Abstract
Purpose: :
Around 10% of human individuals with anophthalmia or severe microphthalmia carry a mutation in the gene encoding the HMG-box transcription factor SOX2. Our lab has previously shown that ablation of SOX2 in mouse optic cup progenitor cells results in cell fate conversion from neural retina (NR) to ciliary epithelium (CE) (Matsushima et al. 2011). Although prospective ciliary epithelial cells can be identified as early as embryonic day 12.5, Sox2-mutant cells do not display all the hallmarks of CE progenitor cells until postnatal stages. The current study seeks to understand the mechanism of this gradual cell fate conversion, providing insight into how the boundary between the neural retina and ciliary epithelium is established.
Methods: :
Two potential mechanisms were investigated. First, the immediate read-out of NR-to-CE cell fate conversion is upregulation of the transcription factor PAX6. PAX6 is highly expressed in the prospective CE and has been shown to be important for maintaining the CE progenitor cell pool. DNA sequence analysis and protein-DNA binding assays were used to determine whether SOX2 can bind and regulate the eye-specific enhancer of the Pax6 gene. Secondly, ablation of SOX2 in optic cup progenitor cells results in expanded Wnt signaling. Wnt signaling is an additional potential mediator of CE fate. Epistasis analysis in the mouse was used to determine the genetic relationship between SOX2 and beta-catenin, the major transcriptional mediator of canonical Wnt signaling.
Results: :
Results from DNA binding assays suggest that SOX2 can physically bind the retina-specific enhancer, alpha, of the Pax6 gene in a genomic locus where it may interact with PAX6 protein. Moreover, genetic epistasis analysis shows that SOX2 acts upstream of beta-catenin in optic cup patterning.
Conclusions: :
These results suggest that SOX2 signaling regionalizes the optic cup into prospective NR and CE through at least two mechanisms: First, SOX2 antagonizes Pax6 expression via direct binding to alpha, thereby mediating transcriptional repression of Pax6. This antagonism may set-up the inverse SOX2-PAX6 gradient, with SOX2 high in prospective neural retinal cells and PAX6 high in prospective ciliary epithelial cells. Secondly, SOX2 may antagonize canonical Wnt signaling upstream of beta-catenin, perhaps ensuring that Wnt signaling acrtivity is primarily confiined to the prospective CE. Together, these studies provide insight into how disrupted SOX2 function can lead to improper eye development.
Keywords: retinal development • ciliary body • transcription factors