Summary Transdifferentiation of the retina into pigmented cells in ocular retardation mice defines a new function of the homeodomain gene Chx 10

The vertebrate retina and retinal pigmented epithelium (RPE) derive from the optic vesicle, an evagination of the developing forebrain. As the optic vesicle grows and encounters signals from the prospective lens ectoderm, the domains that form the neural retina and RPE become established (reviewed by Chow and Lang, 2001). The distal optic vesicle thickens and becomes the neural retina, while the immediate proximal optic vesicle gives rise to the RPE. The interface between these two domains forms the peripheral retina, a region that gives rise to several pigmented and nonpigmented structures, including the iris and ciliary body. Several transcription factors, many of them homeodomain-containing factors, have been shown to play important roles in the specification and development of the optic vesicle (Chow and Lang, 2001). Mutations in these genes in mice or humans frequently lead to anophthalmia or microphthalmia (Graw, 2003). Other transcription factors contribute to the definition of optic vesicle compartments. Two notable transcription factors, Chx10 and Mitf, are expressed in the developing optic vesicle in patterns that are either temporally or spatially mutually exclusive (Bora et al., 1998; Liu et al., 1994; Nguyen and Arnheiter, 2000). Mutations of both Chx10 and Mitf are responsible for classically defined mouse ocular phenotypes affecting development of the retina or RPE, respectively. Chx10, a paired-like homeodomain transcription factor, is expressed in the presumptive neural retina, probably in response to inductive signals from the prospective lens ectoderm (Liu et al., 1994; Nguyen and Arnheiter, 2000). In this respect, Chx10 is the earliest characterized specific marker of retinal progenitor cells. Fibroblast growth factor (Fgf) signals from the surface ectoderm may play a role in patterning the domains of the optic vesicle and may be required for Chx10 expression in the presumptive neural retina (Nguyen and Arnheiter, 2000). In mouse, as well as the chick, Chx10 is xpressed in what appears to be nearly all retinal progenitor cells, but is absent from all of the postmitotic cell types except for bipolar interneurons (Belecky-Adams et al., 1997; Burmeister et al., 1996; Chen and Cepko, 2000) and a subset of Müller glia (Rowan and Cepko, 2004). Mutations in Chx10 cause the ocular retardation phenotype in mice, including the orJ mutant, which has a spontaneous mutation that leads to a premature stop codon and failure to produce Chx10 protein The homeodomain transcription factor Chx10 is one of the earliest markers of the developing retina. It is required for retinal progenitor cell proliferation as well as formation of bipolar cells, a type of retinal interneuron. orJ (ocular retardation) mice, which are Chx10 null mutants, are microphthalmic and show expanded and abnormal peripheral structures, including the ciliary body. We show here, in a mixed genetic background, the progressive appearance of pigmented cells in the neural retina, concomitant with loss of expression of retinal markers. Fate mapping analysis using a multifunctional Chx10 BAC reporter mouse revealed this process to be direct transdifferentiation of retinal cells into pigmented cells. Microarray and in situ hybridization analyses revealed a complex program underlying the transdifferentiation. This program involved the expansion of expression of genes normally found only in the periphery into central regions of the eye. These genes included a transcription factor controlling pigmentation, Mitf, and the related factor Tfec (Tcfec– Mouse Genome Informatics), which can activate a melanogenic gene expression program. Misexpression of Chx10 in the developing retinal pigmented epithelium (RPE) caused downregulation of Mitf, Tfec, and associated pigment markers, leading to a nonpigmented RPE. These data link Chx10 and Mitf to maintenance of the neural retina and RPE fates respectively. Further, they suggest a new role for Chx10 in maintenance of compartment boundaries in the peripheral retina.