Cleavage stimulation factor subunit 2 (CSTF2) is a fundamental factor in the regulation of 3'-end cleavage and alternative polyadenylation of pre-mRNAs. Previous work has identified a tumor-promoting role of CSTF2, suggesting that it may represent a potential therapeutic target. Here, we aimed to elucidate the mechanistic function of CSTF2 in hepatocellular carcinoma (HCC). CSTF2 upregulation was frequent in HCC, and elevated levels of CSTF2 correlated with poor patient prognosis. While CSTF2 inhibition did not suppress HCC growth under non-stress conditions, it supported tolerance and survival of HCC cells under hypoxic conditions. Mechanistically, CSTF2 increased PGK1 protein production to enhance glycolysis, thereby sustaining the energy supply under hypoxic conditions. CSTF2 shortened the 3' untranslated region (3' UTR) of phosphoglycerate kinase 1 (PGK1) pre-mRNA by binding near the proximal polyadenylation site (pPAS). This shortening led to a loss of N6-methyladenosine (m6A) modification sites that are bound by YTH N6-methyladenosine RNA-binding protein F2 (YTHDF2) and increase degradation of PGK1 mRNA. Concurrently, hypoxia increased m6A modification of PGK1 mRNA near the pPAS that was recognized by the YTH N6-methyladenosine RNA-binding protein C1 (YTHDC1), which recruited CSTF2 to enhance the shortening of the PGK1 3’-UTR. A small molecule screen identified masitinib as an inhibitor of CSTF2. Masitinib counteracted PGK1 upregulation by CSTF2 and suppressed the growth of HCC xenograft and patient-derived organoid models. In conclusion, this study revealed a function of CSTF2 in supporting HCC survival under hypoxia conditions through m6A modification evasion and metabolic reprogramming, indicating inhibiting CSTF2 may overcome hypoxia tolerance in HCC.

中文翻译：

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CSTF2 通过使 PGK1 的 m6A 修饰逃避以增强糖酵解来支持肝细胞癌的缺氧耐受性


切割刺激因子亚基 2 （CSTF2） 是调节 3' 端切割和前 mRNA 选择性多聚腺苷酸化的基本因子。以前的工作已经确定了 CSTF2 的肿瘤促进作用，表明它可能代表了一个潜在的治疗靶点。在这里，我们旨在阐明 CSTF2 在肝细胞癌 （HCC） 中的机制功能。CSTF2 在 HCC 中上调频繁，CSTF2 水平升高与患者预后不良相关。虽然 CSTF2 抑制在非应激条件下不抑制 HCC 生长，但它支持 HCC 细胞在缺氧条件下的耐受性和存活。从机制上讲，CSTF2 增加了 PGK1 蛋白的产生以增强糖酵解，从而在缺氧条件下维持能量供应。CSTF2 通过在近端多聚腺苷酸化位点 （pPAS） 附近结合来缩短磷酸甘油酸激酶 1 （PGK1） 前体 mRNA 的 3' 非翻译区 （3' UTR）。这种缩短导致与 YTH N6-甲基腺苷 RNA 结合蛋白 F2 （YTHDF2） 结合的 N6-甲基腺苷 （m6A） 修饰位点丢失，并增加 PGK1 mRNA 的降解。同时，缺氧增加了 pPAS 附近 PGK1 mRNA 的 m6A 修饰，该修饰被 YTH N6-甲基腺苷 RNA 结合蛋白 C1 （YTHDC1） 识别，该蛋白募集 CSTF2 以增强 PGK1 3'-UTR 的缩短。小分子筛选发现马赛替尼是 CSTF2 的抑制剂。Masitinib 抵消 CSTF2 对 PGK1 的上调，并抑制 HCC 异种移植物和患者来源类器官模型的生长。总之，本研究揭示了 CSTF2 通过 m6A 修饰逃避和代谢重编程支持缺氧条件下 HCC 存活的功能，表明抑制 CSTF2 可能克服 HCC 的缺氧耐受。