Depressive disorder (DD) is associated with N6-methyladenosine (m6A) hypermethylation. This study sought to explore the molecular mechanism of Methyltransferase-like 3 (METTL3) in cognitive deficits of chronic unpredictable mild stress (CUMS)-treated rats and provide novel targets for DD treatment. A DD rat model was established via CUMS treatment. Cognitive deficits were assessed via body weighing and behavioral tests. METTL3, microRNA (miR)-221-3p, pri-miR-221, GRB2-associated binding protein 1 (Gab1) expressions in hippocampal tissues were detected via RT-qPCR and Western blotting. m6A, DiGeorge syndrome critical region gene 8 (DGCR8)-bound pri-miR-221 and pri-miR-221 m6A levels were measured. The binding relationship between miR-221-3p and Gab1 was testified by dual-luciferase and RNA pull-down assays. Rescue experiments were designed to confirm the role of miR-221-3p and Gab1. METTL3 was highly expressed in CUMS rats, and silencing METTL3 attenuated cognitive deficits of CUMS rats. METTL3-mediated m6A modification facilitated processing and maturation of pri-miR-221 via DGCR8 to upregulate miR-221-3p. miR-221-3p targeted Gab1. miR-221-3p overexpression or Gab1 downregulation reversed the role of silencing METTL3 in CUMS rats. Overall, METTL3-mediated m6A modification facilitated processing and maturation of pri-miR-221 to upregulate miR-221-3p and then inhibit Gab1, thereby aggravating cognitive deficits of CUMS rats.

中文翻译：

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METTL3介导的m6A修饰在抑郁症引起的认知缺陷中的机制

抑郁症 (DD) 与 N6-甲基腺苷 (m6A) 高甲基化有关。本研究旨在探讨甲基转移酶样 3 (METTL3) 在慢性不可预知轻度应激 (CUMS) 治疗大鼠认知缺陷中的分子机制，并为 DD 治疗提供新的靶点。通过CUMS处理建立DD大鼠模型。通过体重和行为测试评估认知缺陷。通过 RT-qPCR 和蛋白质印迹检测海马组织中 METTL3、microRNA (miR)-221-3p、pri-miR-221、GRB2 相关结合蛋白 1 (Gab1) 的表达。测量了 m6A、DiGeorge 综合征关键区域基因 8 (DGCR8) 结合的 pri-miR-221 和 pri-miR-221 m6A 水平。miR-221-3p 和 Gab1 之间的结合关系通过双荧光素酶和 RNA pull-down 分析得到证实。救援实验旨在确认 miR-221-3p 和 Gab1 的作用。METTL3 在 CUMS 大鼠中高表达，沉默 METTL3 可减轻 CUMS 大鼠的认知缺陷。METTL3 介导的 m6A 修饰通过 DGCR8 促进 pri-miR-221 的加工和成熟，从而上调 miR-221-3p。miR-221-3p 靶向 Gab1。miR-221-3p 过表达或 Gab1 下调逆转了 CUMS 大鼠中 METTL3 沉默的作用。总体而言，METTL3 介导的 m6A 修饰促进了 pri-miR-221 的加工和成熟，从而上调 miR-221-3p，然后抑制 Gab1，从而加重 CUMS 大鼠的认知缺陷。METTL3 介导的 m6A 修饰通过 DGCR8 促进 pri-miR-221 的加工和成熟，从而上调 miR-221-3p。miR-221-3p 靶向 Gab1。miR-221-3p 过表达或 Gab1 下调逆转了 CUMS 大鼠中 METTL3 沉默的作用。总体而言，METTL3 介导的 m6A 修饰促进了 pri-miR-221 的加工和成熟，从而上调 miR-221-3p，然后抑制 Gab1，从而加重 CUMS 大鼠的认知缺陷。METTL3 介导的 m6A 修饰通过 DGCR8 促进 pri-miR-221 的加工和成熟，从而上调 miR-221-3p。miR-221-3p 靶向 Gab1。miR-221-3p 过表达或 Gab1 下调逆转了 CUMS 大鼠中 METTL3 沉默的作用。总体而言，METTL3 介导的 m6A 修饰促进了 pri-miR-221 的加工和成熟，从而上调 miR-221-3p，然后抑制 Gab1，从而加重 CUMS 大鼠的认知缺陷。