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PMID: 7796813 Published · ppublish English Journal Article Research Support, Non-U.S. Gov't

Phosphorylation of human I kappa B-alpha on serines 32 and 36 controls I kappa B-alpha proteolysis and NF-kappa B activation in response to diverse stimuli.

The EMBO journal ·Vol. 14 ·No. 12 ·1995-06-15 ·Pages 2876-83

Traenckner EB, Pahl HL, Henkel T, Schmidt KN, Wilk S, Baeuerle PA

Abstract

Post-translational activation of the higher eukaryotic transcription factor NF-kappa B requires both phosphorylation and proteolytic degradation of the inhibitory subunit I kappa B-alpha. Inhibition of proteasome activity can stabilize an inducibly phosphorylated form of I kappa B-alpha in intact cells, suggesting that phosphorylation targets the protein for degradation. In this study, we have identified serines 32 and 36 in human I kappa B-alpha as essential for the control of I kappa B-alpha stability and the activation of NF-kappa B in HeLa cells. A point mutant substituting serines 32 and 36 by alanine residues was no longer phosphorylated in response to okadaic acid (OA) stimulation. This and various other Ser32 and Ser36 mutants behaved as potent dominant negative I kappa B proteins attenuating kappa B-dependent transactivation in response to OA, phorbol 12-myristate 13-acetate (PMA) and tumor necrosis factor-alpha (TNF). While both endogenous and transiently expressed wild-type I kappa B-alpha were proteolytically degraded in response to PMA and TNF stimulation of cells, the S32/36A mutant of I kappa B-alpha remained largely intact under these conditions. Our data suggest that such diverse stimuli as OA, TNF and PMA use the same kinase system to phosphorylate and thereby destabilize I kappa B-alpha, leading to NF-kappa B activation.

MeSH Terms
Base Sequence Cysteine Endopeptidases/metabolism DNA-Binding Proteins/chemistry,metabolism Enzyme Inhibitors/pharmacology Ethers, Cyclic/pharmacology HeLa Cells Humans I-kappa B Proteins Molecular Sequence Data Multienzyme Complexes/metabolism NF-KappaB Inhibitor alpha NF-kappa B/genetics,metabolism Okadaic Acid Oligopeptides/pharmacology Phosphoprotein Phosphatases/antagonists & inhibitors Phosphorylation/drug effects Point Mutation/physiology Proteasome Endopeptidase Complex Recombinant Fusion Proteins/biosynthesis,metabolism Serine/metabolism Tetradecanoylphorbol Acetate/pharmacology Transcriptional Activation/drug effects,physiology Tumor Necrosis Factor-alpha/pharmacology
Chemicals
DNA-Binding Proteins Enzyme Inhibitors Ethers, Cyclic I-kappa B Proteins Multienzyme Complexes NF-kappa B NFKBIA protein, human Oligopeptides Recombinant Fusion Proteins Tumor Necrosis Factor-alpha benzyloxycarbonyl-isoleucyl-glutamyl(O-tert-butyl)-alanyl-leucinal NF-KappaB Inhibitor alpha Okadaic Acid Serine Phosphoprotein Phosphatases Cysteine Endopeptidases Proteasome Endopeptidase Complex Tetradecanoylphorbol Acetate
Authors & Affiliations
6 authors, click to expand affiliations / ORCID
Traenckner E B
Institute of Biochemistry, Albert-Ludwigs-University, Freiburg, Germany.
Pahl H L
Henkel T
Schmidt K N
Wilk S
Baeuerle P A
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Article Info
Journal
The EMBO journal
Abbr.
EMBO J
ISSN
0261-4189
Published
1995-06-15
Pages
2876-83
Language
English
Region
England
NLM ID
8208664
PMCID
PMC398406
Subset
IM
External Links
PubMed source PMC Full Text View journal details
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