A Yap-dependent mechanoregulatory program sustains cell migration for embryo axis assembly.

The assembly of the embryo's primary axis is a fundamental landmark for the establishment of the vertebrate body plan. Although the morphogenetic movements directing cell convergence towards the midline have been described extensively, little is known on how gastrulating cells interpret mechanical cues. Yap proteins are well-known transcriptional mechanotransducers, yet their role in gastrulation remains elusive. Here we show that the double knockout of yap and its paralog yap1b in medaka results in an axis assembly failure, due to reduced displacement and migratory persistence in mutant cells. Accordingly, we identified genes involved in cytoskeletal organization and cell-ECM adhesion as potentially direct Yap targets. Dynamic analysis of live sensors and downstream targets reveal that Yap is acting in migratory cells, promoting cortical actin and focal adhesions recruitment. Our results indicate that Yap coordinates a mechanoregulatory program to sustain intracellular tension and maintain the directed cell migration for embryo axis development.

Mutation of vsx genes in zebrafish highlights the robustness of the retinal specification network.

Genetic studies in human and mice have established a dual role for Vsx genes in retina development: an early function in progenitors' specification, and a later requirement for bipolar-cells fate determination. Despite their conserved expression patterns, it is currently unclear to which extent Vsx functions are also conserved across vertebrates, as mutant models are available only in mammals. To gain insight into vsx function in teleosts, we have generated vsx1 and vsx2 CRISPR/Cas9 double knockouts (vsxKO) in zebrafish. Our electrophysiological and histological analyses indicate severe visual impairment and bipolar cells depletion in vsxKO larvae, with retinal precursors being rerouted toward photoreceptor or Müller glia fates. Surprisingly, neural retina is properly specified and maintained in mutant embryos, which do not display microphthalmia. We show that although important cis-regulatory remodelling occurs in vsxKO retinas during early specification, this has little impact at a transcriptomic level. Our observations point to genetic redundancy as an important mechanism sustaining the integrity of the retinal specification network, and to Vsx genes regulatory weight varying substantially among vertebrate species.

Analysis of gene network bifurcation during optic cup morphogenesis in zebrafish.

Sight depends on the tight cooperation between photoreceptors and pigmented cells, which derive from common progenitors through the bifurcation of a single gene regulatory network into the neural retina (NR) and retinal-pigmented epithelium (RPE) programs. Although genetic studies have identified upstream nodes controlling these networks, their regulatory logic remains poorly investigated. Here, we characterize transcriptome dynamics and chromatin accessibility in segregating NR/RPE populations in zebrafish. We analyze cis-regulatory modules and enriched transcription factor motives to show extensive network redundancy and context-dependent activity. We identify downstream targets, highlighting an early recruitment of desmosomal genes in the flattening RPE and revealing Tead factors as upstream regulators. We investigate the RPE specification network dynamics to uncover an unexpected sequence of transcription factors recruitment, which is conserved in humans. This systematic interrogation of the NR/RPE bifurcation should improve both genetic counseling for eye disorders and hiPSCs-to-RPE differentiation protocols for cell-replacement therapies in degenerative diseases.

Trap-TRAP, a Versatile Tool for Tissue-Specific Translatomics in Zebrafish.

Developmental and physiological processes depend on the transcriptional and translational activity of heterogeneous cell populations. A main challenge in gene expression studies is dealing with this intrinsic complexity while keeping sequencing efficiency. Translating ribosome affinity purification (TRAP) methods have allowed cell-specific recovery of polyribosome-associated RNAs by genetic tagging of ribosomes in selected cell populations. Here we combined the TRAP approach with adapted enhancer trap methods (trap-TRAP) to systematically generate zebrafish transgenic lines suitable for tissue-specific translatome interrogation. Through the random integration of a GFP-tagged version of the large subunit ribosomal protein L10a (EGFP-Rpl10a), we have generated stable lines driving expression in a variety of tissues, including the retina, skeletal muscle, lateral line primordia, rhombomeres, or jaws. To increase the range of applications, a UAS:TRAP transgenic line compatible with available Gal4 lines was also generated and tested. The resulting collection of lines and applications constitutes a resource for the zebrafish community in developmental genetics, organ physiology and disease modelling.