Functional roles of the gap junction protein, Connexin43.4, during vertebrate development.

Abstract

During early embryonic development, compartments of cells must send and receive signals in order to coordinate activities such as proliferation, migration and differentiation. Cells regulate these activities inpart through cell-to-cell communication by sharing small signaling molecules and ions via Connexin (Cx)-based gap junction (GJ) channels that connect the cytoplasms of adjacent cells. Mouse knockout models of Cx proteins have demonstrated the necessity for GJ communication during development; cx45 knockouts are embryonic lethal and cx43 null embryos exhibit defects in heart development. However, the mechanisms by which GJ communication regulate early embryonic development, particularly in terms of axis specification and patterning, remain largely unexamined. Here, the role of zebrafish Cx43.4 (the ortholog of mammalian Cx45) in patterning the asymmetric left-right (L-R) axis of zebrafish was examined. L-R patterning is directed by ciliary beating that generates a leftward fluid flow in the mammalian node or in Kupffer's vesicle (KV), the related structure in zebrafish. Leftward nodal flow is required for normal asymmetric organ placement and function. Cx43.4 expression was detected in KV and morpholino (MO) knockdown of Cx43.4 resulted in randomized organ distribution and reversed asymmetric gene expression. The major finding of these knockdown experiments is that Cx43.4 is required for the morphogenesis of KV. Additionally, Cx43.4 hemichannels, rather than GJ communication, are sufficient to rescue the defects in asymmetric patterning. Finally, the function of Cx43.4 hemichannels, together with purinergic receptors, is required for the morphogenesis of the KV epithelium. These novel findings have important implications for both the communication and L-R development fields, as well as suggesting new roles for Cx proteins in the early development of a variety of epithelial tissues.