Tracking Distinct RNA Populations Using Efficient and Reversible Covalent Chemistry

被引：143

作者：

Duffy, Erin E. ^{[1
,2
]}

Rutenberg-Schoenberg, Michael ^{[1
,2
]}

Stark, Catherine D. ^{[1
,2
]}

Kitchen, Robert R. ^{[1
]}

Gerstein, Mark B. ^{[1
]}

Simon, Matthew D. ^{[1
,2
]}

机构：

[1] Yale Univ, Dept Biochem & Mol Biophys, New Haven, CT 06511 USA

[2] Yale Univ, Inst Chem Biol, West Haven, CT 06516 USA

来源：

MOLECULAR CELL | 2015年 / 59卷 / 05期

关键词：

IN-VIVO; IDENTIFICATION; TURNOVER; 4-THIOURIDINE; DEGRADATION; TRANSCRIPTS; DYNAMICS; RATES;

D O I：

10.1016/j.molcel.2015.07.023

中图分类号：

Q5 [生物化学]; Q7 [分子生物学];

学科分类号：

071010 ; 081704 ;

摘要：

We describe a chemical method to label and purify 4-thiouridine (s(4)U)-containing RNA. We demonstrate that methanethiosulfonate (MTS) reagents form disulfide bonds with s(4)U more efficiently than the commonly used HPDP-biotin, leading to higher yields and less biased enrichment. This increase in efficiency allowed us to use s(4)U labeling to study global microRNA (miRNA) turnover in proliferating cultured human cells without perturbing global miRNA levels or the miRNA processing machinery. This improved chemistry will enhance methods that depend on tracking different populations of RNA, such as 4-thiouridine tagging to study tissue-specific transcription and dynamic transcriptome analysis (DTA) to study RNA turnover.

引用

页码：858 / 866

页数：9

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