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
Descripción
Sumario: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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