Summary Mutations in the photoreceptor transcription factor gene cone-rod homeobox (CRX) lead to distinct retinopathy phenotypes, including early-onset vision impairment in dominant Leber congenital amaurosis (LCA). Using induced pluripotent stem cells (iPSCs) from a patient with CRX -I138fs48 mutation, we established an in vitro model of CRX -LCA in retinal organoids that showed defective photoreceptor maturation by histology and gene profiling, with diminished expression of visual opsins. Adeno-associated virus (AAV)-mediated CRX gene augmentation therapy partially restored photoreceptor phenotype and expression of phototransduction-related genes as determined by single-cell RNA-sequencing. Retinal organoids derived from iPSCs of a second dominant CRX -LCA patient carrying K88N mutation revealed the loss of opsin expression as a common phenotype, which was alleviated by AAV-mediated augmentation of CRX. Our studies provide a proof-of-concept for developing gene therapy of dominant CRX -LCA and other CRX retinopathies.

Diapause is a programmed dormancy that allows organisms to tolerate predictable periods of unfavorable conditions by temporarily halting development and reducing metabolism. Diapause is widespread among insects and is crucial for allowing organisms to coordinate their growth and reproduction with favorable environmental conditions. While the adaptive significance of diapause is well-understood, the molecular mechanisms underpinning diapause remain unresolved. We performed high-throughput sequencing to investigate the role of microRNAs in the diapause of the Asian tiger mosquito, Aedes albopictus. We first investigate miRNAs in diapause induction by characterizing maternally-provisioned miRNAs in mature oocytes of Ae. albopictus under diapause-inducing and diapause-averting conditions. Second, we investigate miRNAs in diapause maintenance by characterizing miRNAs in diapause and non-diapause pharate larvae. We identified 162 miRNAs, 152 previously known and 10 putatively novel. We identified no differentially abundant miRNAs in mature oocytes and seven differentially abundant miRNAs in pharate larvae. The predicted targets of differentially abundant miRNAs include genes affecting several processes related to diapause maintenance including ecdysone regulation, immune response, lipid metabolism, and regulation of development. Our results suggest that Ae. albopictus does not maternally-provision a unique set of miRNAs during diapause induction but miRNAs are a component of diapause maintenance in this species.

Chromatin organization and enhancer-promoter contacts establish unique spatiotemporal gene expression patterns in distinct cell types. Non-coding genetic variants can influence cellular phenotypes by modifying higher-order transcriptional hubs and consequently gene expression. To elucidate genomic regulation in human retina, we mapped chromatin contacts at high resolution and integrated with super-enhancers (SEs), histone marks, binding of CTCF and select transcription factors. We show that topologically associated domains (TADs) with central SEs exhibit stronger insulation and augmented contact with retinal genes relative to TADs with edge SEs. Merging genome-wide expression quantitative trait loci (eQTLs) with topology map reveals physical links between 100 eQTLs and corresponding eGenes associated with retinal neurodegeneration. Additionally, we uncover candidate genes for susceptibility variants linked to age-related macular degeneration and glaucoma. Our study of high-resolution genomic architecture of human retina provides insights into genetic control of tissue-specific functions, suggests paradigms for missing heritability, and enables the dissection of common blinding disease phenotypes.