TNF-alpha induced p21(WAF1) but not Bax in colon cancer cells WiDr with mutated p53: important role of protein stabilization.
The p21(WAF1)induces cell cycle arrest at G(1)and its expression is regulated by the functional p53. TNF-alpha induced expression of p21(WAF1)at protein and mRNA levels in a dose-dependent fashion with an association with G(1)-arrest in human colon cancer cells WiDr that carry mutated p53 at codon 273 (His(273)). However, TNF-alpha did not affect the levels of the Bax protein, which also has p53-binding sites on its promoter and causes apoptosis. Further experiments suggested that cycloheximide (CHX), a protein synthesis inhibitor, increased the levels of p21(WAF1)mRNA and the induction of p21(WAF1)mRNA by TNF-alpha did not require new protein synthesis. Co-transfection of the p53 His(273)expression construct with a luciferase gene controlled by the p21(WAF1)promoter showed that the p53 His(273)was inactive, although TNF-alpha increased the transcriptional rate of p21(WAF1)in these cells. Further study found that TNF-alpha markedly stabilized the p21(WAF1)protein. These findings suggest that TNF-alpha induces expression of p21(WAF1)through a distinct pathway from Bax and that protein stabilization is an important mechanism in the expression of p21(WAF1)independent of p53.
Kobayashi
N
N
Division of Radiation Health, National Institute of Radiological Sciences, Chiba 263-8555, Japan.
Takada
Y
Y
Hachiya
M
M
Ando
K
K
Nakajima
N
N
Akashi
M
M
D017209:Apoptosis / Q000187:drug effects; D001665:Binding Sites; D015153:Blotting, Western; D002467:Cell Nucleus / Q000378:metabolism; D003110:Colonic Neoplasms / Q000378:metabolism*; D050759:Cyclin-Dependent Kinase Inhibitor p21; D016213:Cyclins / Q000378:metabolism*; D003513:Cycloheximide / Q000494:pharmacology; D004247:DNA / Q000378:metabolism; D018076:DNA, Complementary / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D005434:Flow Cytometry; D016193:G1 Phase / Q000187:drug effects; D016158:Genes, p53; D006639:Histidine / Q000737:chemistry; D006801:Humans; D008156:Luciferases / Q000378:metabolism; D009154:Mutation; D010957:Plasmids / Q000378:metabolism; D011401:Promoter Regions, Genetic; D011499:Protein Processing, Post-Translational; D011500:Protein Synthesis Inhibitors / Q000494:pharmacology; D011518:Proto-Oncogene Proteins / Q000378:metabolism*; D019253:Proto-Oncogene Proteins c-bcl-2; D012333:RNA, Messenger / Q000378:metabolism; D013997:Time Factors; D014158:Transcription, Genetic; D015533:Transcriptional Activation; D014162:Transfection; D014407:Tumor Cells, Cultured; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism* / Q000494:pharmacology; D015854:Up-Regulation; D051028:bcl-2-Associated X Protein
C494826:BAX protein, human; C493956:CDKN1A protein, human; D050759:Cyclin-Dependent Kinase Inhibitor p21; D016213:Cyclins; D018076:DNA, Complementary; D011500:Protein Synthesis Inhibitors; D011518:Proto-Oncogene Proteins; D019253:Proto-Oncogene Proteins c-bcl-2; D012333:RNA, Messenger; D014409:Tumor Necrosis Factor-alpha; D051028:bcl-2-Associated X Protein; D006639:Histidine; D004247:DNA; D003513:Cycloheximide; D008156:Luciferases