Human CRX regulates photoreceptor cells development via bidirectional transcriptional control in retinal organoids.

CRX (cone-rod homeobox) is a key regulator of retinal photoreceptor development, yet its human-specific functions remain poorly understood due to scarce human retinal tissues and significant species differences. Here, we established a human CRX-mCherry fluorescent reporter retinal organoid (RO) model to dissect CRX-mediated gene regulation. Using FACS, RNA sequencing, and Cleavage Under Targets and Tagmentation (CUT&Tag) sequencing, we identified CRX target genes and revealed its dual regulatory role: it activates photoreceptor-specific genes (e.g., RP1L1, linked to inherited retinal degeneration) in a dose-dependent manner, while suppressing non-photoreceptor genes (e.g., PCDH8 and PROX1). Notably, we first generated the human CRX CUT&Tag dataset, providing direct insights into CRX's genome-wide regulatory landscape in photoreceptor cell development. These findings demonstrate that CRX functions as both a transcriptional activator and repressor, ensuring photoreceptor-specific gene expression and preventing aberrant cell fate transitions. Our study provides critical insights into the role of human CRX in retinal development and implications for retinal degenerative diseases.