Tsg101 Is Necessary for the Establishment and Maintenance of Mouse Retinal Pigment Epithelial Cell Polarity.

The retinal pigment epithelium (RPE) forms a monolayer sheet separating the retina and choroid in vertebrate eyes. The polarized nature of RPE is maintained by distributing membrane proteins differentially along apico-basal axis. We found the distributions of these proteins differ in embryonic, post-natal, and mature mouse RPE, suggesting developmental regulation of protein trafficking. Thus, we deleted tumor susceptibility gene 101 (Tsg101), a key component of endosomal sorting complexes required for transport (ESCRT), in embryonic and mature RPE to determine whether ESCRT-mediated endocytic protein trafficking correlated with the establishment and maintenance of RPE polarity. Loss of Tsg101 severely disturbed the polarity of RPE, which forms irregular aggregates exhibiting non-polarized distribution of cell adhesion proteins and activation of epidermal growth factor receptor signaling. These findings suggest that ESCRT-mediated protein trafficking is essential for the development and maintenance of RPE cell polarity.

mTORC1 accelerates retinal development via the immunoproteasome.

The numbers and types of cells constituting vertebrate neural tissues are determined by cellular mechanisms that couple neurogenesis to the proliferation of neural progenitor cells. Here we identified a role of mammalian target of rapamycin complex 1 (mTORC1) in the development of neural tissue, showing that it accelerates progenitor cell cycle progression and neurogenesis in mTORC1-hyperactive tuberous sclerosis complex 1 (Tsc1)-deficient mouse retina. We also show that concomitant loss of immunoproteasome subunit Psmb9, which is induced by Stat1 (signal transducer and activator of transcription factor 1), decelerates cell cycle progression of Tsc1-deficient mouse retinal progenitor cells and normalizes retinal developmental schedule. Collectively, our results establish a developmental role for mTORC1, showing that it promotes neural development through activation of protein turnover via a mechanism involving the immunoproteasome.

Hippo Signaling Circuit and Divergent Tissue Growth in Mammalian Eye.

Vertebrate organ development is accompanied by demarcation of tissue compartments, which grow coordinately with their neighbors. Hence, perturbing the coordinative growth of neighboring tissue compartments frequently results in organ malformation. The growth of tissue compartments is regulated by multiple intercellular and intracellular signaling pathways, including the Hippo signaling pathway that limits the growth of various organs. In the optic neuroepithelial continuum, which is partitioned into the retina, retinal pigment epithelium (RPE) and ciliary margin (CM) during eye development, the Hippo signaling activity operates differentially, as it does in many tissues. In this review, we summarize recent studies that have explored the relationship between the Hippo signaling pathway and growth of optic neuroepithelial compartments. We will focus particularly on the roles of a tumor suppressor, neurofibromin 2 (NF2), whose expression is not only dependent on compartment-specific transcription factors, but is also subject to regulation by a Hippo-Yap feedback signaling circuit.

Differential Expression of NF2 in Neuroepithelial Compartments Is Necessary for Mammalian Eye Development.

The optic neuroepithelial continuum of vertebrate eye develops into three differentially growing compartments: the retina, the ciliary margin (CM), and the retinal pigment epithelium (RPE). Neurofibromin 2 (Nf2) is strongly expressed in slowly expanding RPE and CM compartments, and the loss of mouse Nf2 causes hyperplasia in these compartments, replicating the ocular abnormalities seen in human NF2 patients. The hyperplastic ocular phenotypes were largely suppressed by heterozygous deletion of Yap and Taz, key targets of the Nf2-Hippo signaling pathway. We also found that, in addition to feedback transcriptional regulation of Nf2 by Yap/Taz in the CM, activation of Nf2 expression by Mitf in the RPE and suppression by Sox2 in retinal progenitor cells are necessary for the differential growth of the corresponding cell populations. Together, our findings reveal that Nf2 is a key player that orchestrates the differential growth of optic neuroepithelial compartments during vertebrate eye development.

The Retinal Pigment Epithelium Is a Notch Signaling Niche in the Mouse Retina.

Notch signaling in neural progenitor cell is triggered by ligands expressed in adjacent cells. To identify the sources of active Notch ligands in the mouse retina, we negatively regulated Notch ligand activity in various neighbors of retinal progenitor cells (RPCs) by eliminating mindbomb E3 ubiquitin protein ligase 1 (Mib1). Mib1-deficient retinal cells failed to induce Notch activation in intra-lineage RPCs, which prematurely differentiated into neurons; however, Mib1 in post-mitotic retinal ganglion cells was not important. Interestingly, Mib1 in the retinal pigment epithelium (RPE) also contributed to Notch activation in adjacent RPCs by supporting the localization of active Notch ligands at RPE-RPC contacts. Combining this RPE-driven Notch signaling and intra-retinal Notch signaling, we propose a model in which one RPC daughter receives extra Notch signals from the RPE to become an RPC, whereas its sister cell receives only a subthreshold level of intra-retinal Notch signal and differentiates into a neuron.

Mitochondrial Protection by Exogenous Otx2 in Mouse Retinal Neurons.

OTX2 (orthodenticle homeobox 2) haplodeficiency causes diverse defects in mammalian visual systems ranging from retinal dysfunction to anophthalmia. We find that the retinal dystrophy of Otx2(+/GFP) heterozygous knockin mice is mainly due to the loss of bipolar cells and consequent deficits in retinal activity. Among bipolar cell types, OFF-cone bipolar subsets, which lack autonomous Otx2 gene expression but receive Otx2 proteins from photoreceptors, degenerate most rapidly in Otx2(+/GFP) mouse retinas, suggesting a neuroprotective effect of the imported Otx2 protein. In support of this hypothesis, retinal dystrophy in Otx2(+/GFP) mice is prevented by intraocular injection of Otx2 protein, which localizes to the mitochondria of bipolar cells and facilitates ATP synthesis as a part of mitochondrial ATP synthase complex. Taken together, our findings demonstrate a mitochondrial function for Otx2 and suggest a potential therapeutic application of OTX2 protein delivery in human retinal dystrophy.