The T-box gene eomesodermin (eomes) has been implicated in mesoderm specification and patterning in both zebrafish and frog. Here, we describe an additional function for eomes in the control of morphogenesis. Epiboly, the spreading and thinning of an epithelial cell sheet, is a central component of gastrulation in many species; however, despite its importance, little is known about its molecular control. Here, we show that repression of eomes function in the zebrafish embryo dramatically inhibits epiboly movements. We also show that eomes regulates the expression of a zygotic homeobox transcription factor mtx2. Gene knockdown of mtx2 using antisense morpholino oligonucleotides, likewise, leads to an inhibition of epiboly; moreover, we show that knockdown of mtx2 function in the extraembryonic yolk syncytial layer only is sufficient to cause epiboly defects. Thus, we have identified two components in a molecular pathway controlling epiboly and show that interactions between deep layer cells of the embryo proper and extraembryonic tissues contribute in a coordinated manner to different aspects of epiboly movements. Developmental Dynamics 233:105-114, 2005.

Rhodopsin (rh1) is the visual pigment expressed in rod photoreceptors of vertebrates that is responsible for initiating the critical first step of dim-light vision. Rhodopsin is usually a single copy gene; however, we previously discovered a novel rhodopsin-like gene expressed in the zebrafish retina, rh1-2, which we identified as a functional photosensitive pigment that binds 11-cis retinal and activates in response to light. Here, we localized expression of rh1-2 in the zebrafish retina to a subset of peripheral photoreceptor cells, which indicates a partially overlapping expression pattern with rh1. We also expressed, purified and characterized Rh1-2, including investigation of the stability of the biologically active intermediate. Using fluorescence spectroscopy, we found the half-life of the rate of retinal release of Rh1-2 following photoactivation to be more similar to that of the visual pigment rhodopsin than to the non-visual pigment exo-rhodopsin (exorh), which releases retinal around 5 times faster. Phylogenetic and molecular evolutionary analyses show that rh1-2 has ancient origins within teleost fishes, is under similar selective pressure to rh1, and likely experienced a burst of positive selection following its duplication and divergence from rh1. These findings indicate that rh1-2 is another functional visual rhodopsin gene, which contradicts the prevailing notion that visual rhodopsin is primarily found as a single copy gene within ray-finned fishes. The reasons for retention of this duplicate gene, as well as possible functional consequences for the visual system, are discussed.

The organizer is essential for dorsal-ventral (DV) patterning in vertebrates. Goosecoid (Gsc), a transcriptional repressor found in the organizer, elicits partial secondary axes when expressed ventrally in Xenopus, similar to an organizer transplant. Although gsc is expressed in all vertebrate organizers examined, knockout studies in mouse suggested that it is not required for DV patterning. Moreover, experiments in Xenopus and zebrafish suggest a role in head formation, although a function in axial mesoderm formation is less clear. To clarify the role of Gsc in vertebrate development, we used gain-and loss-of-function approaches in zebrafish. Ventral injection of low doses of gsc produced incomplete secondary axes, which we propose results from short-range repression of BMP signaling. Higher gsc doses resulted in complete secondary axes and long-range signaling, correlating with repression of BMP and Wnt signals. In striking contrast to Xenopus, the BMP inhibitor Chordin (Chd) is not required for Gsc function. Gsc produced complete secondary axes in chd null mutant embryos and gsc-morpholino knockdown in chd mutants enhanced the mutant phenotype, suggesting that Gsc has Chd-independent functions in DV patterning. Even more striking was that Gsc elicited complete secondary axes in the absence of three secreted BMP antagonists, Chd, Follistatin-like 1b and Noggin 1, suggesting that Gsc functions in parallel with secreted BMP inhibitors. Our findings suggest that Gsc has dose dependent effects on axis induction and provide new insights into molecularly distinct short-and long-range signaling activities of the organizer.