Connexin37 protects against atherosclerosis by regulating monocyte adhesion.

A genetic polymorphism in the human gene encoding connexin37 (CX37, encoded by GJA4, also known as CX37) has been reported as a potential prognostic marker for atherosclerosis. The expression of this gap-junction protein is altered in mouse and human atherosclerotic lesions: it disappears from the endothelium of advanced plaques but is detected in macrophages recruited to the lesions. The role of CX37 in atherogenesis, however, remains unknown. Here we have investigated the effect of deleting the mouse connexin37 (Cx37) gene (Gja4, also known as Cx37) on atherosclerosis in apolipoprotein E-deficient (Apoe(-/-)) mice, an animal model of this disease. We find that Gja4(-/-)Apoe(-/-) mice develop more aortic lesions than Gja4(+/+)Apoe(-/-) mice that express Cx37. Using in vivo adoptive transfer, we show that monocyte and macrophage recruitment is enhanced by eliminating expression of Cx37 in these leukocytes but not by eliminating its expression in the endothelium. We further show that Cx37 hemichannel activity in primary monocytes, macrophages and a macrophage cell line (H36.12j) inhibits leukocyte adhesion. This antiadhesive effect is mediated by release of ATP into the extracellular space. Thus, Cx37 hemichannels may control initiation of the development of atherosclerotic plaques by regulating monocyte adhesion. H36.12j macrophages expressing either of the two CX37 proteins encoded by a polymorphism in the human GJA4 gene show differential ATP-dependent adhesion. These results provide a potential mechanism by which a polymorphism in CX37 protects against atherosclerosis.

Cooperation between snail and LEF-1 transcription factors is essential for TGF-beta1-induced epithelial-mesenchymal transition.

Transforming growth factor beta 1 (TGF-beta1) has been shown to induce epithelial-mesenchymal transition (EMT) during various stages of embryogenesis and progressive disease. This alteration in cellular morphology is typically characterized by changes in cell polarity and loss of adhesion proteins such as E-cadherin. Here we demonstrate that EMT is associated with loss of claudin-1, claudin-2, occludin, and E-cadherin expression within 72 h of exposure to TGF-beta1 in MDCKII cells. It has been suggested that this expression loss occurs through TGF-beta1 in a Smad-independent mechanism, involving MEK and PI3K pathways, which have previously been shown to induce expression of the Snail (SNAI-1) gene. Here we show that these pathways are responsible for loss of tight junctions and a partial loss of E-cadherin. However, our results also demonstrate that a complete loss of E-cadherin and transformation to the mesenchymal phenotype are dependent on Smad signaling, which subsequently stimulates formation of beta-catenin/LEF-1 complexes that induce EMT.

Paracellular ion channel at the tight junction.

The tight junction of epithelial cells excludes macromolecules but allows permeation of ions. However, it is not clear whether this ion-conducting property is mediated by aqueous pores or by ion channels. To investigate the permeability properties of the tight junction, we have developed paracellular ion flux assays for four major extracellular ions, Na(+), Cl(-), Ca(2+), and Mg(2+). We found that the tight junction shares biophysical properties with conventional ion channels, including size and charge selectivity, dependency of permeability on ion concentration, competition between permeant molecules, anomalous mole-fraction effects, and sensitivity to pH. Our results support the hypothesis that discrete ion channels are present at the tight junction. Unlike conventional ion channels, which mediate ion transport across lipid bilayers, the tight junction channels must orient parallel to the plane of the plasma membranes to support paracellular ion movements. This new class of paracellular-tight junction channels (PTJC) facilitates the transport of ions between separate extracellular compartments.

Xwnt-2 (Xwnt-2b) is maternally expressed in Xenopus oocytes and embryos.

Xwnt-2 (formerly Xwnt-2b) is a member of the Xwnt-8 class of axis-inducing Wnts. Its zygotic expression is at the prosencephalic-mesencephalic border of the early tadpole brain and above the heart primordium [Mech. Dev. 63 (1997) 199]. Here, we report that Xwnt-2 has an earlier, maternal pattern of expression. It is detected in the oocyte, egg and the developing embryo. Studies of the spatial localization of maternal Xwnt-2 show transcripts in both vegetal and animal blastomeres with enrichment in the animal hemisphere. The identification of maternal Xwnt-2 raises questions about possible roles of dorsalizing Xwnts in axial patterning of the Xenopus embryo.

Nonreceptor tyrosine kinase c-Yes interacts with occludin during tight junction formation in canine kidney epithelial cells.

Occludin is an integral membrane protein that is tyrosine phosphorylated when localized at tight junctions. When Ca(2+) was depleted from the culture medium, occludin tyrosine phosphorylation was diminished from Madin-Darby canine kidney epithelial cells in 2 min. This dephosphorylation was correlated with a significant reduction in transepithelial electrical resistance (TER), indicating a global loss of the tight junction barrier function. Reconstitution of Ca(2+) resulted in a robust tyrosine rephosphorylation of occludin that was temporally associated with an increase in TER. Moreover, we demonstrate in this study that occludin was colocalized with the nonreceptor tyrosine kinase c-Yes at cell junction areas and formed an immunoprecipitable complex with c-Yes in vivo. This complex dissociated when the cells were incubated in medium without Ca(2+) or treated with a c-Yes inhibitor, CGP77675. In the presence of CGP77675 after Ca(2+) repletion, occludin tyrosine phosphorylation was completely abolished and both tight junction formation and the increase of the TER were inhibited. Our study thus provides strong evidence that occludin tyrosine phosphorylation is tightly linked to tight junction formation in epithelial cells, and that the nonreceptor tyrosine kinase c-Yes is involved in the regulation of this process.