Facile detection of RNA phospho-methylation in cells and tissues.

Facile detection of RNA phospho-methylation in cells and tissues.
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DOI:
10.1016/bs.mie.2021.06.002
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发表时间:
2021
影响因子:
--
通讯作者:
Xhemalçe B
Xhemalçe B
中科院分区:
生物学4区
文献类型:
--
作者:
Devanathan SK;Debnath TK;Xhemalçe B

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来自各种细胞和组织的 RNA 以近 200 种不同的化学方式进行修饰。这些修饰可以沉积在 5' 或 3' 末端,也可以沉积在核碱基或糖主链内部。 5' 端修饰对于保护 RNA 免受不合时宜的降解/加工、调节其细胞功能或区分内源性 RNA 和致病性 RNA 至关重要。 5'-末端磷酸甲基化是一种显着的 RNA 修饰,它通过人 BCDIN3/MePCE 酶促沉积在新生三磷酸化 RNA 的 γ-磷酸上,或通过人 BCDIN3D 酶促沉积在加工的单磷酸化 RNA 的 α-磷酸上。这些 5'-磷酸甲基转移酶是从粟酒裂殖酵母到人类中保守的 O-甲基转移酶 BIN3 家族的一部分,发挥重要的细胞和生物学作用,其中许多作用有待进一步阐明。在这里,我们快速回顾了检测 5' 端磷酸甲基修饰的历史方法,并更具体地关注可用于检测和定量来自细胞或组织的低至 10-100 ng 总 RNA 的 α-单磷酸甲基化的方法。该方法对于破译 BCDIN3D 及其同源物在物种间的作用非常重要,并且可以作为开发检测 5' 末端修饰的新方法的起点。
RNAs from various cells and tissues are modified in nearly 200 chemically distinct ways. These modifications can be deposited either on the 5′ or 3′ ends, or internally on the nucleobases or sugar backbone. 5′-end modifications are crucial for protecting RNAs from untimely degradation/processing, regulating their cellular functions, or discriminating endogenous RNAs from pathogenic RNAs. 5′-end phospho-methylation is a remarkable RNA modification that is enzymatically deposited either on the γ-phosphate of nascent triphosphorylated RNAs by human BCDIN3/MePCE, or on the α-phosphate of processed monophosphorylated RNAs by human BCDIN3D. These 5′-phosphomethyltransferases are part of the BIN3 family of O-methyltransferases conserved from S. pombe to humans and play important cellular and biological roles, many of which await further elucidation. Here, we quickly recapitulate historical methods for the detection of 5′-end phospho-methyl modifications, and focus more specifically on a method that can be used to detect and quantify α-monophosphate methylation from as low as 10–100 ng of total RNA from cells or tissues. This method is important for deciphering the roles of BCDIN3D and its homologs across species, as well as serves as starting point for the development of new methods for detection of 5′-end modifications.
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