Sensing of Lys 63-linked polyubiquitination by NEMO is a key event in NF-kappaB activation [corrected].
The transcription factor NF-kappaB is sequestered in the cytoplasm in a complex with IkappaB. Almost all NF-kappaB activation pathways converge on IkappaB kinase (IKK), which phosphorylates IkappaB resulting in Lys 48-linked polyubiquitination of IkappaB and its degradation. This allows migration of NF-kappaB to the nucleus where it regulates gene expression. IKK has two catalytic subunits, IKKalpha and IKKbeta, and a regulatory subunit, IKKgamma or NEMO. NEMO is essential for NF-kappaB activation, and NEMO dysfunction in humans is the cause of incontinentia pigmenti and hypohidrotic ectodermal dysplasia and immunodeficiency (HED-ID). The recruitment of IKK to occupied cytokine receptors, and its subsequent activation, are dependent on the attachment of Lys 63-linked polyubiquitin chains to signalling intermediates such as receptor-interacting protein (RIP). Here, we show that NEMO binds to Lys 63- but not Lys 48-linked polyubiquitin, and that single point mutations in NEMO that prevent binding to Lys 63-linked polyubiquitin also abrogates the binding of NEMO to RIP in tumour necrosis factor (TNF)-alpha-stimulated cells, the recruitment of IKK to TNF receptor (TNF-R) 1, and the activation of IKK and NF-kappaB. RIP is also destabilized in the absence of NEMO binding and undergoes proteasomal degradation in TNF-alpha-treated cells. These results provide a mechanism for NEMO's critical role in IKK activation, and a key to understanding the link between cytokine-receptor proximal signalling and IKK and NF-kappaB activation.
Laboratory of Immune Cell Biology, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892, USA.
Chuan-Jin
CJ
Wu
Dietrich B
DB
Conze
Tao
T
Li
Srinivasa M
SM
Srinivasula
Jonathan D
JD
Ashwell
D015374:Biosensing Techniques; D024202:Electrophoretic Mobility Shift Assay; D006367:HeLa Cells; D006801:Humans; D051550:I-kappa B Kinase / Q000235:genetics* / Q000378:metabolism; D047468:Immunoprecipitation; D008239:Lysine / Q000235:genetics / Q000378:metabolism*; D016328:NF-kappa B / Q000235:genetics / Q000378:metabolism*; D017354:Point Mutation; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism; D053422:Receptor-Interacting Protein Serine-Threonine Kinases; D047888:Receptors, Tumor Necrosis Factor, Type I / Q000378:metabolism; D012441:Saccharomyces cerevisiae; D015398:Signal Transduction; D047988:Tumor Necrosis Factor Receptor-Associated Peptides and Proteins / Q000378:metabolism; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology; D020798:Two-Hybrid System Techniques; D025801:Ubiquitin / Q000378:metabolism*
C491731:IKBKG protein, human; D016328:NF-kappa B; D047888:Receptors, Tumor Necrosis Factor, Type I; D047988:Tumor Necrosis Factor Receptor-Associated Peptides and Proteins; D014409:Tumor Necrosis Factor-alpha; D025801:Ubiquitin; D017346:Protein Serine-Threonine Kinases; C506249:RIPK1 protein, human; D053422:Receptor-Interacting Protein Serine-Threonine Kinases; D051550:I-kappa B Kinase; D008239:Lysine