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“NAD-capQ” detection and quantitation of NAD caps

RNA 5′ cap structures comprising the metabolic effector nicotinamide adenine dinucleotide (NAD) have been identified in diverse organisms. Here we report a simple, two-step procedure to detect and quantitate NAD-capped RNA, termed “NAD-capQ.” By use of NAD-capQ we quantitate NAD-capped RNA levels in...

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Detalles Bibliográficos
Autores principales: Grudzien-Nogalska, Ewa, Bird, Jeremy G., Nickels, Bryce E., Kiledjian, Megerditch
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Cold Spring Harbor Laboratory Press 2018
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6140466/
https://www.ncbi.nlm.nih.gov/pubmed/30045887
http://dx.doi.org/10.1261/rna.067686.118
Descripción
Sumario:RNA 5′ cap structures comprising the metabolic effector nicotinamide adenine dinucleotide (NAD) have been identified in diverse organisms. Here we report a simple, two-step procedure to detect and quantitate NAD-capped RNA, termed “NAD-capQ.” By use of NAD-capQ we quantitate NAD-capped RNA levels in Escherichia coli, Saccharomyces cerevisiae, and human cells, and we measure increases in NAD-capped RNA levels in cells from all three organisms harboring disruptions in their respective “deNADding” enzymes. We further show that NAD-capped RNA levels in human cells respond to changes in cellular NAD concentrations, indicating that NAD capping provides a mechanism for human cells to directly sense and respond to alterations in NAD metabolism. Our findings establish NAD-capQ as a versatile, rapid, and accessible methodology to detect and quantitate 5′-NAD caps on endogenous RNA in any organism.