April N. Smith

Co-operative roles for E-cadherin and N-cadherin during lens vesicle separation and lens epithelial cell survival.

The classical cadherins are known to have both adhesive and signaling functions. It has also been proposed that localized regulation of cadherin activity may be important in cell assortment during development. In the context of eye development, it has been suggested that cadherins are important for separation of the invaginated lens vesicle from the surface ectoderm. To test this hypothesis, we conditionally deleted N-cadherin or E-cadherin from the presumptive lens ectoderm of the mouse. Conditional deletion of either cadherin alone did not produce a lens vesicle separation defect. However, these conditional mutants did exhibit common structural deficits, including microphthalmia, severe iris hyperplasia, persistent vacuolization within the fibre cell region, and eventual lens epithelial cell deterioration. To assess the co-operative roles of E-cadherin and N-cadherin within the developing lens, double conditional knockout embryos were generated. These mice displayed distinct defects in lens vesicle separation and persistent expression of another classical cadherin, P-cadherin, within the cells of the persistent lens stalk. Double mutant lenses also exhibited severe defects in lens epithelial cell adhesion and survival. Finally, the severity of the lens phenotype was shown to be sensitive to the number of wild-type E- and N-cadherin alleles. These data suggest that the co-operative expression of both E- and N-cadherin during lens development is essential for normal cell sorting and subsequent lens vesicle separation.

Stage-dependent modes of Pax6-Sox2 epistasis regulate lens development and eye morphogenesis

The transcription factors Pax6 and Sox2 have been implicated in early events in lens induction and have been proposed to cooperate functionally. Here, we investigated the activity of Sox2 in lens induction and its genetic relationship to Pax6 in the mouse. Conditional deletion of Sox2 in the lens placode arrests lens development at the pit stage. As previously shown, conditional deletion of Pax6 in the placode eliminates placodal thickening and lens pit invagination. The cooperative activity of Sox2 and Pax6 is illustrated by the dramatic failure of lens and eye development in presumptive lens conditional, compound Sox2, Pax6 heterozygotes. The resulting phenotype resembles that of germ line Pax6 inactivation, and the failure of optic cup morphogenesis indicates the importance of ectoderm-derived signals for all aspects of eye development. We further assessed whether Sox2 and Pax6 were required for N-cadherin expression at different stages of lens development. N-cadherin was lost in Sox2-deficient but not Pax6-deficient pre-placodal ectoderm. By contrast, after the lens pit has formed, N-cadherin expression is dependent on Pax6. These data support a model in which the mode of Pax6-Sox2 inter-regulation is stage-dependent and suggest an underlying mechanism in which DNA binding site availability is regulated.

Optic cup and facial patterning defects in ocular ectoderm β-catenin gain-of-function mice

BackgroundThe canonical Wnt signaling pathway has a number of critical functions during embryonic development and, when activated aberrantly, in the genesis of cancer. Current evidence suggests that during eye development, regulation of Wnt signaling is critical for patterning the surface ectoderm that will contribute to multiple components of the eye. Wnt signaling loss-of-function experiments show that a region of periocular ectoderm will form ectopic lentoid bodies unless the Wnt pathway modifies its fate towards other structures. Consistent with this, Wnt signaling gain of function in the ocular region ectoderm results in a suppression of lens fate.ResultsHere we demonstrate that ectoderm-specific Wnt signaling gain-of-function embryos exhibit additional defects besides those noted in the lens. There are profound facial defects including a foreshortened snout, malformation of the nasal region, and clefting of the epidermis along the ocular-nasal axis. Furthermore, despite the restriction of Wnt pathway gain-of-function to the surface ectoderm, the optic cup is inappropriately patterned and ultimately forms a highly convoluted, disorganized array of epithelium with the characteristics of retina and retinal pigmented epithelium.ConclusionWe suggest that activation of the Wnt pathway in surface ectoderm may disrupt the normal exchange of signals between the presumptive lens and retina that coordinate development of a functional eye.

Eye formation in the absence of retina.

