Functional interactions of genes mediating convergent extension, knypek and trilobite, during the partitioning of the eye primordium in zebrafish

Vertebrate eye development in the anterior region of the neural plate involves a series of inductive interactions dependent on the underlying prechordal plate and signals from the midline of the neural plate, including Hedgehog. The mechanisms controlling the spatiotemporal expression pattern of hedgehog genes are currently not understood. Cyclopia is observed in trilobite (tri) and knypek (kny) mutants with affected convergent extension of the embryonic axis during gastrulation. Here, we demonstrate that tri mutants show a high frequency of partial or complete cyclopia, kny mutants exhibit cyclopia infrequently, while kny(m119) tri(m209) double-mutant embryos have dramatically reduced convergent extension and are completely cyclopic. We analyzed the relationships between the convergent extension defect, the expression of hedgehog and prechordal plate genes, and the formation of cyclopia in kny(m119) and tri(m209) mutants. Our results correlate the cyclopia phenotype with the abnormal location of hh-expressing cells with respect to the optic primordium. We show that cyclopia in these mutants is not due to an incompetence of tri and kny cells to respond to Hedgehog signaling. Rather, it is a consequence of exceeding a critical distance (>40-50 μm) between hedgehog-expressing cells and the prospective eye field. We hypothesize that at this distance, midline cells are not in an appropriate position to physically separate the eye field and that HH and other signals do not reach the appropriate target cells. Furthermore, tri and kny have overlapping functions in establishing proper alignment of the anterior neural plate and midline cells expressing shh and twhh genes when the partitioning of the eye primordium takes place.