In vivo identification of inducible phosphoacceptors in the IKKgamma/NEMO subunit of human IkappaB kinase.
Transcription factor NF-kappaB plays a pivotal regulatory role in the genetic programs for cell cycle progression and inflammation. Nuclear translocation of NF-kappaB is controlled by an inducible protein kinase called IKK, which earmarks cytoplasmic inhibitors of NF-kappaB for proteolytic destruction. IKK contains two structurally related catalytic subunits termed IKKalpha and IKKbeta as well as a noncatalytic subunit called IKKgamma/NEMO. Mutations in the X-linked gene encoding IKKgamma can interfere with NF-kappaB signaling and lead to immunodeficiency disease. Although its precise mechanism of action remains unknown, IKKgamma is phosphorylated in concert with the induction of NF-kappaB by the viral oncoprotein Tax and the proinflammatory cytokine tumor necrosis factor alpha (TNF). We now demonstrate that TNF-induced phosphorylation of IKKgamma is blocked in cells deficient for IKKbeta but not IKKalpha. Phosphopeptide-mapping experiments with metabolically radiolabeled cells indicate that IKKbeta phosphorylates human IKKgamma at Ser-31, Ser-43, and Ser-376 following the enforced expression of either the Tax oncoprotein or the type 1 TNF receptor. Inducible phosphorylation of IKKgamma is attenuated following the deletion of its COOH-terminal zinc finger domain (amino acids 397-419), a frequent target for mutations that occur in IKKgamma-associated immunodeficiencies. As such, IKKbeta-mediated phosphorylation of IKKgamma at these specific serine targets may facilitate proper regulation of NF-kappaB signaling in the immune system.
Carter
Robert S
RS
Department of Microbiology and Immunology, Vanderbilt University School of Medicine, Nashville, Tennessee 37232-0295, USA.
Pennington
Kevin N
KN
Ungurait
Bradley J
BJ
Ballard
Dean W
DW
D002352:Carrier Proteins; D002460:Cell Line; D051550:I-kappa B Kinase; D020928:Mitogen-Activated Protein Kinases / Q000737:chemistry*; D010766:Phosphorylation; D017346:Protein Serine-Threonine Kinases / Q000737:chemistry* / Q000378:metabolism*
D002352:Carrier Proteins; C491731:IKBKG protein, human; D017346:Protein Serine-Threonine Kinases; D051550:I-kappa B Kinase; D020928:Mitogen-Activated Protein Kinases