N6-甲基腺苷的生物学功能及检测技术研究进展

doi:10.3969/j.issn.1673-8640.2022.09.016

摘要/Abstract

摘要：

N6-甲基腺苷（m6A）是哺乳动物细胞mRNA中丰度最高的一种动态可逆的修饰,由甲基化酶[甲基转移酶（METTL）3、甲基转移酶（METTL）14、肾母细胞瘤1关联蛋白（WTAP）]、去甲基化酶[肥胖基因相关蛋白（FTO）、α-酮戊二酸依赖性双加氧酶同源物（ALKBH）5]、甲基结合蛋白[YTH N6甲基腺苷RNA结合蛋白1（YTHDF1）、YTH结构域蛋白（YTHDC）1、YTHDC2]等协同调控。m6A甲基化修饰与人体的生长发育和多种疾病的发生发展密切相关。文章对m6A的分子机制、生物学功能和检测方法的最新进展进行综述,并对m6A检测的临床应用价值进行展望。

关键词: N6-甲基腺苷, RNA修饰, 肿瘤

Abstract:

N6-methyladenosine（m6A） is a dynamic reversible modification with the highest concentration of mRNA in mammalian cells. It is composed of methylases [methyltransferase like（METTL）3,METTL14,recombinant Wilms tumor 1 associated protein（WTAP）],demethylases [fat-mass and obesity-associated protein（FTO）,alk B homolog（ALKBH）5],methyl-binding proteins [YTH N6-methyladenosine RNA binding protein 1（YTHDF1）,YTH domain-containing（YTHDC）1,YTHDC2] and so on. The m6A methylation is related to the growth and development of human body and the occurrence and development of many diseases. In this review,the molecular mechanism,biological function,detection methods and the latest research progress of m6A are described,and the clinical application significance of m6A detection is prospected.

Key words: N6-methyladenosine, RNA modification, Tumor

中图分类号:

R446.7

孔玉洁, 王晨, 何炳. N6-甲基腺苷的生物学功能及检测技术研究进展[J]. 检验医学, 2022, 37(9): 872-876.

KONG Yujie, WANG Chen, HE Bing. Research progress of N6-methyladenosine in biological function and detection technology[J]. Laboratory Medicine, 2022, 37(9): 872-876.

图/表 2

参考文献 37

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肿瘤类型	m6A调控蛋白	表达趋势	下游基因	肿瘤发展
GBM^①[28]	METTL3	下调	ADAM19、EPHA3、KLF4、CDKN2A、BRCA2、TP53	抑制
	METTL14	下调	ADAM19	抑制
	ALKBH5	上调	FOXM1	促进
AML^②[29]	METTL3	上调	c-MYC、BCL2、PTEN	促进
	METTL14	上调	MYB、MYC	促进
	FTO	上调	ASB2、RARA	促进
肝细胞肝癌^[30]	METTL3	上调	SOCS2	促进
	METTL14	下调	DGCR8	抑制
	YTHDF2	上调	未知	促进
乳腺癌^[31]	METTL3	上调	HBXIP	促进
	ALKBH5	上调	NANOG、KLF4	促进
胰腺癌^[32]	ALKBH5	下调	KCNK15-AS1	抑制
	YTHDF2	下调	YAP	抑制
前列腺癌^[33]	YTHDF2	上调	miR-493-3p	促进
宫颈癌^[34]	FTO	上调	β-catenin	促进
子宫内膜癌^[35]	METTL3	下调	PHLPP2、mTORC2	抑制

肿瘤类型	m6A调控蛋白	表达趋势	下游基因	肿瘤发展
GBM^①[28]	METTL3	下调	ADAM19、EPHA3、KLF4、CDKN2A、BRCA2、TP53	抑制
	METTL14	下调	ADAM19	抑制
	ALKBH5	上调	FOXM1	促进
AML^②[29]	METTL3	上调	c-MYC、BCL2、PTEN	促进
	METTL14	上调	MYB、MYC	促进
	FTO	上调	ASB2、RARA	促进
肝细胞肝癌^[30]	METTL3	上调	SOCS2	促进
	METTL14	下调	DGCR8	抑制
	YTHDF2	上调	未知	促进
乳腺癌^[31]	METTL3	上调	HBXIP	促进
	ALKBH5	上调	NANOG、KLF4	促进
胰腺癌^[32]	ALKBH5	下调	KCNK15-AS1	抑制
	YTHDF2	下调	YAP	抑制
前列腺癌^[33]	YTHDF2	上调	miR-493-3p	促进
宫颈癌^[34]	FTO	上调	β-catenin	促进
子宫内膜癌^[35]	METTL3	下调	PHLPP2、mTORC2	抑制

检测方法	检测水平	优点	缺点
m6A-Seq	修饰相对丰度	高通量	对原始材料质量、数量要求高,受抗体影响大,分辨率低
MeRIP-Seq	全转录组	操作流程完善,灵敏度高,通量高	对研究目标质量、数量要求高,易受抗体影响,分辨率较低,mRNA需求量较大,免疫沉淀/抗体可能会有假阳性
2D-TLC	m6A水平	定量,灵敏度高,操作流程完善	有放射性,精确度不高
斑点杂交	m6A水平	成本低,没有放射性要求	易出现非特异性抗体结合,灵敏度低
LC-MS/MS	m6A水平	定量,灵敏度高,检测限低	对设备要求高,成本高,实验过程较为繁琐
SCARLET	特定部位甲基化	高度定量,通量低,准确度高	成本高,有放射性,过程繁琐,不能实现高通量,不能用于转录组分析
PA-m6A-Seq	单核苷酸分辨率	分辨率比MeRIP-Seq高,通量高	只适用于细胞,不能用于组织或临床样本,免疫沉淀/抗体会产生假阳性
mi-CLIP-Seq	单核苷酸分辨率	通量高	对特异性抗体具有高度依赖性
m6A-LAIC-Seq	m6A水平	通量高,可量化甲基化与未甲基化RNA的百分比	无法检测到特定的m6A位点
DA-m6A-seq	m6A水平	更高的灵敏度和测序深度	成本较高,实验过程较复杂

检测方法	检测水平	优点	缺点
m6A-Seq	修饰相对丰度	高通量	对原始材料质量、数量要求高,受抗体影响大,分辨率低
MeRIP-Seq	全转录组	操作流程完善,灵敏度高,通量高	对研究目标质量、数量要求高,易受抗体影响,分辨率较低,mRNA需求量较大,免疫沉淀/抗体可能会有假阳性
2D-TLC	m6A水平	定量,灵敏度高,操作流程完善	有放射性,精确度不高
斑点杂交	m6A水平	成本低,没有放射性要求	易出现非特异性抗体结合,灵敏度低
LC-MS/MS	m6A水平	定量,灵敏度高,检测限低	对设备要求高,成本高,实验过程较为繁琐
SCARLET	特定部位甲基化	高度定量,通量低,准确度高	成本高,有放射性,过程繁琐,不能实现高通量,不能用于转录组分析
PA-m6A-Seq	单核苷酸分辨率	分辨率比MeRIP-Seq高,通量高	只适用于细胞,不能用于组织或临床样本,免疫沉淀/抗体会产生假阳性
mi-CLIP-Seq	单核苷酸分辨率	通量高	对特异性抗体具有高度依赖性
m6A-LAIC-Seq	m6A水平	通量高,可量化甲基化与未甲基化RNA的百分比	无法检测到特定的m6A位点
DA-m6A-seq	m6A水平	更高的灵敏度和测序深度	成本较高,实验过程较复杂