Live-cell imaging of microRNA expression with post-transcriptional feedback control

被引:7
作者
Sano, Masayuki [1 ]
Morishita, Kana [1 ]
Oikawa, Satoshi [2 ]
Akimoto, Takayuki [2 ]
Sumaru, Kimio [1 ]
Kato, Yoshio [3 ]
机构
[1] Natl Inst Adv Ind Sci & Technol, Cellular & Mol Biotechnol Res Inst, Cent 5,1-1-1 Higashi, Tsukuba, Ibaraki 3058565, Japan
[2] Waseda Univ, Fac Sport Sci, 2-579-15 Mikajima, Tokorozawa, Saitama 3591192, Japan
[3] Natl Inst Adv Ind Sci & Technol, Biomed Res Inst, Cent 6,1-1-1 Higashi, Tsukuba, Ibaraki 3058566, Japan
来源
MOLECULAR THERAPY-NUCLEIC ACIDS | 2021年 / 26卷
关键词
TRANSGENE EXPRESSION; ENDOGENOUS MICRORNA; IN-VITRO; RNA; SYSTEM; VECTOR; DIFFERENTIATION; CRISPR/CAS; MECHANISM; MIRNAS;
D O I
10.1016/j.omtn.2021.08.018
中图分类号
R-3 [医学研究方法]; R3 [基础医学];
学科分类号
1001 ;
摘要
MicroRNAs (miRNAs) are small noncoding RNAs that regulate complex gene expression networks in eukaryotic cells. Because of their unique expression patterns, miRNAs are potential molecular markers for specific cell states. Although a system capable of imaging miRNA in living cells is needed to visually detect miRNA expression, very few fluorescence signal-on sensors that respond to expression of target miRNA (miR-ON sensors) are available. Here we report an miR-ON sensor containing a bidirectional promoter-driven Csy4 endoribonuclease and green fluorescent protein, ZsGreen1, for live-cell imaging of miRNAs with post-transcriptional feedback control. Csy4-assisted miR-ON (Csy4-miR-ON) sensors generate negligible background but respond sensitively to target miRNAs, allowing high-contrast fluorescence detection of miRNAs in various human cells. We show that Csy4-miRON sensors enabled imaging of various miRNAs, including miR-21, miR-302a, and miR-133, in vitro as well as in vivo. This robust tool can be used to evaluate miRNA expression in diverse biological and medical applications.
引用
收藏
页码:547 / 556
页数:10
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