Single-cell transcriptomics reveals that RNA pseudouridylation-related gene NHP2 promotes the progression of uveal melanoma and validated in vivo and in vitro.

Uveal melanoma (UM) stands as the predominant primary intraocular malignancy encountered among adults. There is still limited research on the function of RNA pseudouridylation-associated genes in UM. This study identified RNA modification-related genes that play crucial role in UM by single-cell RNA sequencing analysis. We validated their expression in UM tissues and cell lines using immunohistochemistry and Western blotting experiments. Gene knockdown and overexpression experiments were conducted in UM cell lines, and their effects on UM proliferation, migration, invasion and apoptosis were verified using CCK-8, colony formation, Edu, scratch wound healing, Transwell assays and flow cytometry. RNA pseudouridylation (RNA-Ψ) modification-related genes NHP2 were identified to be highly expressed in UM tissues and cells. Cox and Lasso regression analysis suggested that NHP2 had a significant impact on the overall survival probability of UM patients. Knockdown of NHP2 inhibited the proliferation, migration, and invasion of UM cells, and promoted apoptosis, while overexpression resulted in the opposite effects. In vivo experiments confirmed that the elevated expression of NHP2 promoted the growth of UM tumors. The molecular subtypes of UM obtained through unsupervised clustering based on the gene set related to high NHP2 expression effectively distinguished the survival statuses of patients. There was a relatively strong immune response and interaction among cytokines in UM with high-risk state, which may be related to the RNA-Ψ modification mediated by NHP2. The prognostic model constructed relied on NHP2-related molecular subtypes efficiently predicted the survival probability and the metastasis risk of UM patients, and experimental validation confirmed the increased expression of the prognostic genes NENF, ILKAP, and SRD5A3 in clinical tissue samples of UM. NHP2 serves as a biomarker related to RNA-Ψ modification in UM, with increased expression promoting disease progression, indicating poor prognosis for patients, and suggesting its potential role as a molecular target for UM treatment.