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Abstract：
BACKGROUND: KIAA1429, a regulatory subunit of the N6-methyladenosine (m6A) methyltransferase complex, has been implicated in the progression of various cancers. However, the role of KIAA1429 in gastric cancer (GC) and its underlying mechanisms remain elusive. This study aimed to investigate the role of KIAA1429 in GC and to elucidate the underlying mechanisms.
METHODS: The expression patterns and clinical relevance of KIAA1429 in GC were assessed using quantitative real-time PCR (qRT-PCR), Western blotting, immunohistochemistry (IHC), and bioinformatic analysis. In vitro and in vivo loss- and gain-of-function assays, m6A dot blot assays, methylated RNA immunoprecipitation sequencing (MeRIP-seq), RNA-seq, MeRIP-qPCR, dual luciferase reporter assays, RNA stability assays, RNA immunoprecipitation (RIP) assays, and RNA pull-down assays were performed to investigate the biological functions and underlying molecular mechanisms of KIAA1429 in GC.
RESULTS: Both the mRNA and protein expression of KIAA1429 were greater in GC tissues than in normal gastric tissues. High KIAA1429 expression correlated positively with poor prognosis in GC patients. KIAA1429 not only promoted GC cell proliferation, colony formation, G2/M cell cycle transition, migration, and invasion in vitro but also enhanced GC tumor growth and metastasis in vivo. Mechanistically, KIAA1429 increased the m6A level of RASD1 mRNA and enhanced its stability in an m6A-YTHDF2-dependent manner, thereby upregulating its expression. RASD1 knockdown partially rescued the KIAA1429 knockdown-induced impairment of pro‑oncogenic ability in GC cells. The expression levels of KIAA1429 and RASD1 were negatively correlated in GC tissues.
CONCLUSIONS: KIAA1429 plays a pro‑oncogenic role in GC by downregulating RASD1 expression through destabilizing RASD1 mRNA in an m6A-YTHDF2-dependent manner. KIAA1429 may serve as a prognostic biomarker and therapeutic target for GC.
METHODS: The expression patterns and clinical relevance of KIAA1429 in GC were assessed using quantitative real-time PCR (qRT-PCR), Western blotting, immunohistochemistry (IHC), and bioinformatic analysis. In vitro and in vivo loss- and gain-of-function assays, m6A dot blot assays, methylated RNA immunoprecipitation sequencing (MeRIP-seq), RNA-seq, MeRIP-qPCR, dual luciferase reporter assays, RNA stability assays, RNA immunoprecipitation (RIP) assays, and RNA pull-down assays were performed to investigate the biological functions and underlying molecular mechanisms of KIAA1429 in GC.
RESULTS: Both the mRNA and protein expression of KIAA1429 were greater in GC tissues than in normal gastric tissues. High KIAA1429 expression correlated positively with poor prognosis in GC patients. KIAA1429 not only promoted GC cell proliferation, colony formation, G2/M cell cycle transition, migration, and invasion in vitro but also enhanced GC tumor growth and metastasis in vivo. Mechanistically, KIAA1429 increased the m6A level of RASD1 mRNA and enhanced its stability in an m6A-YTHDF2-dependent manner, thereby upregulating its expression. RASD1 knockdown partially rescued the KIAA1429 knockdown-induced impairment of pro‑oncogenic ability in GC cells. The expression levels of KIAA1429 and RASD1 were negatively correlated in GC tissues.
CONCLUSIONS: KIAA1429 plays a pro‑oncogenic role in GC by downregulating RASD1 expression through destabilizing RASD1 mRNA in an m6A-YTHDF2-dependent manner. KIAA1429 may serve as a prognostic biomarker and therapeutic target for GC.
摘要：
背景：KIAA1429，N6-甲基腺苷（m6A）甲基转移酶复合物的调节亚基，与各种癌症的进展有关。然而,KIAA1429在胃癌（GC）中的作用及其潜在机制仍然难以捉摸。本研究旨在探讨KIAA1429在GC中的作用并阐明其潜在机制。
方法：使用定量实时PCR（qRT-PCR）评估KIAA1429在GC中的表达模式和临床相关性，西方印迹,免疫组织化学（IHC），和生物信息学分析。体外和体内功能损失和增益测定，M6A斑点印迹分析，甲基化RNA免疫沉淀测序（MeRIP-seq），RNA-seq,MeRIP-qPCR,双荧光素酶报告分析，RNA稳定性分析,RNA免疫沉淀（RIP）测定，并进行了RNA下拉测定，以研究KIAA1429在GC中的生物学功能和潜在的分子机制。
结果：GC组织中KIAA1429的mRNA和蛋白表达均高于正常胃组织。KIAA1429高表达与GC患者预后不良呈正相关。KIAA1429不只增进GC细胞增殖，菌落形成,G2/M细胞周期过渡，迁移,和体外侵袭，但也增强了体内GC肿瘤的生长和转移。机械上,KIAA1429以m6A-YTHDF2依赖性方式增加RASD1mRNA的m6A水平并增强其稳定性，从而提高其表达。RASD1敲低部分挽救了KIAA1429敲低诱导的GC细胞原癌能力受损。GC组织中KIAA1429和RASD1的表达水平呈负相关。
结论：KIAA1429通过以m6A-YTHDF2依赖性方式使RASD1mRNA不稳定而下调RASD1表达，从而在GC中发挥原癌作用。KIAA1429可作为GC的预后生物标志物和治疗靶标。
方法：使用定量实时PCR（qRT-PCR）评估KIAA1429在GC中的表达模式和临床相关性，西方印迹,免疫组织化学（IHC），和生物信息学分析。体外和体内功能损失和增益测定，M6A斑点印迹分析，甲基化RNA免疫沉淀测序（MeRIP-seq），RNA-seq,MeRIP-qPCR,双荧光素酶报告分析，RNA稳定性分析,RNA免疫沉淀（RIP）测定，并进行了RNA下拉测定，以研究KIAA1429在GC中的生物学功能和潜在的分子机制。
结果：GC组织中KIAA1429的mRNA和蛋白表达均高于正常胃组织。KIAA1429高表达与GC患者预后不良呈正相关。KIAA1429不只增进GC细胞增殖，菌落形成,G2/M细胞周期过渡，迁移,和体外侵袭，但也增强了体内GC肿瘤的生长和转移。机械上,KIAA1429以m6A-YTHDF2依赖性方式增加RASD1mRNA的m6A水平并增强其稳定性，从而提高其表达。RASD1敲低部分挽救了KIAA1429敲低诱导的GC细胞原癌能力受损。GC组织中KIAA1429和RASD1的表达水平呈负相关。
结论：KIAA1429通过以m6A-YTHDF2依赖性方式使RASD1mRNA不稳定而下调RASD1表达，从而在GC中发挥原癌作用。KIAA1429可作为GC的预后生物标志物和治疗靶标。