Eye development is a complex process that involves the formation of the retina and the lens, collectively called the eyeball, as well as the formation of auxiliary eye structures such as the eyelid, lacrimal gland, cornea and conjunctiva. The developmental requirements for the formation of each individual structure are only partially understood. We have shown previously that the homeobox-containing gene Rx is a key component in eye formation, as retinal structures do not develop and retina-specific gene expression is not observed in Rx-deficient mice. In addition, Rx-/- embryos do not develop any lens structure, despite the fact that Rx is not expressed in the lens. This demonstrates that during normal mammalian development, retina-specific gene expression is necessary for lens formation. In this paper we show that lens formation can be restored in Rx-deficient embryos experimentally, by the elimination of beta-catenin expression in the head surface ectoderm. This suggests that beta-catenin is involved in lens specification either through Wnt signaling or through its function in cell adhesion. In contrast to lens formation, we demonstrate that the development of auxiliary eye structures does not depend on retina-specific gene expression or retinal morphogenesis. These results point to the existence of two separate developmental processes involved in the formation of the eye and its associated structures. One involved in the formation of the eyeball and the second involved in the formation of the auxiliary eye structures.

Which FGF ligands are involved in lens induction

The optic cup ablation experiments of Hans Spemann (Spemann, 1901) introduced the concept of embryonic induction and established that presumptive retina provided signals required for lens development. Since then, lens induction has been a favored subject for developmental biologists wishing to understand the molecular mechanisms of inductive signaling. Much has been learned about the genetic regulation of lens induction and Fig. 1 summarizes some of the advances. The transcription factor Pax6 is centrally involved. It is both necessary (Ashery-Padan et al., 2000; Collinson et al., 2000) and sufficient (Altmann et al., 1997; Chow et al., 1999) for lens development and is induced by the fibroblast growth factor (FGF)(Faber et al., 2001; Gotoh et al., 2004) and bone morphogenetic protein 7 (BMP7) (Wawersik et al., 1999) signaling pathways that are required for early lens formation. Meis and Six family transcription factors have also been implicated in lens induction through their action at the Ectoderm Enhancer of the Pax6 gene (Zhang et al., 2002; Liu et al., 2006). The Sry family transcription factor Sox2 is involved in lens development (Kamachi et al., 2001; Kondoh et al., 2004) and has an essential, parallel function to Pax6 at pre-placodal stages (Smith et al., 2009). Sox2 is up-regulated by BMP4, the first signaling ligand to be implicated in lens induction (Furuta and Hogan, 1998). In a recent analysis, the Grainger group has provided evidence that the broadly expressed transcription factor Otx2 cooperates with the locally expressed Notch pathway transcriptional regulator suppressor of hairless (Su(H)) to up-regulate expression of FoxE3 and define lens placode ectoderm (Ogino et al., 2008). Since Su(H) is dependent on the Notch pathway ligand Delta2 that is expressed in the optic vesicle, this is an example of the type of lens induction signaling that would be anticipated from classical studies.

09-P038 Pax6 roles in lens development

right ventricle and the outflow tract of the heart. Here we have investigated the role of tailup (tup), the Drosophila homologue of vertebrate islet-1 in the development of the dorsal vessel. In Drosophila, cardiac progenitor cells are specified bilaterally in the dorsal mesoderm, which requires complex interactions between the transcription factors Tinman/Nkx2.5, Pannier/GATA factor, and Dorsocross/Tbx factor. Additionally, the growth factors Wingless and Decapentaplegic provide crucial signals for the induction and maintenance of the cardiac regulatory network. Tup mutants are characterized by reduced expression of the essential transcription factors Tin, Pnr and Doc in early cardiogenesis. Conversely, Tup expression depends on each of these cardiac factors, as well as on the early inductive signals Dpp and Wg. Genetic interactions show that tup cooperates with tin, pnr and Doc in cardiogenesis. Lossand gain-of-function analyses for Tup and Pnr suggest that a proper balance of these factors is essential for cardiac specification. Inhibition of Tup in differentiated cardiac cells shows, that later in development, Tup is required to maintain cardiac identities. In summary, our data add tup as a critical factor to the early cardiac regulatory network that determines a cardiac fate.