1. Single-Cell Imaging of m 6 A Modified RNA Using m 6 A-Specific In Situ Hybridization Mediated Proximity Ligation Assay (m 6 AISH-PLA).
- Author
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Ren X, Deng R, Zhang K, Sun Y, Li Y, and Li J
- Subjects
- Adenosine chemistry, Cell Line, HSP70 Heat-Shock Proteins genetics, HSP70 Heat-Shock Proteins metabolism, Humans, RNA chemistry, RNA, Messenger metabolism, Single-Cell Analysis, Adenosine analogs & derivatives, In Situ Hybridization methods, RNA metabolism
- Abstract
N
6 -methyladenosine (m6 A) modification-the most prevalent mammalian RNA internal modification-plays key regulatory roles in mRNA metabolism. Current approaches for m6 A modified RNA analysis limit at bulk-population level, resulting in a loss of spatiotemporal and cell-to-cell variability information. Here we proposed a m6 A-specific in situ hybridization mediated proximity ligation assay (m6 AISH-PLA) for cellular imaging of m6 A RNA, allowing to identify m6 A modification at specific location in RNAs and image m6 A RNA with single-cell and single-molecule resolution. Using m6 AISH-PLA, we investigated the m6 A level and subcellular location of HSP70 RNA103-m6 A in response to heat shock stress, and found an increased m6 A modified ratio and an increased distribution ratio in cytoplasm under heat shock. m6 AISH-PLA can serve in the study of m6 A RNA in single cells for deciphering epitranscriptomic mechanisms and assisting clinical diagnosis., (© 2021 Wiley-VCH GmbH.)- Published
- 2021
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