Elucidating the Role of the Electron Transport Chain in Retinal Progenitor Cells and Neuro‐ regeneration

Abstract

In the retinas of non‐mammalian vertebrates, such as zebrafish, neural regeneration is mediated by the molecular reprogramming of MGs into progenitor‐like cells. While mammalian MGs do not exhibit the regenerative potential of the zebrafish, it has been shown that MGs of mammals display limited‐ and transient‐entry into the cell cycle in response to damage. Therefore, an intrinsic proliferative and/or regenerative block likely prevents mammalian MGs from undergoing sustained cell cycle re‐entry and acquiring a multipotential retinal progenitor‐like state. Using a gain‐of‐function strategy we bypassed the Hippo signaling pathway and induced expression of a mutant version of YAP (YAP5SA) in MGs. Lineage tracing and histological analyses of these YAP5SA‐mutant retinas showed that MGs that expressed YAP5SA entered the cell cycle and formed radial clusters of proliferative cells strikingly similar to clonally expanding MG‐derived progenitors of the regenerating zebrafish retina. This finding indicated that YAP5SA is capable of spontaneously driving MGs into the cell cycle. Whether this YAP‐mediated reprogramming of MGs is sufficient to induce differentiation of MGs into fully functional neurons is undetermined. Subsequent single cell RNA sequencing analysis of the YAP5SA‐mutant retinas revealed that the proliferative event of YAP5SA‐positive cells occurred coincident with a dramatic downregulation of genes required for mitochondrial metabolic processes indicating that a metabolic shift is part of the MG reprogramming event. In a variety of cellular contexts, mitochondrial function and the electron transport chain (ETC) activity have emerged as integral regulators of the cell cycle and cell fate. The factors regulating a mitochondrial metabolic reprogramming and cell cycle are coupled but unknown. We generated a mouse line to conditionally knock out (CKO) the mitochondrial transcription factor A (Tfam) to target the ETC. In the CKO developing retinas, loss of Tfam results in loss of ETC activity. Preliminary histological analysis of CKO developing retinas show that loss of Tfam delays progenitor cell cycle exit and reduces neurogenesis resulting in a hypoplastic adult retina. Our findings so far indicate that unless retinal progenitor cells have a functional ETC‐driven oxidative activity, they remain within the cell cycle compromising the normal neural developmental program. The overall goal of this studies will be to determine how the RPC cell cycle machinery interfaces with ETC to drive cell cycle exit and neural differentiation. A better understanding of these requirements will have broad implications for the entire field of neural regeneration.