Ciliary margin transdifferentiation from neural retina is controlled by canonical Wnt signaling

Abstract

The epithelial layers of the ciliary body (CB) and iris are non-neural structures that differentiate from the anterior region of the eyecup, the ciliary margin (CM). We show here that activation of the canonical Wnt signaling pathway is sufficient and necessary for the normal development of anterior eye structures. Pharmacological activation of beta-catenin signaling with lithium (Li(+)) treatment in retinal explants in vitro induced the ectopic expression of the CM markers Otx1 and Msx1. Cre-mediated stabilization of beta-catenin expression in the peripheral retina in vivo induced a cell autonomous upregulation of CM markers at the expense of neural retina (NR) markers and inhibited neurogenesis. Consistent with a cell autonomous conversion to peripheral eye fates, the proliferation index in the region of the retina that expressed stabilized beta-catenin was identical to the wild-type CM and there was an expansion of CB-like structures at later stages. Conversely, Cre-mediated inactivation of beta-catenin reduced CM marker expression as well as the size of the CM and CB/iris. Aberrant CB development in both mouse models was also associated with a reduction in the number of retinal stem cells in vitro. In summary, activation of canonical Wnt signaling is sufficient to promote the development of peripheral eyecup fates at the expense of the NR and is also required for the normal development of anterior eyecup structures.

Loss of adenomatous polyposis coli (apc) results in an expanded ciliary marginal zone in the zebrafish eye

“…As these studies relied on global overexpression studies, they did not differentiate between proliferation rates in the CMZ vs. the central retina. In contrast, other studies performed in chick and mice describe a reduction in proliferation upon constitutive activation of the Wnt/b-catenin pathway (Cho and Cepko, 2006;Liu et al, 2007). Our results that Wnt/b-catenin pathway activation leads to a loss of cyclinD1 and myca expression accompanied by a loss or reduction in BrdU incorporation is in agreement with these later studies.…”

“…In mouse and chick, constitutive activation of the Wnt/b-catenin pathway causes expansion of the ciliary body and iris demonstrating that Wnt/ b-catenin signaling is required for the induction of peripheral fates (Cho and Cepko, 2006;Liu et al, 2007;Kubo and Nakagawa, 2009). We observed that in zebrafish ectopic expression of Wnt target genes leads to the upregulation and expansion of otx1, a marker for the future ciliary epithelium and iris in mice that is normally expressed in the distal tip of the eye …”

“…2A,B; Martinez-Morales et al, 2001;Liu et al, 2007). It is not known if otx1-positive cells only differentiate into peripheral structures, such as the ciliary body and the iris or if they are the stem cells that give rise to peripheral structures as well as the neural retina cells.…”

“…We observed that, in zebrafish apc mutant eyes, cells that express Wnt/ b-catenin pathway members do not co-express retinal identity and proneural genes, such as sox2 and atoh7 suggesting that Wnt/b-catenin signaling inhibits these genes (Kubo et al, 2003(Kubo et al, , 2005Cho and Cepko, 2006;Liu et al, 2007). The neural competence factor Sox2 plays a crucial role in retina development as it is essential for neural specification maintenance of neural progenitor cells (Mizuseki et al, 1998;Bylund et al, 2003;Pevny and Placzek, 2005;Van Raay et al, 2005;Taranova et al, 2006;Agathocleous et al, 2009).…”

“…The Wnt/bcatenin pathway regulates stem cells, proliferation and differentiation in many other systems and also plays an important role in the regulation of the CMZ in vertebrates (Van Raay and Vetter, 2004;Van Raay et al, 2005). Studies in mice and chick have emphasized that activation of the Wnt pathway is required for the development of peripheral structures, such as the ciliary body and the iris, whereas other studies propose that the Wnt pathway also serves an important function in the maintenance of stem cells (Kubo et al, 2003(Kubo et al, , 2005Liu et al, 2007;Denayer et al, 2008;Kubo and Nakagawa, 2009). Here we propose that the zebrafish adenomatous polyposis coli (apc mcr ) mutation, which leads to constitutive up-regulation of the Wnt/b-catenin pathway, is an excellent model to decipher the roles of signaling pathways in the developing eye.…”