Methods to detect NF-κB acetylation and methylation

Methods in Molecular Biology
JinJing Chen, Lin-feng Chen

Abstract

Posttranslational modifications of NF-κB, including acetylation and methylation, have emerged as an important regulatory mechanism for determining the duration and strength of NF-κB nuclear activity as well as its transcriptional output. Within the seven NF-κB family proteins, the RelA subunit of NF-κB is the most studied for its regulation by lysine acetylation and methylation. Acetylation or methylation at different lysine residues modulates distinct functions of NF-κB, including DNA-binding and transcription activity, protein stability, and its interaction with NF-κB modulators. Here, we describe the experimental methods to monitor the in vitro and in vivo acetylated or methylated forms of NF-κB. These methods include radiolabeling the acetyl or methyl groups and immunoblotting with pan- or site-specific acetyl- or methyl-lysine antibodies. Radiolabeling is useful in the initial validation of the modifications. Immunoblotting with antibodies provides a rapid and powerful approach to detect and analyze the functions of these modifications in vitro and in vivo.

Citations

Aug 10, 2016·The Journal of General Virology·Ariana G Bravo Cruz, Joanna L Shisler
Apr 20, 2018·Biochemistry and Cell Biology = Biochimie Et Biologie Cellulaire·Emma Bondy-ChorneyMichael Downey

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