CircMYH9 通过调节丝氨酸代谢和依赖 p53 的氧化还原平衡来驱动结直肠癌的生长。

CircMYH9 drives colorectal cancer growth by regulating serine metabolism and redox homeostasis in a p53-dependent manner.

机构信息

Department of Endocrinology, Affiliated Hospital of Xuzhou Medical University, Xuzhou, 221000, China.

Institute of Digestive Diseases, Xuzhou Medical University, Xuzhou, 221002, China.

出版信息

Mol Cancer. 2021 Sep 8;20(1):114. doi: 10.1186/s12943-021-01412-9.

DOI:10.1186/s12943-021-01412-9

PMID:34496888

原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC8424912/

Abstract

BACKGROUND

Circular RNAs (circRNAs) play important roles in cancer progression and metabolism regulation. Serine/glycine metabolism supports the growth of cancer cells by contributing to their anabolic demands and epigenome as well as by regulating their redox state. However, the role of circRNA in the regulation of serine/glycine metabolism has not been well elucidated.

METHODS

Microarray analysis was used to screen differentially expressed novel circRNAs. qRT-PCR and FISH were utilized to analyzed the expression of circMYH9. CCK8, colony formation and FACS were used to analyze proliferation of colorectal cancer (CRC) cells. Xenograft experiments were used to analyze tumor growth in vivo. RNA-sequencing, immunoblot and LC-MS were used to identify the downstream metabolic pathway of circMYH9. ChIRP, Mass Spectrometry, RIP and RNA pulldown were utilized to test the interaction between circMYH9, hnRNPA2B1 and p53 pre-mRNA. ChIP-qPCR was used to analyze the binding sites of HIF-1α. Chemically-induced CRC mice were generated to evaluate the role of circMYH9 in tumorigenesis.

RESULTS

We identified an intron-derived circRNA, circMYH9, which was significantly upregulated in CRC tissues. A higher circMYH9 level correlated with shorter relapse-free survival and overall survival of CRC patients. CircMYH9 promoted serine/glycine metabolism, the NAD + /NADH ratio, and glutathione recycling and inhibited reactive oxygen species (ROS) in a p53-dependent manner, impacting tumour growth. Mechanistically, circMYH9 destabilized the pre-mRNA of p53 by recruiting hnRNPA2B1 in the nucleus. hnRNPA2B1 bound to N6-methyladenosine sites on the 3' untranslated region of p53 pre-mRNA and maintained its stability. Moreover, a lack of amino acids led to an elevated level of ROS, resulting in increased HIF1α, which promoted circMYH9 expression by binding to the promoter region. Furthermore, in vivo AAV9-mediated transfection of circMYH9 could drive chemically-induced carcinogenesis by suppressing p53 in mice.

CONCLUSIONS

The overexpression of circMYH9 promotes CRC proliferation though modulating serine/glycine metabolism and redox homeostasis in a p53-dependent manner, and targeting circMYH9 and its pathway may be an effective strategy for the treatment of CRC.

摘要

背景

环状 RNA（circRNAs）在癌症进展和代谢调控中发挥重要作用。丝氨酸/甘氨酸代谢通过为癌细胞的合成需求以及表观基因组提供支持，并调节其氧化还原状态，从而促进癌细胞的生长。然而，circRNA 在调节丝氨酸/甘氨酸代谢中的作用尚未得到充分阐明。

方法

采用微阵列分析筛选差异表达的新型 circRNAs。qRT-PCR 和 FISH 用于分析 circMYH9 的表达。CCK8、集落形成和 FACS 用于分析结直肠癌细胞（CRC）的增殖。异种移植实验用于分析体内肿瘤生长。RNA 测序、免疫印迹和 LC-MS 用于鉴定 circMYH9 的下游代谢途径。ChIRP、质谱、RIP 和 RNA 下拉用于测试 circMYH9、hnRNPA2B1 和 p53 前体 mRNA 之间的相互作用。ChIP-qPCR 用于分析 HIF-1α 的结合位点。通过化学诱导的 CRC 小鼠生成来评估 circMYH9 在肿瘤发生中的作用。

结果

我们鉴定了一种内含子衍生的 circRNA，circMYH9，其在 CRC 组织中显著上调。circMYH9 水平较高与 CRC 患者的无复发生存期和总生存期较短相关。circMYH9 以 p53 依赖的方式促进丝氨酸/甘氨酸代谢、NAD+/NADH 比和谷胱甘肽再循环，并抑制活性氧（ROS），从而影响肿瘤生长。机制上，circMYH9 通过在核内募集 hnRNPA2B1 来稳定 p53 的前体 RNA。hnRNPA2B1 结合到 p53 前体 RNA 的 3'非翻译区的 N6-甲基腺苷位点，并维持其稳定性。此外，缺乏氨基酸会导致 ROS 水平升高，从而导致 HIF1α 增加，HIF1α 通过结合启动子区域促进 circMYH9 的表达。此外，体内 AAV9 介导的 circMYH9 转染可以通过抑制小鼠中的 p53 来驱动化学诱导的致癌作用。