我校新桥医院杨仕明教授等在《Molecular Cancer》发表研究成果—— CircMRPS35通过招募KAT7调控靶基因FOXO1/3a的组蛋白修饰进而抑
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CircMRPS35通过招募KAT7调控靶基因FOXO1/3a的组蛋白修饰进而抑制胃癌进展的研究

节梦梦 吴亚冉 胡长江 杨仕明

环状RNA(circRNA)是一类具有共价闭合环状结构的新型非编码RNA,其在多种生物信号通路中起着至关重要的调控作用。与线性mRNA不同,circRNA没有5’-3’极性和poly A尾,可耐受RNase R酶的降解,且在体内具有高度稳定性,在哺乳动物细胞中丰富、保守、具有组织特异性和发育特异性。既往研究表明circRNA可以通过与miRNA、蛋白等相互作用参与调控复杂生物过程。CircRNA在不同肿瘤细胞中具有高度特异性表达模式,提示其可能是恶性肿瘤的潜在诊断或预后标志物。文献表明circRNA在多种肿瘤中表达失调,其表达异常可影响肿瘤的发生发展。深入研究circRNA在胃癌发生发展中的可能作用及机制,将为评价胃癌诊断标志物、治疗靶点及预后评估提供理论依据。

本研究通过circRNA-seq技术检测人的胃癌和相应癌旁组织中circRNA的表达谱,筛选出一条在胃癌组织中显著低表达的circMRPS35 (hsa_circ_0000384),其由MRPS35基因的2,3,4,5号外显子反向剪切环化形成,并通过发散与内敛引物扩增、Oligo dT与随机引物反转录、RNase R处理、放线菌素D处理及测序等方法验证其环状特性。进一步分析circMRPS35的表达与160位胃癌患者临床病理特征的关系,发现circMRPS35表达水平与TNM分期、淋巴结转移及肿瘤大小呈负相关,且低表达circMRPS35的胃癌患者生存时间明显缩短。体内体外实验均表明过表达circMRPS35显著抑制胃癌细胞的增殖和侵袭转移。RNA-seq及GO/Pathway信号通路分析发现circMRPS35对FOXO通路影响最显著。荧光原位杂交及细胞核浆分离实验证实circMRPS35主要在细胞核中富集,萤光素酶报告基因实验进一步表明circMRPS35可通过激活FOXO1/3a的转录及其下游靶基因表达,从而发挥抗胃癌增殖及侵袭转移作用。RNA pull-down结合质谱分析、蛋白截短实验证实circMRPS35可与组蛋白乙酰转移酶KAT7(256-315aa)结合。ChIP实验发现circMRPS35可上调FOXO1/3a启动子区H4K5的乙酰化水平,而敲降KAT7可逆转上述变化。

本研究结果表明circMRPS35可通过招募组蛋白乙酰转移酶KAT7,促进FOXO1和FOXO3a启动子区的H4K5乙酰化水平,从而上调FOXO1、FOXO3a的转录活性,最终抑制胃癌的增殖和侵袭转移。该研究首次发现circRNA直接特异性影响靶基因调控区域的乙酰化水平,为抗胃癌策略提供新思路。

该研究成果于2020年03月发表于《Molecular Cancer》(IF=10.679)。陆军军医大学第二附属医院消化内科胡长江副教授和杨仕明教授为该文通讯作者,节梦梦博士和吴亚冉硕士为该文第一作者。该研究得到国家重点研发项目和国家自然科学基金等研究项目的资助。

附:英文摘要

CircMRPS35 Suppresses Gastric Cancer Progression via Recruiting KAT7 to Govern Histone Modification

Mengmeng Jie 1, Yaran Wu 1, Mengyuan Gao 1, Xinzhe Li 1, Cheng Liu 1, Qin Ouyang 2, Qingyun Tang 1, Changyu Shan 2, Yangfan Lv 3, Kebin Zhang 4, Qian Dai 4, Yang Chen 1, Shuo Zeng 1, Chenglin Li 1, Liting Wang 5, Fengtian He 6, Changjiang Hu 7, Shiming Yang 8

affiliations

1Department of Gastroenterology, Xinqiao Hospital, Third Military Medical University, Chongqing, 400037, China.

2Department of Medicinal Chemistry, College of Pharmacy, Third Military Medical University, Chongqing, 400038, China.

3Department of Pathology, Xinqiao Hospital, Third Military Medical University, Chongqing, 400037, China.

4Central Laboratory, Xinqiao Hospital, Third Military Medical University, Chongqing, 400037, China.

5Biomedical Analysis Center, Third Military Medical University, Chongqing, 400038, China.

6Department of Biochemistry and Molecular Biology, College of Basic Medical Sciences, Third Military Medical University, Chongqing, 400038, China.

7Department of Gastroenterology, Xinqiao Hospital, Third Military Medical University, Chongqing, 400037, China. hcj_888@126.com.

8Department of Gastroenterology, Xinqiao Hospital, Third Military Medical University, Chongqing, 400037, China. Yangshiming@tmmu.edu.cn.

PMID: 32164722

PMCID: PMC7066857

DOI: 10.1186/s12943-020-01160-2

Background: Aberrant expression of circular RNAs contributes to the initiation and progression of cancers, but the underlying mechanism remains elusive.

Methods: RNA-seq and qRT-PCR were performed to screen differential expressed circRNAs between gastric cancer tissues and adjacent normal tissues. Candidate circRNA (circMRPS35) was screened out and validated by qRT-PCR. Cell proliferation and invasion ability were determined by CCK-8 and cell invasion assays. RNA-seq, GO-pathway, RNA pull-down and ChIRP were further applied to search for detailed mechanism.

Results: Here, a novel circRNA named circMRPS35, was screened out by RNA-seq in gastric cancer tissues, whose expression is related to clinicopathological characteristics and prognosis in gastric cancer patients. Biologically, circMRPS35 suppresses the proliferation and invasion of gastric cancer cells in vitro and in vivo. Mechanistically, circMRPS35 acts as a modular scaffold to recruit histone acetyltransferase KAT7 to the promoters of FOXO1 and FOXO3a genes, which elicits acetylation of H4K5 in their promoters. Particularly, circMRPS35 specifically binds to FOXO1/3a promoter regions directly. Thus, it dramatically activates the transcription of FOXO1/3a and triggers subsequent response of their downstream target genes expression, including p21, p27, Twist1 and E-cadherin, resulting in the inhibition of cell proliferation and invasion. Moreover, circMRPS35 expression positively correlates with that of FOXO1/3a in gastric cancer tissues.

Conclusions: Our findings not only reveal the pivotal roles of circMRPS35 in governing histone modification in anticancer treatment, but also advocate for triggering circMRPS35/KAT7/FOXO1/3a pathway to combat gastric cancer.

Keywords: Acetylation; Circular RNA; Gastric cancer; Histone modification.

全文链接:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7066857/

 


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