R102W mutation of RS1 gene regulates cell fates of neural retinal cells with abnormal cell communication in early XLRS retinal organoids

Abstract

Purpose : This study is to decipher how the patient-specific mutation R102W in RS1 regulated the differentiation and maturation of retinal cells in patient iPSC-derived retinal organoids (ROs) at an early stage.

Methods : The iPSCs were reprogrammed from the urine cells of XLRS patients and controls, then differentiated into retinal organoids with the 3D differentiation method. Bulk RNA-seq and single-cell RNA-seq (scRNA-seq) were performed on D50 ROs.

Results : ScRNA-Seq analysis showed that, in the R102W group, the proportion of retinal progenitor cells (RPC) and photoreceptor precursors (PR-P) were higher than control, while retinal ganglion cells (RGC) and amacrine cells (AC)/horizon cells (HC) were lower than control. The expression of CRX and NRL, the key transcription factors of photoreceptors, was lower in R102W than those of the control (Fig.1A, B). Bulk RNA-seq showed that 2916 DEGs were identified in R102W versus control. The core genes of 4 representative GO BP terms were selected to obtain heatmap (Fig. 1C) and confirmed in scRNA-seq data (Fig.1D). SNAI2 and FN1, COL3A1 and COL1A1, and S100A11 and HLA-DRB1 were upregulated in R102W, and these markers were enriched in mesenchymal cell cluster. While axonogenesis-related genes MAP2 and GAP43 were downregulated, and the markers were enriched in neural cell clusters (Fig.1C-E).In secreted signaling category, the intensity of the neurotrophin (NT) signaling pathway was higher in control. In the cell-cell contact category, JAM signaling, which plays an important role in retinal development, was significantly higher in control. However, in the ECM category, fibronectin (FN1) signaling, which was associated with fibrogenesis, was not significant in control, while was mainly concentrated in PR-P and RPC in the R102W group (Fig.1F-H).

Conclusions : The R102W mutation of RS1 is sociated with the delay of neurodevelopment and aberrant organization of extracellular structures such as fibrogenesis at the early stage of XLRS ROs.

This abstract was presented at the 2023 ARVO Annual Meeting, held in New Orleans, LA, April 23-27, 2023.

 

Figure1: D50 R0s bulk RNA-seq and scRNA-seq analysis. A: Proportion of each cell type. B: Expression level of key factors in each cell type. C: Heatmap of core DEGs in bulk RNA-seq. D: The position of DEGs in scRNA-seq clusters. E: The qPCR results. F-H: Alterations of intercellular communication patterns include NT(F), JAM(G), and FN1(H) signaling pathway network, between control and R102W ROs.