目的：探讨RNA结合基序蛋白15（RBM15）是否通过含跨膜Bax抑制基序蛋白6（TMBIM6）/TGF-β/Smad轴促进宫颈癌细胞增殖、迁移及侵袭。方法：收集2023年1月至2025年1月在苏州大学附属张家港医院接受手术治疗的82例宫颈癌患者的肿瘤组织与癌旁组织。应用R语言分析癌症基因组图谱（TCGA）数据库中RBM15、TMBIM6 mRNA的表达；采用免疫组织化学法和Western blotting法检测RBM15、TMBIM6蛋白表达；采用Kaplan-Meier曲线分析RBM15、TMBIM6 mRNA表达与宫颈癌患者预后的关系；采用CCK-8法、划痕实验及Transwell侵袭实验检测RBM15、TMBIM6对宫颈癌细胞增殖、迁移及侵袭的影响；利用基于序列的RNA腺苷甲基化位点预测工具（SRAMP）预测TMBIM6 mRNA的N6-甲基腺苷（m6A）修饰位点；采用RNA免疫共沉淀（RIP）实验、RNA衰变和挽救实验鉴定RBM15与TMBIM6 mRNA的相互作用；采用甲基化RNA免疫共沉淀定量PCR（MeRIP-qPCR）检测TMBIM6 mRNA的m6A富集；采用Western blotting法检测RBM15通过TMBIM6对宫颈癌细胞TGF-β/Smad通路相关蛋白表达的影响。结果：宫颈癌组织中RBM15 mRNA、TMBIM6 mRNA表达均高于癌旁组织（均P < 0.05），癌组织RBM15、TMBIM6阳性率均高于癌旁组织（均P < 0.001）。宫颈癌细胞系HeLa、MS-751、SiHa中RBM15、TMBIM6蛋白表达均高于正常宫颈上皮细胞系Ect1/E6E7（均P < 0.05）。国际妇产科联盟（FIGO）分期ⅠB2～ⅡB期宫颈癌组织中RBM15、TMBIM6蛋白阳性率高于ⅠA～ⅠB1期（P均 < 0.05）。RBM15 mRNA高表达组、TMBIM6 mRNA高表达组5年无进展生存率分别低于各自的低表达组（均P < 0.05）。宫颈癌组织中RBM15 mRNA与TMBIM6 mRNA表达呈正相关（P < 0.05）。SRAMP预测结果显示，TMBIM6 mRNA序列的960、24876、25203位点为高可信度m6A修饰位点。在HeLa、SiHa细胞中过表达RBM15后，TMBIM6mRNA和蛋白表达升高（P < 0.05）；敲低RBM15后，TMBIM6 mRNA和蛋白表达降低（P < 0.05）。与IgG组相比，RBM15抗体免疫沉淀物中TMBIM6 mRNA显著富集。MeRIP-qPCR结果显示，TMBIM6 mRNA的960、24876、25203位点均存在明显的m6A富集。敲低RBM15可缩短TMBIM6 mRNA半衰期并降低其稳定性。敲低RBM15可抑制宫颈癌细胞增殖、迁移及侵袭，过表达TMBIM6可逆转上述影响。在HeLa、SiHa细胞中过表达TMBIM6后，TGF-β1、Smad2、Smad3蛋白表达升高；敲低RBM15后，上述蛋白表达降低，过表达TMBIM6可逆转该影响。结论：RBM15可能通过m6A修饰提高TMBIM6 mRNA稳定性并激活TGF-β/Smad通路，从而促进宫颈癌细胞增殖、迁移及侵袭。

Objective: To investigate whether RNA binding motif protein 15 (RBM15) promotes cervical cancer cell proliferation, migration, and invasion through the transmembrane Bax inhibitor motif-containing 6 (TMBIM6)/TGF-β/Smad axis. Methods: Tumor tissues and adjacent tissues were collected from 82 cervical cancer patients who underwent surgical treatment at the Affiliated Zhangjiagang Hospital of Soochow University from January 2023 to January 2025. R was used to analyze RBM15 and TMBIM6 mRNA expression in The Cancer Genome Atlas (TCGA) database. Immunohistochemistry and Western blotting assays were used to detect RBM15 and TMBIM6 protein expression. Kaplan-Meier curves were used to analyze the associations of RBM15 and TMBIM6 mRNA expression with the prognosis of patients with cervical cancer. Cell Counting Kit-8 (CCK-8), scratch, and Transwell invasion assays were used to evaluate the effects of RBM15 and TMBIM6 on cervical cancer cell proliferation, migration, and invasion. The sequence-based RNA adenosine methylation site predictor (SRAMP) was used to predict N6-methyladenosine (m6A) modification sites in TMBIM6 mRNA. RNA immunoprecipitation (RIP), RNA decay, and rescue assays were performed to investigate the interaction between RBM15 and TMBIM6 mRNA. Methylated RNA immunoprecipitation quantitative PCR (MeRIP-qPCR) was used to detect m6A enrichment in TMBIM6 mRNA. Western blotting assays were used to assess the effects of RBM15 and TMBIM6 on TGF-β/Smad pathway-related protein expression. Results: RBM15 and TMBIM6 mRNA expression levels in cervical cancer tissues were higher than those in adjacent noncancerous tissues (both P < 0.05), and the positivity rates of RBM15 and TMBIM6 were higher in cancer tissues than in adjacent noncancerous tissues (both P < 0.001). RBM15 and TMBIM6 protein expression levels in HeLa, MS-751, and SiHa cells were higher than those in Ect1/E6E7 cells (all P < 0.05). The positivity rates of RBM15 and TMBIM6 in cervical cancer tissues at FIGO stages IB2-IIB were higher than those at stages IA-IB1 (both P < 0.05). The 5-year progression-free survival rates in the RBM15 and TMBIM6 high-expression groups were lower than those in their respective low-expression groups (both P < 0.05). RBM15 mRNA expression was positively correlated with TMBIM6 mRNA expression in cervical cancer tissues (P < 0.05). SRAMP predicted high-confidence m6A modification sites at positions 960, 24876, and 25203 in TMBIM6 mRNA. RBM15 overexpression increased, whereas RBM15 knockdown decreased, TMBIM6 mRNA and protein expression in HeLa and SiHa cells (P < 0.05). TMBIM6 mRNA was significantly enriched in RBM15 immunoprecipitates compared with the IgG group. MeRIP-qPCR showed significant m6A enrichment at positions 960, 24876, and 25203 of TMBIM6 mRNA. RBM15 knockdown shortened the half-life and reduced the stability of TMBIM6 mRNA. RBM15 knockdown inhibited cervical cancer cell proliferation, migration, and invasion, whereas TMBIM6 overexpression reversed these effects. TMBIM6 overexpression increased TGF-β1, Smad2, and Smad3 protein expression, whereas RBM15 knockdown decreased their expression; TMBIM6 overexpression reversed the effects of RBM15 knockdown. Conclusion: The results suggest that RBM15 may increase TMBIM6 mRNA stability through m6A modification and activate the TGF-β/Smad pathway, thereby promoting cervical cancer cell proliferation, migration, and invasion.

江苏省妇幼健康科研项目（F202167）