Protein/DNA arrays identify nitric oxide-regulated cis-element and trans-factor activities some of which govern neuroblastoma cell viability.
Toxic nitric oxide (NO) levels can regulate gene expression. Using a novel protein/DNA array, we show that toxic NO levels regulate the binding of trans-factors to various cis-elements in neuroblastoma cells, including CRE and those recognized by the transcription factors AP1, AP2, Brn-3a, EGR, E2F1 and SP1. Functionality of some of the cis-elements was confirmed by electro mobility shift and reporter assays. Interestingly, CREB, AP-1, Brn-3a, EGR and E2F1 can control mammalian cell viability. NO induced the anti-apoptotic Bcl-2 protein and its mRNA prior to the onset of death of 30-60% of the cells. Promoter analysis of the bcl-2 gene confirmed the involvement of a CRE in NO-dependent bcl-2 transcription. Neuroblastoma cells over-expressing bcl-2 became much more resistant to NO-induced apoptosis; conversely, Bcl-2 knockdown cells were rendered markedly more sensitive to NO. Together these results suggest that Bcl-2 counteracts NO-induced apoptosis in a fraction of the cell population. Thus, NO stimulates the binding of many trans-factors to their cognate cis-elements, some of which can regulate cell viability through transcriptional activation of target genes. Our results emphasize that a DNA/protein array approach can reveal novel, global transcription factor activities stimulated by cell death-regulating molecules.
Dhakshinamoorthy
Saravanakumar
S
Cell Death and Human Diseases Group, Division of Genomics and Genetics, Institute of Molecular & Cell Biology, Proteos, 61 Biopolis Drive, Singapore 138673, Republic of Singapore.
Sridharan
Shiva Ranjani
SR
Li
Lei
L
Ng
Poh Yong
PY
Boxer
Linda M
LM
Porter
Alan G
AG
D017209:Apoptosis / Q000235:genetics*; D001665:Binding Sites; D045744:Cell Line, Tumor; D002470:Cell Survival; D000242:Cyclic AMP / Q000378:metabolism; D019254:Genes, bcl-2; D006801:Humans; D009447:Neuroblastoma; D009569:Nitric Oxide / Q000378:metabolism*; D020411:Oligonucleotide Array Sequence Analysis; D040081:Protein Array Analysis; D020218:Response Elements; D014157:Transcription Factors / Q000378:metabolism*; D015533:Transcriptional Activation
D014157:Transcription Factors; D009569:Nitric Oxide; D000242:Cyclic AMP
Cell Death Differ. 1999 Oct;6(10):969-75
10556974
Pharmacol Rev. 2000 Sep;52(3):375-414
10977868
FASEB J. 2000 Jan;14(1):166-73
10627291
Mol Med. 1999 Nov;5(11):721-30
10656874
Nat Cell Biol. 2000 Jun;2(6):339-45
10854324
J Biol Chem. 2000 Dec 22;275(51):40134-41
11013233
Trends Cell Biol. 2001 Feb;11(2):66-75
11166214
Nat Rev Mol Cell Biol. 2002 Mar;3(3):214-20
11994742
Oncogene. 2002 May 30;21(24):3898-908
12032828
Mol Cell Biol. 2002 Aug;22(15):5357-66
12101231
J Immunol. 2002 Sep 1;169(5):2619-26
12193733
Oncogene. 2002 Sep 5;21(39):6123-31
12203124
J Neurochem. 2002 Sep;82(5):1282-9
12358775
Curr Biol. 2002 Sep 17;12(18):R620-2
12372267
Mol Biotechnol. 2002 Oct;22(2):123-7
12405260
J Biol Chem. 2002 Dec 20;277(51):49896-902
12368293
Mol Pharmacol. 2003 Mar;63(3):565-73
12606763
Free Radic Biol Med. 2003 May 1;34(9):1136-45
12706494
Cell Death Differ. 2003 Apr;10(4):468-76
12719724
Oncogene. 2003 May 15;22(19):2857-68
12771937
Cell Death Differ. 2003 Aug;10(8):864-9
12867993
Curr Med Chem. 2003 Oct;10(20):2147-74
12871090
Cell Death Differ. 2003 Sep;10(9):956-68
12934070
Gene. 2003 Oct 2;315:33-41
14557062
J Biol Chem. 2003 Oct 24;278(43):42321-9
12907672
Circ Res. 2003 Nov 28;93(11):1034-46
14645134
J Biol Chem. 2003 Dec 26;278(52):52093-101
14551210
J Biol Chem. 2004 Feb 6;279(6):4058-65
14617628
J Biol Chem. 2004 May 7;279(19):20096-107
14985350
Cancer Res. 2004 May 1;64(9):3022-9
15126337
Proc Natl Acad Sci U S A. 2004 Jun 15;101(24):8841-2
15187230
J Biol Chem. 2004 Sep 10;279(37):38480-5
15247296
J Cell Sci. 2004 Sep 15;117(Pt 20):4727-37
15331636
Proc Natl Acad Sci U S A. 1993 Jun 1;90(11):5282-6
8506376
EMBO J. 1995 Apr 3;14(7):1508-19
7729426
FASEB J. 1995 Apr;9(7):552-8
7737465
Nature. 1996 May 16;381(6579):235-8
8622765
Nucleic Acids Res. 1996 Jun 15;24(12):2236-42
8710491
FEBS Lett. 1997 Feb 24;403(3):273-8
9091316
J Biol Chem. 1997 May 30;272(22):14244-50
9162057
Genes Dev. 1997 Aug 1;11(15):1938-48
9271117
J Clin Invest. 1997 Nov 1;100(9):2146-52
9410890
Neurochem Int. 1998 Jan;32(1):93-102
9460707
J Immunol. 1998 Mar 15;160(6):2590-6
9510156
J Exp Med. 1998 May 4;187(9):1361-5
9565628
EMBO J. 1998 Aug 3;17(15):4358-69
9687504
Nat Rev Mol Cell Biol. 2005 Feb;6(2):150-66
15688001
J Biol Chem. 2005 Mar 25;280(12):11851-8
15598651
Mol Immunol. 2005 Jul;42(11):1303-10
15950726
Nat Cell Biol. 2005 Jul;7(7):645-6
15990893
Cancer Lett. 2005 Aug 8;226(1):1-15
16004928
Cancer Res. 2005 Jul 15;65(14):6097-104
16024610
Oncogene. 2005 Oct 6;24(44):6689-98
16007171
Nucleic Acids Res. 2006;34(22):6640-52
17145718
Cancer Biol Ther. 2004 Dec;3(12):1208-11
15662116
Blood. 2000 Jan 1;95(1):189-97
10607702