Thyroid hormone signaling specifies cone subtypes in human retinal organoids

The mechanisms underlying specification of neuronal subtypes within the human nervous system are largely unknown. The blue/S, green/M and red/L cones of the retina enable high-acuity daytime and color vision. To determine the mechanism controlling S vs. L/M fates, we studied the differentiation of human retinal organoids. Organoids and retinas have similar distributions, expression profiles, and morphologies of cone subtypes. S cones are specified first, followed by L/M cones, and thyroid hormone signaling controls this temporal switch. Dynamic expression of thyroid hormone-degrading and activating proteins within the retina ensures low signaling early to specify S cones and high signaling late to produce L/M cones. This work establishes organoids as a model for determining mechanisms of human development with promising utility for therapeutics and vision repair. EP1 iPSC-derived organoids were analyzed at time points ranging from day 10 to day 250 of differentiation. We took samples at day 10 (n=3), day 20 (n=2), day 35 (n=3), day 69 (n=3), day 111 (n=3), day 128 (n=3), day 158 (n=2), day 173 (n=3), day 181 (n=3), day 200 (n=3), and day 250 7 (n=3). RNA from individual organoids was extracted using the Zymo Direct-zol RNA Microprep Kit (Zymo Research) according to manufacturer’s instructions. Individual libraries were prepared for each sample using the Illumina TruSeq stranded mRNA kit and sequenced on an Illumina NextSeq 500 with single 200 bp reads.