title: A single mutant, A276S of p53, turns the switch to apoptosis.
abstract: The tumor suppressor protein p53 induces apoptosis, cell cycle arrest, and DNA repair along with other functions in a transcription-dependent manner [Vousden, K. H. Cell 2000, 103(5), 691-694]. The selection of these functions depends on sequence-specific recognition of p53 to a target decameric sequence of gene promoters [Kitayner, M.; et al. Mol. Cell 2006, 22(6), 741-753]. Amino acid residues in p53 that directly bind to DNA were analyzed, and the replacement of A276 in p53 with selected amino acids elucidated its importance in promoter transcription. For most apoptotic and cell cycle gene promoters, position 9 of the target decameric sequence is a cytosine, while for DNA repair gene promoters, thymine is found instead. Therefore, selective binding to the cytosine at the ninth position may transcribe apoptotic gene promoters and thus can induce apoptosis and cell cycle arrest. Molecular modeling with PyMOL indicated that substitution of a hydrophilic residue, A276S, would prefer binding to cytosine at the ninth position of the target decameric sequence, whereas substitution of a hydrophobic residue (A276F) would fail to do so. Correspondingly, A276S demonstrated higher transcription of PUMA, PERP, and p21(WAF1/CIP1)gene promoters containing a cytosine at the ninth position and lower transcription of GADD45 gene promoter containing a thymine at the ninth position compared to wild-type p53. Cell cycle analysis showed that A276S maintained similar G1/G0 phase arrest as wild-type p53. Additionally, A276S induced higher apoptosis than wild-type p53 as measured by DNA segmentation and 7-AAD assay. Since the status of endogenous p53 can influence the activity of the exogenous p53, we examined the activity of A276S in HeLa cells (wild-type endogenous p53) in addition to T47D cells (mutated and mislocalized endogenous p53). The same apoptotic trend in both cell lines suggested A276S can induce cell death regardless of endogenous p53 status. Cell proliferation assay depicted that A276S efficiently reduced the viability of T47D cells more than wild-type p53 over time. We conclude that the predicted preferred binding of A276S to cytosine at the ninth position better transactivates a number of apoptotic gene promoters. Higher induction apoptosis than wild-type p53 makes A276S an attractive candidate for therapy to eradicate cancer.
authors: Department of Pharmaceutics and Pharmaceutical Chemistry, University of Utah, Salt Lake City, Utah 84112, USA.
authors: Shams
authors: S
authors: Reaz
authors: Mohanad
authors: M
authors: Mossalam
authors: Abood
authors: A
authors: Okal
authors: Carol S
authors: CS
authors: Lim
mesh_terms: D017209:Apoptosis; D002453:Cell Cycle; D049109:Cell Proliferation; D002470:Cell Survival; D050759:Cyclin-Dependent Kinase Inhibitor p21 / Q000378:metabolism; D004247:DNA / Q000737:chemistry; D015316:Genetic Therapy / Q000379:methods; D006367:HeLa Cells; D006801:Humans; D009154:Mutation; D009336:Necrosis; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D014158:Transcription, Genetic; D015533:Transcriptional Activation; D016159:Tumor Suppressor Protein p53 / Q000235:genetics* / Q000502:physiology
chemical_list: D050759:Cyclin-Dependent Kinase Inhibitor p21; C495901:TP53 protein, human; D016159:Tumor Suppressor Protein p53; D004247:DNA
references: Mol Cell Biol. 1992 Jun;12(6):2866-71
references: 1588974
references: Mol Pharm. 2012 May 7;9(5):1449-58
references: 22380534
references: Science. 1994 Nov 25;266(5189):1376-80
references: 7973727
references: Blood. 2005 Feb 1;105(3):1187-94
references: 15466931
references: Biochemistry. 1998 Jun 23;37(25):8965-72
references: 9636038
references: Science. 1994 Jul 15;265(5170):346-55
references: 8023157
references: Curr Med Chem. 2009;16(33):4419-44
references: 19835564
references: Cell. 2000 Nov 22;103(5):691-4
references: 11114324
references: Brief Funct Genomics. 2013 Jan;12(1):46-57
references: 23242178
references: J Biol Chem. 2011 Aug 5;286(31):27751-60
references: 21659527
references: Mol Cell. 2001 Mar;7(3):673-82
references: 11463391
references: J Pept Sci. 1995 Sep-Oct;1(5):319-29
references: 9223011
references: Protein Eng Des Sel. 2009 Jul;22(7):421-30
references: 19515728
references: J Biol Chem. 2003 Jul 11;278(28):25395-400
references: 12734185
references: Mol Cell. 2006 Dec 28;24(6):827-39
references: 17189186
references: Cell. 1992 Nov 13;71(4):587-97
references: 1423616
references: Genes Dev. 2000 Mar 15;14(6):704-18
references: 10733530
references: Mol Cell. 2006 Jun 23;22(6):741-753
references: 16793544
references: DNA Repair (Amst). 2002 Sep 4;1(9):755-62
references: 12509279
references: J Biol Chem. 2009 Nov 6;284(45):30974-80
references: 19700401
references: J Biol Chem. 2004 Jan 9;279(2):1291-6
references: 14534297
references: Trends Cell Biol. 2012 Feb;22(2):97-106
references: 22154076
references: Mol Cancer Res. 2003 Dec;1(14):1048-57
references: 14707288
references: Pharm Res. 2012 Dec;29(12):3455-63
references: 22869106
references: J Histochem Cytochem. 2001 Jul;49(7):821-32
references: 11410607
references: Genes Dev. 1998 Jul 15;12(14):2102-7
references: 9679054
references: J Histochem Cytochem. 1999 Sep;47(9):1101-10
references: 10449530
references: Mol Pharm. 2012 Jan 1;9(1):187-95
references: 22136227
references: Cell. 1993 Nov 19;75(4):817-25
references: 8242752
references: Oncol Lett. 2012 May;3(5):978-982
references: 22783376
references: Oncogene. 2002 Nov 7;21(51):7901-11
references: 12420228
references: Cytometry. 1983 Mar;3(5):323-7
references: 6188586
references: J Biol Chem. 2008 Jan 4;283(1):388-397
references: 17959603
references: Nat Genet. 1992 Apr;1(1):45-9
references: 1301998
references: Science. 2002 Apr 19;296(5567):530-4
references: 11964479