State Key Laboratory of Ophthalmology, Zhongshan Ophthalmic Center, Sun Yat-sen University, Guangzhou City, China.
Department of Chemistry, New York University, New York, New York, United States.
Invest Ophthalmol Vis Sci. 2021 Aug 2;62(10):3. doi: 10.1167/iovs.62.10.3.
Retinoblastoma (RB) is the most common primary malignant intraocular cancer. The etiology of RB is complex, and the mechanisms driving its progression remain unclear. Here, we used a series of bioinformatics approaches and experimental methods to investigate the potential regulatory mechanism involved in RB progression.
The common differentially expressed genes were obtained from the public dataset GSE97508. Protein-protein interaction (PPI) network, correlation, and functional enrichment analyses were carried out. The candidate genes were verified in different RB cell lines, and ARPE19 cells served as control. miRNA-mRNA interaction analysis was performed and confirmed by real-time PCR. The CCK-8 assay was conducted to detect cell viability, and the transwell assay was utilized for evaluating the abilities of cell migration and invasion.
Overall, a total of 258 common differentially expressed genes associated with RB progression were screened out. The PPI network analysis further identified eight downregulated genes mainly enriched in the protein ubiquitination pathway. Moreover, we confirmed UBE2E1, SKP1, FBXO9, FBXO15, and RNF14 from among eight genes through experimental validation in vitro. Furthermore, miRNA-mRNA interaction and real-time PCR analysis of five hub genes revealed that ubiquitination-related miR-548k was involved in RB progression. Loss- and gain-of-function experiments demonstrated that miR-548k and its targets were essential for cell viability, migration, and invasion in the RB cells.
Our data indicate that the dysregulation of protein ubiquitination may play an important role in RB progression, and ubiquitination-related miR-548k may be a promising therapeutic target for RB.
视网膜母细胞瘤（RB）是最常见的原发性眼内恶性肿瘤。RB 的病因复杂，其进展的机制尚不清楚。在这里，我们使用了一系列生物信息学方法和实验方法来研究 RB 进展中涉及的潜在调控机制。
从公共数据集 GSE97508 中获得常见的差异表达基因。进行蛋白质-蛋白质相互作用（PPI）网络、相关性和功能富集分析。在不同的 RB 细胞系中验证候选基因，ARPE19 细胞作为对照。进行 miRNA-mRNA 相互作用分析，并通过实时 PCR 进行验证。通过 CCK-8 检测细胞活力，通过 Transwell 检测细胞迁移和侵袭能力。
总共筛选出与 RB 进展相关的 258 个常见差异表达基因。PPI 网络分析进一步确定了 8 个下调基因，这些基因主要富集在蛋白质泛素化途径中。此外，我们通过体外实验验证了这 8 个基因中的 UBE2E1、SKP1、FBXO9、FBXO15 和 RNF14。此外，通过对 5 个关键基因的 miRNA-mRNA 相互作用和实时 PCR 分析发现，泛素化相关的 miR-548k 参与了 RB 的进展。失活和功能获得实验表明，miR-548k 及其靶基因对于 RB 细胞的活力、迁移和侵袭至关重要。
我们的数据表明，蛋白质泛素化的失调可能在 RB 的进展中发挥重要作用，泛素化相关的 miR-548k 可能是 RB 的一个有前途的治疗靶点。
