The Transcription Factor 12 of Basic Helix-Loop-Helix Plays an Essential Role in Retinal Health.

Purpose: To study a non-redundant role of Tcf12 in retinal health. Methods: A loss-of-function mutation in Tcf12 was identified by applying optical coherence tomography (OCT) to a forward genetic pipeline. CRISPR/Cas9-generated Tcf12ra/ra mice, expressing a replacement allele ("ra") were used to validate the findings from the mutagenesis screen. Retinal morphology was assessed using OCT, fundus photography, histology, and immunohistochemistry. Retinal function was evaluated by electroretinography (ERG). Bulk RNA sequencing and proteomic analyses were performed, with select targets validated by RT-qPCR and immunoblotting. Results: Tcf12ra/ra mice exhibited outer nuclear layer thinning on OCT, which was confirmed by histology. Fundus imaging revealed age-dependent accumulation of retinal fundus spots. Subretinal accumulation of Iba1⁺/Tmem119⁺ cells was observed, many of which stained positively for Gal3, suggestive of activated resident microglia. ERG deficits were noticed at 12 to 15 months of age in Tcf12ra/ra mice. Transcriptomic and proteomic profiling using two independent pathway analyses identified dysregulated pathways related to protein and RNA metabolism, mitochondrial and energy metabolism, cell cycle, signal transduction, cellular response to stimuli, and inflammation. RT-qPCR results showed upregulation of Tmem233 and downregulation of Doc2b, Alpk2, Agr2, and Apobec2 in Tcf12ra/ra retinas. Western blot analysis demonstrated upregulation of Selenbp1 and downregulation of Neurod1 and Faah in Tcf12ra/ra retinas. Conclusions: Deficiency in Tcf12 induces early-onset retinal structural alterations, and late-onset subretinal microglial activation and functional decline. Widespread dysregulation in metabolic and signaling pathways was documented in transcriptomic and proteomic analyses. These findings establish Tcf12 as a key contributor to retinal development and homeostasis.