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METTL3 promotes the osteogenic differentiation of human periodontal ligament cells by increasing YAP activity via IGF2BP1 and YTHDF1-mediated m6A modification

IF 3.4 3区医学 Q1 DENTISTRY, ORAL SURGERY & MEDICINE

Journal of periodontal research Pub Date : 2024-06-04 DOI:10.1111/jre.13297

Xuefei Sun, Xiujiao Meng, Yu Piao, Shaojie Dong, Qianqian Dong

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引用次数: 0

Abstract

Aims

N6-Methyladenosine (m⁶A) has been confirmed to play a dynamic role in osteoporosis and bone metabolism. However, whether m⁶A is involved in the osteogenic differentiation of human periodontal ligament cells (hPDLCs) remains unclear. The present study aimed to verify the role of methyltransferase-like 3 (METTL3)-mediated m⁶A modification in the osteogenic differentiation of hPDLCs.

Methods

The METTL3, Runx2, Osx, and YAP mRNA expression was determined by qPCR. METTL3, RUNX2, OSX, YTHDF1, YAP, IGF2BP1, and eIF3a protein expression was measured by Western blotting and immunofluorescence assays. The levels of m⁶A modification were evaluated by methylated RNA immunoprecipitation (MeRIP) and dot blot analyses. MeRIP-seq and RNA-seq were used to screen potential candidate genes. Nucleic acid and protein interactions were detected by immunoprecipitation. Alizarin red staining was used to evaluate the osteogenic differentiation of hPDLCs. Gene transcription and promoter activities were assessed by luciferase reporter assays (n ≥ 3).

Results

The expression of METTL3 and m⁶A modifications increased synchronously with the osteogenic differentiation of hPDLCs (p = .0016). YAP was a candidate gene identified by MeRIP-seq and RNA-seq, and its mRNA and protein expression levels were simultaneously increased. METTL3 increased the m⁶A methylated IGF2BP1-mediated stability of YAP mRNA (p = .0037), which in turn promoted osteogenic differentiation (p = .0147). Furthermore, METTL3 increased the translation efficiency of YAP by recruiting YTHDF1 and eIF3a to the translation initiation complex (p = .0154), thereby promoting the osteogenic differentiation of hPDLCs (p = .0012).

Conclusion

Our study revealed that METTL3-initiated m⁶A mRNA methylation promotes osteogenic differentiation of hPDLCs by increasing IGF2BP1-mediated YAP mRNA stability and recruiting YTHDF1 and eIF3a to the translation initiation complex to increase YAP mRNA translation. Our findings reveal the mechanism of METTL3-mediated m⁶A modification during hPDLC osteogenesis, providing a potential therapeutic target for periodontitis and alveolar bone defects.

Abstract Image

查看原文本刊更多论文

METTL3 通过 IGF2BP1 和 YTHDF1 介导的 m6A 修饰提高 YAP 活性，从而促进人类牙周韧带细胞的成骨分化。

目的：N6-甲基腺苷（m6A）已被证实在骨质疏松症和骨代谢中发挥动态作用。然而，m6A是否参与人牙周韧带细胞（hPDLCs）的成骨分化仍不清楚。本研究旨在验证甲基转移酶样3（METTL3）介导的m6A修饰在hPDLCs成骨分化中的作用：方法：通过qPCR检测METTL3、Runx2、Osx和YAP mRNA的表达。METTL3、RUNX2、OSX、YTHDF1、YAP、IGF2BP1和eIF3a蛋白的表达通过Western印迹和免疫荧光测定。甲基化 RNA 免疫沉淀（MeRIP）和点印迹分析评估了 m6A 修饰的水平。MeRIP-seq 和 RNA-seq 被用来筛选潜在的候选基因。通过免疫沉淀检测核酸和蛋白质的相互作用。茜素红染色用于评估hPDLCs的成骨分化。基因转录和启动子活性通过荧光素酶报告实验进行评估（n ≥ 3）：结果：METTL3和m6A修饰的表达与hPDLCs的成骨分化同步增加（p = .0016）。YAP是MeRIP-seq和RNA-seq鉴定出的候选基因，其mRNA和蛋白表达水平同时升高。METTL3 增加了 YAP mRNA 的 m6A 甲基化 IGF2BP1 介导的稳定性（p = .0037），这反过来又促进了成骨分化（p = .0147）。此外，METTL3通过招募YTHDF1和eIF3a进入翻译起始复合物，提高了YAP的翻译效率（p = .0154），从而促进了hPDLCs的成骨分化（p = .0012）：我们的研究发现，METTL3引发的m6A mRNA甲基化通过增加IGF2BP1介导的YAP mRNA稳定性，并招募YTHDF1和eIF3a进入翻译起始复合物以增加YAP mRNA翻译，从而促进hPDLCs的成骨分化。我们的研究结果揭示了 METTL3 在 hPDLC 成骨过程中介导 m6A 修饰的机制，为牙周炎和牙槽骨缺损提供了一个潜在的治疗靶点。

本文章由计算机程序翻译，如有差异，请以英文原文为准。

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来源期刊

Journal of periodontal research

Journal of periodontal research 医学-牙科与口腔外科

CiteScore

6.90

自引率

5.70%

发文量

103

审稿时长

6-12 weeks

期刊介绍： The Journal of Periodontal Research is an international research periodical the purpose of which is to publish original clinical and basic investigations and review articles concerned with every aspect of periodontology and related sciences. Brief communications (1-3 journal pages) are also accepted and a special effort is made to ensure their rapid publication. Reports of scientific meetings in periodontology and related fields are also published. One volume of six issues is published annually.

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