PDF(Pubmed)
Abstract：
UNASSIGNED: N6-methyladenosine (m6A) methylation is a chemical modification that occurs on RNA molecules, where the hydrogen atom of adenine (A) nucleotides is replaced by a methyl group, forming N6-methyladenosine. This modification is a dynamic and reversible process that plays a crucial role in regulating various biological processes, including RNA stability, transport, translation, and degradation. Currently, there is a lack of research on the role of m6A modifications in maintaining the characteristics of RPE cells. m6A readers play a crucial role in executing the functions of m6A modifications, which prompted our investigation into their regulatory roles in the RPE.
UNASSIGNED: Phagocytosis assays, immunofluorescence staining, flow cytometry experiments, β-galactosidase staining, and RNA sequencing (RNA-seq) were conducted to assess the functional and cellular characteristics changes in retinal pigment epithelium (RPE) cells following short-hairpin RNA-mediated knockdown of insulin-like growth factor 2 mRNA-binding protein 2 (IGF2BP2). RNA-seq and ultraviolet crosslinking immunoprecipitation with high-throughput sequencing (HITS-CLIP) were employed to identify the target genes regulated by IGF2BP2. adeno-associated virus (AAV) subretinal injection was performed in 6- to 8-week-old C57 mice to reduce IGF2BP2 expression in the RPE, and the impact of IGF2BP2 knockdown on mouse visual function was assessed using immunofluorescence, quantitative real-time PCR, optical coherence tomography, and electroretinography.
UNASSIGNED: IGF2BP2 was found to have a pronounced effect on RPE phagocytosis. Subsequent in-depth exploration revealed that IGF2BP2 modulates the mRNA stability of PAX6 and OTX2, and the loss of IGF2BP2 induces inflammatory and aging phenotypes in RPE cells. IGF2BP2 knockdown impaired RPE function, leading to retinal dysfunction in vivo.
UNASSIGNED: Our data suggest a crucial role of IGF2BP2 as an m6A reader in maintaining RPE homeostasis by regulating the stability of PAX6 and OTX2, making it a potential target for preventing the occurrence of retinal diseases related to RPE malfunction.
UNASSIGNED: Phagocytosis assays, immunofluorescence staining, flow cytometry experiments, β-galactosidase staining, and RNA sequencing (RNA-seq) were conducted to assess the functional and cellular characteristics changes in retinal pigment epithelium (RPE) cells following short-hairpin RNA-mediated knockdown of insulin-like growth factor 2 mRNA-binding protein 2 (IGF2BP2). RNA-seq and ultraviolet crosslinking immunoprecipitation with high-throughput sequencing (HITS-CLIP) were employed to identify the target genes regulated by IGF2BP2. adeno-associated virus (AAV) subretinal injection was performed in 6- to 8-week-old C57 mice to reduce IGF2BP2 expression in the RPE, and the impact of IGF2BP2 knockdown on mouse visual function was assessed using immunofluorescence, quantitative real-time PCR, optical coherence tomography, and electroretinography.
UNASSIGNED: IGF2BP2 was found to have a pronounced effect on RPE phagocytosis. Subsequent in-depth exploration revealed that IGF2BP2 modulates the mRNA stability of PAX6 and OTX2, and the loss of IGF2BP2 induces inflammatory and aging phenotypes in RPE cells. IGF2BP2 knockdown impaired RPE function, leading to retinal dysfunction in vivo.
UNASSIGNED: Our data suggest a crucial role of IGF2BP2 as an m6A reader in maintaining RPE homeostasis by regulating the stability of PAX6 and OTX2, making it a potential target for preventing the occurrence of retinal diseases related to RPE malfunction.
摘要：
■N6-甲基腺苷（m6A）甲基化是发生在RNA分子上的化学修饰，其中腺嘌呤（A）核苷酸的氢原子被甲基取代，形成N6-甲基腺苷。这种修饰是一个动态和可逆过程，在调节各种生物过程中起着至关重要的作用，包括RNA稳定性，运输,翻译,和退化。目前,缺乏对m6A修饰在维持RPE细胞特性中的作用的研究。M6A阅读器在执行M6A修改的功能中起着至关重要的作用，这促使我们调查他们在RPE中的监管作用。
■吞噬作用测定，免疫荧光染色，流式细胞术实验,β-半乳糖苷酶染色,和RNA测序（RNA-seq）进行评估短发夹RNA介导的胰岛素样生长因子2mRNA结合蛋白2（IGF2BP2）敲低后视网膜色素上皮（RPE）细胞的功能和细胞特征变化。采用RNA-seq和高通量测序的紫外线交联免疫沉淀（HITS-CLIP）来鉴定IGF2BP2调节的靶基因。在6至8周龄的C57小鼠中进行腺相关病毒（AAV）视网膜下注射，以减少RPE中IGF2BP2的表达，使用免疫荧光评估IGF2BP2敲低对小鼠视觉功能的影响，实时定量PCR,光学相干层析成像，和视网膜电图。
■发现IGF2BP2对RPE吞噬作用具有明显作用。随后的深入探索表明，IGF2BP2调节PAX6和OTX2的mRNA稳定性，IGF2BP2的缺失诱导RPE细胞的炎症和衰老表型。IGF2BP2敲低受损的RPE功能，导致体内视网膜功能障碍。
■我们的数据表明，IGF2BP2作为m6A阅读器在通过调节PAX6和OTX2的稳定性来维持RPE稳态方面的关键作用，使其成为预防视网膜疾病发生的潜在目标与RPE功能障碍有关。
■吞噬作用测定，免疫荧光染色，流式细胞术实验,β-半乳糖苷酶染色,和RNA测序（RNA-seq）进行评估短发夹RNA介导的胰岛素样生长因子2mRNA结合蛋白2（IGF2BP2）敲低后视网膜色素上皮（RPE）细胞的功能和细胞特征变化。采用RNA-seq和高通量测序的紫外线交联免疫沉淀（HITS-CLIP）来鉴定IGF2BP2调节的靶基因。在6至8周龄的C57小鼠中进行腺相关病毒（AAV）视网膜下注射，以减少RPE中IGF2BP2的表达，使用免疫荧光评估IGF2BP2敲低对小鼠视觉功能的影响，实时定量PCR,光学相干层析成像，和视网膜电图。
■发现IGF2BP2对RPE吞噬作用具有明显作用。随后的深入探索表明，IGF2BP2调节PAX6和OTX2的mRNA稳定性，IGF2BP2的缺失诱导RPE细胞的炎症和衰老表型。IGF2BP2敲低受损的RPE功能，导致体内视网膜功能障碍。
■我们的数据表明，IGF2BP2作为m6A阅读器在通过调节PAX6和OTX2的稳定性来维持RPE稳态方面的关键作用，使其成为预防视网膜疾病发生的潜在目标与RPE功能障碍有关。
