Examination of Gbx2 function in zebrafish development [abstract]

Abstract

Gbx2 is a member of the Gbx class of homeobox genes which encode DNA-binding transcription factors. The amino acid sequence of Gbx2 is highly conserved across multiple species (e.g. mice, zebrafish, chicken, and frogs) with 100% sequence identity between the Gbx2 homeodomains of the species examined. The function of Gbx2 in mice and zebrafish has been studied with loss-of-function and hypomorphic (reduced expression) models in both species. The results of these studies have demonstrated a requirement for Gbx2 in normal development of the mid/hindbrain organizer (isthmus) and anterior hindbrain. The hindbrain controls many basic life functions such as breathing and heartbeat. In early vertebrate development, the hindbrain is organized into eight distinct segments called rhombomeres. Rhombomeres give rise to hindbrain regions such as the cerebellum, pons, and medulla oblongata. In situ hybridization studies in Gbx2 null and hypomorphic mice have shown that cranial nerve V, which is derived from rhombomeres 2 and 3, fails to develop normally without wild-type levels of Gbx2. Mice lacking wild-type levels of Gbx2 die immediately after birth. To examine if Gbx2 has a similar impact on cranial nerve V development in zebrafish, we injected a morpholino specific for zebrafish Gbx2 into zebrafish embryos to silence the Gbx2 gene early in development. Our findings have shown that zebrafish subjected to the morpholino have a similar phenotype in the hindbrain as Gbx2 hypomorphic mice. Our present research will attempt to rescue the normal hindbrain phenotype in zebrafish embryos by simultaneously injecting the Gbx2 morpholino with synthesized zebrafish Gbx2 mRNA. We will also attempt to rescue the normal phenotype with synthesized mouse Gbx1 mRNA and mouse Gbx2 mRNA.