Synergistic inhibition of Notch signaling and forced cell cycle re-entry drive Müller glia reprogramming in uninjured mouse retina

Regenerative organisms, like teleost fish, possess the ability for Müller glia (MG) cells to enter the cell cycle independently following injury and generate functional retinal neurons. However, mammalian MG cells exhibit limited proliferative and regenerative abilities, resulting in irreversible loss of retinal neurons. Various methods have been employed to stimulate proliferation in adult mouse MG, yet most resulting daughter cells retain their glial identity.

The study found that MG daughter cells consistently maintained high Notch signaling, which could limit their neurogenic potential. Deleting Rbpj, the central transcriptional factor of Notch, triggered limited conversion of MG into neurons without any proliferation. Nonetheless, deleting Rbpj in conjunction with the forced proliferation of MG cells through increased expression of Ccnd1 and suppression of Cdkn1b led to significant enhancement of MG dedifferentiation and the expression of neuronal marker Otx2 in MG offspring cells within uninjured mouse retinas.

By combining Notch inhibition with MG cell cycle reactivation, the researchers observed an increase in the production of bipolar and amacrine-like cells derived from MG. Moreover, this approach promoted differentiation into ON-cone, OFF-cone, and rod-bipolar subtypes. Single-nucleus RNA and ATAC sequencing indicated that Notch inhibition facilitated the formation of MG-derived progenitor-like cells, while MG proliferation increased chromatin accessibility of neurogenic genes.

Interestingly, most MG-derived cells persisted long-term with incomplete maturation. Overall, these findings elucidate the roles of Notch inhibition and MG proliferation, singly or in combination, in modulating the regenerative capacity of MG cells in the mammalian retina.