Molecular Anatomy of Functional Differentination Affected by Combination of Gap Junctions, Connexins

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Cx45-deficient ES cells were induced to differentiate into contracting cardiac myocytes in vitro. Their contractions were uncoordinated due to defective intercellular communication. In vivo, we created mice lacking Cx45 in all the somatic cells (Cx45-KO), in cardiac myocytes (Cx45-CA), and in vascular endothelial cells (Cx45-Tie2). We found intriguing differences of phenotypes in each mutant. The Cx45-KO mice died at the E10 stage, showing atrioventricular conduction block and an endocardial cushion defect in the early heart. The Ca2+-dependent transcription factor Nfatcl was in inactive, cytoplasmic form in the Cx45-KO endocardial endothelium. The Cx45-CA mice died at the E10 stage, showing atrioventricular conduction block, but without the endocardial cushion defect. Nfatcl was in active nuclear form in the Cx45-CA endocardial endothelium. The lethal stage shows that the conduction block is the primary cause of death in the Cx45-KO mice. The Cx45-Tie2 mice, in contrast, did not show … More any abnormality. Moreover, we created mice lacking another cardiac connexin Cx43, in the absence of Cx45. They were identical to the Cx45-KO mice, indicating that Cx45 constitutes the most significant gap junction protein in the early heart.Next, we created cardiac troponin T (cTnT)-deficient mice. Because it is an essential component of the contracting apparatus, the cTnT-deficient mice did not show any contractions. Their lethal stage was the same as the above Cx45-mutants. They showed the endocardial cushion defect due to retarded cardiac development. Nfatcl localization, however, indicated active nuclear form.The endocardial cushion defect was only apparent when Cx45 was absent in both of the myocardial and the endocardial layer. Our model, in which Cx45 regulates the epithelial-mesenchymal transformation in the endocardial endothelium, needs further revise by the future study, though the defect is not caused by growth retardation. Collaborations with the overseas coworkers are in progress, revealing the tissue specific functions of Cx45 in neuronal/glial/neural crest cells. Roles of Cx45 in these cell types will be evident in the near future. Less