NF-kappaB RelA phosphorylation regulates RelA acetylation.
The nuclear functions of NF-kappaB p50/RelA heterodimers are regulated in part by posttranslational modifications of its RelA subunit, including phosphorylation and acetylation. Acetylation at lysines 218, 221, and 310 differentially regulates RelA's DNA binding activity, assembly with IkappaBalpha, and transcriptional activity. However, it remains unclear whether the acetylation is regulated or simply due to stimulus-coupled nuclear translocation of NF-kappaB. Using anti-acetylated lysine 310 RelA antibodies, we detected p300-mediated acetylation of RelA in vitro and in vivo after stimulation of cells with tumor necrosis factor alpha (TNF-alpha). Coexpression of catalytically inactive mutants of the catalytic subunit of protein kinase A/mitogen- and stress-activated kinase 1 or IKK1/IKK2, which phosphorylate RelA on serine 276 or serine 536, respectively, sharply inhibited RelA acetylation on lysine 310. Furthermore, phosphorylation of RelA on serine 276 or serine 536 increased assembly of phospho-RelA with p300, which enhanced acetylation on lysine 310. Reconstitution of RelA-deficient murine embryonic fibroblasts with RelA S276A or RelA S536A decreased TNF-alpha-induced acetylation of lysine 310 and expression of the endogenous NF-kappaB-responsive E-selectin gene. These findings indicate that the acetylation of RelA at lysine 310 is importantly regulated by prior phosphorylation of serines 276 and 536. Such phosphorylated and acetylated forms of RelA display enhanced transcriptional activity.
Chen
Lin-Feng
LF
Gladstone Institute of Virology and Immunology, 1650 Owens St., San Francisco, CA 94158, USA. lchen@gladstone.ucsf.edu
Williams
Samuel A
SA
Mu
Yajun
Y
Nakano
Hiroyasu
H
Duerr
James M
JM
Buckbinder
Leonard
L
Greene
Warner C
WC
D000107:Acetylation; D000818:Animals; D002478:Cells, Cultured; D050881:E1A-Associated p300 Protein; D006801:Humans; D008239:Lysine / Q000378:metabolism; D051379:Mice; D009154:Mutation; D016328:NF-kappa B / Q000235:genetics / Q000378:metabolism*; D009687:Nuclear Proteins / Q000378:metabolism*; D010766:Phosphorylation; D012694:Serine / Q000378:metabolism; D015534:Trans-Activators / Q000378:metabolism*; D051996:Transcription Factor RelA; D014158:Transcription, Genetic
D016328:NF-kappa B; D009687:Nuclear Proteins; D015534:Trans-Activators; D051996:Transcription Factor RelA; D012694:Serine; D050881:E1A-Associated p300 Protein; C494361:Ep300 protein, mouse; D008239:Lysine