title: p53-Dependent p21-mediated growth arrest pre-empts and protects HCT116 cells from PUMA-mediated apoptosis induced by EGCG.
abstract: The tumor suppressor protein p53 plays a key role in regulation of negative cellular growth in response to EGCG. To further explore the role of p53 signaling and elucidate the molecular mechanism, we employed colon cancer HCT116 cell line and its derivatives in which a specific transcriptional target of p53 is knocked down by homologous recombination. Cells expressing p53 and p21 accumulate in G1 upon treatment with EGCG. In contrast, same cells lacking p21 traverse through the cell cycle and eventually undergo apoptosis as revealed by TUNEL staining. Treatment with EGCG leads to induction of p53, p21 and PUMA in p21 wild-type, and p53 and PUMA in p21(-/-) cells. Ablation of p53 by RNAi protects p21(-/-) cells, thus indicating a p53-dependent apoptosis by EGCG. Furthermore, analysis of cells lacking PUMA or Bax with or without p21 but with p53 reveals that all the cells expressing p53 and p21 survived after EGCG treatment. More interestingly, cells lacking both PUMA and p21 survived ECGC treatment whereas those lacking p21 and Bax did not. Taken together, our results present a novel concept wherein p21-dependent growth arrest pre-empts and protects cells from otherwise, in its absence, apoptosis which is mediated by activation of pro-apoptotic protein PUMA. Furthermore, we find that p53-dependent activation of PUMA in response to EGCG directly leads to apoptosis with out requiring Bax as is the case in response to agents that induce DNA damage. p21, thus can be used as a molecular switch for therapeutic intervention of colon cancer.
authors.affiliation: Department of Genetics, Case Western Reserve University, Cleveland, OH 44106, USA.
authors.forename: Vijay S
authors.initials: VS
authors.lastname: Thakur
authors.forename: A R M
authors.initials: AR
authors.lastname: Ruhul Amin
authors.forename: Rajib K
authors.initials: RK
authors.lastname: Paul
authors.forename: Kalpana
authors.initials: K
authors.lastname: Gupta
authors.forename: Kedar
authors.initials: K
authors.lastname: Hastak
authors.forename: Mukesh K
authors.initials: MK
authors.lastname: Agarwal
authors.forename: Mark W
authors.initials: MW
authors.lastname: Jackson
authors.forename: David N
authors.initials: DN
authors.lastname: Wald
authors.forename: Hasan
authors.initials: H
authors.lastname: Mukhtar
authors.forename: Munna L
authors.initials: ML
authors.lastname: Agarwal
mesh_terms: D000818:Animals; D016588:Anticarcinogenic Agents / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects*; D051017:Apoptosis Regulatory Proteins / Q000172:deficiency / Q000235:genetics* / Q000502:physiology; D002392:Catechin / Q000031:analogs & derivatives* / Q000494:pharmacology; D002453:Cell Cycle / Q000187:drug effects; D050759:Cyclin-Dependent Kinase Inhibitor p21 / Q000378:metabolism*; D004249:DNA Damage; D016158:Genes, p53 / Q000187:drug effects; D045325:HCT116 Cells / Q000473:pathology / Q000502:physiology*; D006801:Humans; D011518:Proto-Oncogene Proteins / Q000172:deficiency / Q000235:genetics* / Q000502:physiology; D014162:Transfection; D016159:Tumor Suppressor Protein p53 / Q000235:genetics* / Q000378:metabolism
chemical_list: D016588:Anticarcinogenic Agents; D051017:Apoptosis Regulatory Proteins; C438378:BBC3 protein, human; C493956:CDKN1A protein, human; D050759:Cyclin-Dependent Kinase Inhibitor p21; D011518:Proto-Oncogene Proteins; D016159:Tumor Suppressor Protein p53; D002392:Catechin; C045651:epigallocatechin gallate
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