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Qiutang Li, Patrick A Scott, Eric Vukmanic, Lei Xue, Douglas C Dean, Henry J Kaplan, Qingxian Lu; Yap1 is required for maintaining adult RPE differentiation. Invest. Ophthalmol. Vis. Sci. 2017;58(8):1043. doi: https://doi.org/.
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© ARVO (1962-2015); The Authors (2016-present)
Transcriptional coactivator YAP1 is a major downstream effector of the evolutionarily conserved Hippo pathway that controls organ size by regulating stem cell/progenitor cell proliferation and apoptosis during early development. The importance of YAP1 to the eye was further illustrated by the discovery of the heterozygous mutations in YAP1 causing coloboma in man, and the mutation in YAP1 binding domain of TEAD1 responsible for Sveinsson's chorioretinal atrophy (SCRA). However, YAP1 function in maintenance of RPE phenotype has not been investigated. This study aims to dissect YAP1 function and its regulatory mechanism in RPE cells.
To directly test the role of YAP1 in the differentiated RPE, we crossed Yap1 floxed mice with these carrying a Cre recombinase driven by the Best1 promoter, which is activated in the differentiated RPE cells. We investigated the phenotypes progressively developed in the Yap1-/- RPE in a timecourse up to 12 months old, using (1) EM for ultrastructure; (2) Immunohistochemistry for markers delineating RPE differentiation status, cell polarity, cell contact, microvilli organization, and cytoskeletal arrangement; (3) RPE barrier assay with fluorescent angiography; and (4) ERG for visual function. Additionally, we also performed the qPCR and Western blot to characterize molecular changes including these in the Wnt/β-catenin signaling pathway.
Yap1 mutation in differentiated RPE caused diminished tight junctions, cell depolarization and RPE65 loss, retraction of microvilli and loss of pigment. These phenotypes are classic features of epithelial-mesenchymal transition (EMT), which are also evident in RPE during proliferative vitreoretinopathy (PVR). When getting older, the RPE-conditional Yap1 knockout mice exhibited impaired barrier function of RPE cells, disrupted and diminished choriocapillaris, outer segment loss in the adjacent photoreceptors, and abnormal electrophysiology. The photoreceptors in the conditional knockout mice eventually died between 6-12 months of age. Interestingly, β-catenin expression and its transcriptional activity was significantly increased in the mutant RPE.
YAP1 plays a critical role in maintenance of the RPE differentiating phenotype by downregulation of nuclear β-catenin; and thus, maintaining a constant inhibition of Wnt/β-catenin signaling prevents dedifferentiating EMT in normal adult RPE.
This is an abstract that was submitted for the 2017 ARVO Annual Meeting, held in Baltimore, MD, May 7-11, 2017.
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