TY - JOUR
T1 - Live-cell imaging of microRNA expression with post-transcriptional feedback control
AU - Sano, Masayuki
AU - Morishita, Kana
AU - Oikawa, Satoshi
AU - Akimoto, Takayuki
AU - Sumaru, Kimio
AU - Kato, Yoshio
N1 - Funding Information:
This work was supported in part by JSPS KAKENHI (grants 19K06501 to M.S., 20K20620 to T.A., and 19H02578 to K.S.).
Publisher Copyright:
© 2021 The Author(s)
PY - 2021/12/3
Y1 - 2021/12/3
N2 - 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-miR-ON 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.
AB - 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-miR-ON 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.
KW - Csy4
KW - fluorescent protein
KW - live-cell imaging
KW - microRNA
KW - post-transcriptional feedback control
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U2 - 10.1016/j.omtn.2021.08.018
DO - 10.1016/j.omtn.2021.08.018
M3 - Article
AN - SCOPUS:85115760171
VL - 26
SP - 547
EP - 556
JO - Molecular Therapy - Nucleic Acids
JF - Molecular Therapy - Nucleic Acids
SN - 2162-2531
ER -