosphorylated form of Cx43 in MC3T3-E1 cells.