COP9 signalosome-specific phosphorylation targets p53 to degradation by the ubiquitin system.
In higher eukaryotic cells, the p53 protein is degraded by the ubiquitin-26S proteasome system mediated by Mdm2 or the human papilloma virus E6 protein. Here we show that COP9 signalosome (CSN)-specific phosphorylation targets human p53 to ubiquitin-26S proteasome-dependent degradation. As visualized by electron microscopy, p53 binds with high affinity to the native CSN complex. p53 interacts via its N-terminus with CSN subunit 5/Jab1 as shown by far-western and pull-down assays. The CSN-specific phosphorylation sites were mapped to the core domain of p53 including Thr155. A phosphorylated peptide, Deltap53(145-164), specifically inhibits CSN-mediated phosphorylation and p53 degradation. Curcumin, a CSN kinase inhibitor, blocks E6-dependent p53 degradation in reticulocyte lysates. Mutation of Thr155 to valine is sufficient to stabilize p53 against E6-dependent degradation in reticulocyte lysates and to reduce binding to Mdm2. The p53T155V mutant accumulates in both HeLa and HL 60 cells and exhibits a mutant (PAb 240+) conformation. It induces the cyclin-dependent inhibitor p21. In HeLa and MCF-7 cells, inhibition of CSN kinase by curcumin or Deltap53(145-164) results in accumulation of endogenous p53.
Bech-Otschir
D
D
Division of Molecular Biology, Department of Surgery and Institute of Biochemistry, Medical Faculty Charité, Humboldt University, Monbijoustrasse 2, 10117 Berlin, Germany.
Kraft
R
R
Huang
X
X
Henklein
P
P
Kapelari
B
B
Pollmann
C
C
Dubiel
W
W
D000595:Amino Acid Sequence; D001665:Binding Sites; D000075686:COP9 Signalosome Complex; D004247:DNA / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000378:metabolism; D018922:HL-60 Cells; D006367:HeLa Cells; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins; D008969:Molecular Sequence Data; D046912:Multiprotein Complexes; D016297:Mutagenesis, Site-Directed; D010447:Peptide Hydrolases / Q000378:metabolism*; D010766:Phosphorylation; D046988:Proteasome Endopeptidase Complex; D011506:Proteins / Q000378:metabolism*; D016283:Proto-Oncogene Proteins p21(ras) / Q000378:metabolism; D015398:Signal Transduction; D013912:Threonine / Q000235:genetics; D014157:Transcription Factors / Q000378:metabolism; D016159:Tumor Suppressor Protein p53 / Q000235:genetics / Q000378:metabolism*; D014452:Ubiquitins / Q000378:metabolism*; D014633:Valine / Q000235:genetics
D004268:DNA-Binding Proteins; D047908:Intracellular Signaling Peptides and Proteins; D046912:Multiprotein Complexes; D011506:Proteins; D014157:Transcription Factors; D016159:Tumor Suppressor Protein p53; D014452:Ubiquitins; D013912:Threonine; D004247:DNA; D010447:Peptide Hydrolases; C501488:COPS5 protein, human; D000075686:COP9 Signalosome Complex; D046988:Proteasome Endopeptidase Complex; C061553:ATP dependent 26S protease; C501009:HRAS protein, human; D016283:Proto-Oncogene Proteins p21(ras); D014633:Valine