Role of Otx2 in mature retinal photoreceptors

Although the role of Otx2 transcription factor in retinal development is well understood, its function in the adult retina remains unclear. Otx2 expression is maintained in Retinal Pigmented Epithelium (RPE), photoreceptors (PRs) and bipolar cells throughout life. Previous works from our lab showed that Otx2 ablation in the adult retina leads to photoreceptor degeneration, suggesting a role in their maintenance. It was then showed that RPE-restricted Otx2 expression was both necessary and sufficient for photoreceptor maintenance, indicating a non-cell-autonomous neuroprotective function. This left unexplained the function of endogenous Otx2 in PRs. In order to elucidate it, we performed PR-specific Otx2 knockout, using the inducible PR-specific Crx-CreERT2 mouse line. Histology confirmed that endogenous Otx2 is not required for PR survival or maintenance of their cell identity in the adult. However, we found that Otx2 is implicated in visual adaptation, by regulating the light induced movement of arrestin in photoreceptors. Arrestin shuttles between PR outer segments, where it accumulates in the light, and inner segments and soma, where it accumulates in the dark, to modulate PR sensitivity, in response to different light intensities. Such trafficking is compromised in PR-specific Otx2 KO mice, where arrestin remains blocked in inner segments and soma, even after prolonged light exposure. Behavioural tests showed that PR-specific Otx2 KO mice are strongly photophobic. The detailed mechanisms remain to be elucidated.To decipher the genetic network controlled by Otx2 in PRs, we carried out time series RNA-seq analyses following PR-specific Otx2 knockout. No photo-transduction gene was found to be deregulated. Instead of that, we found down-regulation of extracellular matrix genes and, surprisingly, of a group of RPE-specific genes involved in melanogenesis, genes that were already known to be direct targets of Otx2. Careful examination of PR-specific Otx2 KO mice confirmed that Otx2 protein level in the RPE nuclei was reduced while some Otx2protein could now be detected in PR outer segments. According to such data and to the neuroprotective effect of RPE-restricted Otx2, we hypothesized a direct Otx2 protein transfer from RPE to PRs. To prove this hypothesis, viral vectors driving tagged-Otx2 expression specifically in RPE cells were generated and used for sub-retinal injection in mice. After RPE transduction, tagged-Otx2 could be detected in the PRs. We found that in wild type retina, constitutive Otx2 transfer is active at low rate. After KO induction in the PRs, the transfer rate is increased, probably acting as a neuroprotective response. Such increment in the transfer explains well the reduction of Otx2 protein level in the RPE and the subsequent downregulation of melanogenesis genes. Transferred Otx2 appears to travel across PR cells from the inner and outer segments to the synapse in the outer plexiform layer, suggesting its neuroprotective function involves mechanisms that are different from its classical genomic function of transcription factor.In conclusion, this study disclosed a new function of endogenous Otx2 in PR light adaptation and demonstrated the existence of a transfer of Otx2 from RPE to PR cells, with a putative neuroprotective role.