Wang Dan, Guan Hongbo, Xia Yajun
Department of Obstetrics and Gynecology, Shengjing Hospital of China Medical University, Shenyang 110001, Liaoning Province, China.
Department of Obstetrics and Gynecology, Shengjing Hospital of China Medical University, Shenyang 110001, Liaoning Province, China.
Biochem Pharmacol. 2023 Apr;210:115456. doi: 10.1016/j.bcp.2023.115456. Epub 2023 Feb 11.
N6-methyladenosine (m6A) is the most abundant mRNA internal modification in eukaryotic mRNAs. This study focuses on the effect of circMPP1 on placental villi function and the molecular mechanism. First, differentially expressed circular RNAs (circRNAs) in placenta tissues of large-for-gestational-age(LGA) neonates were screened by m6A-circRNA Epitranscriptomic Microarray and bioinformatics analyses. The abnormal expression of circMPP1 in placental tissues and cell lines was validated by RT-qPCR. In-vitro and in-vivo functional experiments were performed to evaluate the role of circMPP1 in placental impairment and fetal dysplasia. The interacting proteins of circMPP1 were identified and validated using RNA pull-down, RNA immunoprecipitation, fluorescence in situ hybridization, and immunofluorescence experiments. Protein interactions and expression levels were detected by Co-immunoprecipitation and western blot analysis. The m6A modification in circMPP1 was verified by methylated RNA immunoprecipitation assay. Bioinformatics analyses showed that circMPP1 was highly expressed in tissues with disordered placental function. In-vitro and in-vivo functional experiments showed that circMPP1 inhibited the function of placental villi. Further mechanism analyses showed that circMPP1 activated the NF-kappa B and MAPK3 signaling pathways. In addition, the m6A "reader" protein YTHDC1 was found to reduce circMPP1 expression via m6A modification. In conclusion, this study demonstrates that YTHDC1 maintains trophoblasts function by promoting degradation of m6A-mediated circMPP1.
N6-甲基腺苷（m6A）是真核生物mRNA中最丰富的mRNA内部修饰。本研究聚焦于circMPP1对胎盘绒毛功能的影响及其分子机制。首先，通过m6A-环状RNA表观转录组微阵列和生物信息学分析筛选出大于胎龄（LGA）新生儿胎盘组织中差异表达的环状RNA（circRNA）。通过RT-qPCR验证了circMPP1在胎盘组织和细胞系中的异常表达。进行体外和体内功能实验以评估circMPP1在胎盘损伤和胎儿发育异常中的作用。使用RNA下拉、RNA免疫沉淀、荧光原位杂交和免疫荧光实验鉴定并验证了circMPP1的相互作用蛋白。通过免疫共沉淀和蛋白质印迹分析检测蛋白质相互作用和表达水平。通过甲基化RNA免疫沉淀测定法验证了circMPP1中的m6A修饰。生物信息学分析表明circMPP1在胎盘功能紊乱的组织中高表达。体外和体内功能实验表明circMPP1抑制胎盘绒毛的功能。进一步的机制分析表明circMPP1激活了NF-κB和MAPK3信号通路。此外，发现m6A“阅读器”蛋白YTHDC1通过m6A修饰降低circMPP1的表达。总之，本研究表明YTHDC1通过促进m6A介导的circMPP1降解来维持滋养层细胞功能。
