Protocol to analyze and quantify protein-methylated RNA interactions in mammalian cells with a combination of RNA immunoprecipitation and nucleoside mass spectrometry

Cellular RNAs are modified by both physiological factors and exogenous agents, such as methyl methanesulfonate (MMS). However, techniques for analyzing how proteins may interact with these modified RNAs are limited. Here, we provide a protocol combining RNA immunoprecipitation (RIP) with mass spectr...

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Detalles Bibliográficos
Autores principales: Tsao, Ning, Soll, Jennifer M., Mosammaparast, Nima
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Elsevier 2022
Materias:
Protocol
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8980960/
https://www.ncbi.nlm.nih.gov/pubmed/35391937
http://dx.doi.org/10.1016/j.xpro.2022.101268
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author Tsao, Ning
Soll, Jennifer M.
Mosammaparast, Nima
author_facet Tsao, Ning
Soll, Jennifer M.
Mosammaparast, Nima
author_sort Tsao, Ning
collection PubMed
description Cellular RNAs are modified by both physiological factors and exogenous agents, such as methyl methanesulfonate (MMS). However, techniques for analyzing how proteins may interact with these modified RNAs are limited. Here, we provide a protocol combining RNA immunoprecipitation (RIP) with mass spectrometry (MS) to analyze the methylation state of the RNAs bound by Flag-tagged proteins in mammalian cells. The approach is highly quantitative and can simultaneously detect several methylated nucleosides in a single experiment. For complete details on the use and execution of this protocol, please refer to Tsao et al. (2021).
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spelling pubmed-89809602022-04-06 Protocol to analyze and quantify protein-methylated RNA interactions in mammalian cells with a combination of RNA immunoprecipitation and nucleoside mass spectrometry Tsao, Ning Soll, Jennifer M. Mosammaparast, Nima STAR Protoc Protocol Cellular RNAs are modified by both physiological factors and exogenous agents, such as methyl methanesulfonate (MMS). However, techniques for analyzing how proteins may interact with these modified RNAs are limited. Here, we provide a protocol combining RNA immunoprecipitation (RIP) with mass spectrometry (MS) to analyze the methylation state of the RNAs bound by Flag-tagged proteins in mammalian cells. The approach is highly quantitative and can simultaneously detect several methylated nucleosides in a single experiment. For complete details on the use and execution of this protocol, please refer to Tsao et al. (2021). Elsevier 2022-04-01 /pmc/articles/PMC8980960/ /pubmed/35391937 http://dx.doi.org/10.1016/j.xpro.2022.101268 Text en © 2022 The Author(s) https://creativecommons.org/licenses/by-nc-nd/4.0/This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/).
spellingShingle Protocol
Tsao, Ning
Soll, Jennifer M.
Mosammaparast, Nima
Protocol to analyze and quantify protein-methylated RNA interactions in mammalian cells with a combination of RNA immunoprecipitation and nucleoside mass spectrometry
title Protocol to analyze and quantify protein-methylated RNA interactions in mammalian cells with a combination of RNA immunoprecipitation and nucleoside mass spectrometry
title_full Protocol to analyze and quantify protein-methylated RNA interactions in mammalian cells with a combination of RNA immunoprecipitation and nucleoside mass spectrometry
title_fullStr Protocol to analyze and quantify protein-methylated RNA interactions in mammalian cells with a combination of RNA immunoprecipitation and nucleoside mass spectrometry
title_full_unstemmed Protocol to analyze and quantify protein-methylated RNA interactions in mammalian cells with a combination of RNA immunoprecipitation and nucleoside mass spectrometry
title_short Protocol to analyze and quantify protein-methylated RNA interactions in mammalian cells with a combination of RNA immunoprecipitation and nucleoside mass spectrometry
title_sort protocol to analyze and quantify protein-methylated rna interactions in mammalian cells with a combination of rna immunoprecipitation and nucleoside mass spectrometry
topic Protocol
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8980960/
https://www.ncbi.nlm.nih.gov/pubmed/35391937
http://dx.doi.org/10.1016/j.xpro.2022.101268
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