Article ID | Journal | Published Year | Pages | File Type |
---|---|---|---|---|
5531862 | Developmental Biology | 2017 | 11 Pages |
â¢Regulatory regions 5â² of the six6 gene drive optic cup and mature retinal expression.â¢A 3â² regulatory region is sufficient for eye field and optic vesicle six6 expression.â¢Several trans-acting factors that regulate endogenous six6 expression were identified.â¢Deletion of the 3â² enhancer reduces eye size.
The eye field transcription factor, Six6, is essential for both the early (specification and proliferative growth) phase of eye formation, as well as for normal retinal progenitor cell differentiation. While genomic regions driving six6 optic cup expression have been described, the sequences controlling eye field and optic vesicle expression are unknown. Two evolutionary conserved regions 5â² and a third 3â² to the six6 coding region were identified, and together they faithfully replicate the endogenous X. laevis six6 expression pattern. Transgenic lines were generated and used to determine the onset and expression patterns controlled by the regulatory regions. The conserved 3â² region was necessary and sufficient for eye field and optic vesicle expression. In contrast, the two conserved enhancer regions located 5â² of the coding sequence were required together for normal optic cup and mature retinal expression. Gain-of-function experiments indicate endogenous six6 and GFP expression in F1 transgenic embryos are similarly regulated in response to candidate trans-acting factors. Importantly, CRISPR/CAS9-mediated deletion of the 3â² eye field/optic vesicle enhancer in X. laevis, resulted in a reduction in optic vesicle size. These results identify the cis-acting regions, demonstrate the modular nature of the elements controlling early versus late retinal expression, and identify potential regulators of six6 expression during the early stages of eye formation.