DIO3 coordinates photoreceptor development timing and fate stability in human retinal organoids

How neuronal subtypes are generated at distinct times and proportions during human retinal development is poorly understood. Here, we investigated how rod and cone photoreceptor subtypes develop in human retinal organoids. We find that type 3 iodothyronine deiodinase (DIO3), an enzyme that degrades thyroid hormone (TH), is a master regulator of human photoreceptor developmental timing and cell fate stability. DIO3 is highly expressed in retinal progenitor cells (RPCs) and decreases as cells asynchronously differentiate into neurons, progressively lowering TH degradation during development. DIO3 mutant organoids display precocious S cone, L/M cone, and rod development, increased photoreceptor (PR) density, and conversion of S cones to L/M fate. Cell autonomous and non-autonomous mechanisms regulate DIO3 expression to maintain and locally coordinate levels among cells. In a computational model, a mechanism that couples TH levels and fate specification provides robustness to photoreceptor development compared to a probabilistic, cell-intrinsic mechanism. Our findings suggest a population-based mechanism for photoreceptor subtype developmental timing in which the relative proportion of progenitors to neurons decreases over time to relieve degradation of TH signaling, which advances photoreceptor development. Overall design: Wildtype human retinal organoids or DIO3?33 human retinal organoids were lysed into a single nuclei suspension and run on the 10x Genomics Chromium Single Cell System platform using the Single Cell Multiome ATAC + Gene Expression chemistry. Libraries were sequenced on Illumina NovaSeq 6000 and processed through the Cell Ranger pipeline using the GRCh38 reference genome.