[Show abstract][Hide abstract] ABSTRACT: The transcription factor Pax6 is required for eye morphogenesis in humans, mice and insects, and can induce ectopic eye formation in vertebrate and invertebrate organisms. Although the role of Pax6 has intensively been studied, only a limited number of genes have been identified that depend on Pax6 activity for their expression in the mammalian visual system. Using a large-scale in situ hybridization screen approach, we have identified a novel gene expressed in the mouse optic vesicle. This gene, Necab, encodes a putative cytoplasmic Ca(2+)-binding protein and coincides with Pax6 expression pattern in the neural ectoderm of the optic vesicle and in the forebrain pretectum. Remarkably, Necab expression is absent in both structures in Pax6 mutant embryos. By contrast, the optic vesicle-expressed homeobox genes Rx, Six3, Otx2 and Lhx2 do not exhibit an altered expression pattern. Using gain-of-function experiments, we show that Pax6 can induce ectopic expression of Necab, suggesting that Necab is a direct or indirect transcriptional target of Pax6. In addition, we have found that Necab misexpression can induce ectopic expression of the homeobox gene Chx10, a transcription factor implicated in retina development. Taken together, our results provide evidence that Necab is genetically downstream of Pax6 and that it is a part of a signal transduction pathway in retina development.