Molecular cloning, expression analysis, and functional characterization ofconnexin44.1: A zebrafish lens gap junction protein

The connexin family of genes codes for proteins that oligomerize into a con nexon of six subunits to form one half of the gap junction channel. Gap jun ctions are plasma membrane structures that mediate intercellular communicat ion by joining the cytoplasm of two cells, allowing the passage of small mo lecules and metabolites, and contributing significantly to the maintenance of tissue homeostasis. The signaling mediated by these junctions appears to be necessary for the correct timing of key developmental events. This comm unication is especially important in the avascular lens where the intercell ular passage of metabolites, second messengers, and ions is necessary to ma intain the correct ionic balance in the lens fibre cells, and prevent catar act formation. To characterize the role that the connexin genes play in dev elopment; a novel connexin was cloned from zebrafish. A genomic clone was i solated that contained a 1,173 base open reading frame. The nucleotide sequ ence in this open reading frame shows extensive sequence similarity to mous e connexin50 (Cx50), chicken Cx45.6, sheep Cx49, and human Cx50, The protei n encoded by this open reading frame contains 391 amino acids, with a predi cted molecular weight of 44.1 kDa and a typical connexin transmembrane topo logy. By using the LN54 radiation hybrid panel, the Cx44.1 gene was mapped to linkage group 1, Whole-mount in situ hybridization and Northern blot ana lyses were performed on zebrafish embryos at various developmental stages t o characterize the developmental expression of the Cx44.1 message. The ocul ar lens was the only tissue in which Cx44.1 transcripts were detected. The transcripts were first detected in the lens around 24 hr post fertilization and remained detectable until 120 hr post fertilization. Electrophysiologi cal analysis of Cx44.1 channels revealed gating properties that were virtua lly identical to the mouse and chicken orthologues of Cx44.1. (C) 2001 Wile y-Liss, Inc.