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During embryogenesis, the germ layers, including the endoderm, undergo convergence and extension movements to narrow and elongate the body plan. In zebrafish, the dorsal migration of endodermal cells during gastrulation is controlled by chemokine signaling, but little is known about how they migrate during segmentation. Here, we show that glypican 4 (Gpc4), a member of the heparin sulfate proteoglycan family, is required for efficient migration of anterior endodermal cells during early segmentation, regulating Rac activation to maintain polarized actin-rich lamellipodia. An endoderm transplantation assay showed that Gpc4 regulates endoderm migration in a non-cell-autonomous fashion. Further analyses revealed that the impaired endoderm migration in gpc4 mutants results from increases in the expression and assembly of fibronectin and laminin, major components of the extracellular matrix (ECM). Notably, we found that matrix metalloproteinase 14 (Mmp14a/b) is required for the control of ECM expression during endoderm migration, with Gpc4 acting through Mmp14a/b to limit ECM expression. Our results suggest that Gpc4 is crucial for generating the environment required for efficient migration of endodermal cells, uncovering a novel function of Gpc4 during development.

Here, we describe an optogenetic gene expression system optimized for use in zebrafish. This system overcomes the limitations of current inducible expression systems by enabling robust spatial and temporal regulation of gene expression in living organisms. Because existing optogenetic systems show toxicity in zebrafish, we re-engineered the blue-light-activated EL222 system for minimal toxicity while exhibiting a large range of induction, fine spatial precision and rapid kinetics. We validate several strategies to spatially restrict illumination and thus gene induction with our new TAEL (TA4-EL222) system. As a functional example, we show that TAEL is able to induce ectopic endodermal cells in the presumptive ectoderm via targeted sox32 induction. We also demonstrate that TAEL can be used to resolve multiple roles of Nodal signaling at different stages of embryonic development. Finally, we show how inducible gene editing can be achieved by combining the TAEL and CRISPR/Cas9 systems. This toolkit should be a broadly useful resource for the fish community.