Grape seed extract upregulates p21 (Cip1) through redox-mediated activation of ERK1/2 and posttranscriptional regulation leading to cell cycle arrest in colon carcinoma HT29 cells.
Abnormalities in cell cycle progression provide unlimited replicative potential to cancer cells, and therefore targeting of key cell cycle regulators could be a sound cancer chemopreventive strategy. Earlier, we found that grape seed extract (GSE) increases Cip/p21 protein level and inhibits growth and induces apoptosis in human colon carcinoma HT29 cells both in vitro and in vivo. However, the mechanism of GSE-induced p21 upregulation and its role in biological efficacy of GSE are not known, which were investigated here. GSE treatment of HT29 cells resulted in a strong dose- and time-dependent phosphorylation of extracellular signal regulated kinase 1/2 (ERK1/2), consistent with p21 induction. The inhibition of sustained ERK1/2 activation by GSE using pharmacological inhibitors abrogated GSE-induced p21 upregulation. Furthermore, pretreatment of cells with N-acetylcysteine inhibited GSE-induced ERK1/2 phosphorylation as well as p21 upregulation, suggesting the involvement of GSE-induced oxidative stress as an upstream event. Consistent with this, GSE also decreased intracellular level of reduced glutathione. Next, we determined whether GSE-induced signaling regulates p21 expression at transcriptional and/or translational levels. GSE was found to increase the stability of p21 message with resultant increase in p21 protein level, but it did not alter the protein stability to a great extent. Importantly, knock-down of p21 abrogated GSE-induced G(1) arrest suggesting that p21 induction by GSE is essential for its G(1) arrest effect. Together, our results for the first time identify a central role of p21 induction and associated mechanism in GSE-induced cell cycle arrest in HT29 cells.
Department of Pharmaceutical Sciences, School of Pharmacy, University of Colorado Denver, Aurora, USA.
Manjinder
M
Kaur
Alpna
A
Tyagi
Rana P
RP
Singh
Robert A
RA
Sclafani
Rajesh
R
Agarwal
Chapla
C
Agarwal
D001483:Base Sequence; D015153:Blotting, Western; D002453:Cell Cycle / Q000187:drug effects*; D003110:Colonic Neoplasms / Q000201:enzymology / Q000378:metabolism* / Q000473:pathology; D050759:Cyclin-Dependent Kinase Inhibitor p21 / Q000378:metabolism*; D017931:DNA Primers; D004789:Enzyme Activation; D056604:Grape Seed Extract / Q000494:pharmacology*; D019073:HT29 Cells; D006801:Humans; D019950:Mitogen-Activated Protein Kinase 1 / Q000378:metabolism*; D048052:Mitogen-Activated Protein Kinase 3 / Q000378:metabolism*; D010084:Oxidation-Reduction; D016133:Polymerase Chain Reaction; D012323:RNA Processing, Post-Transcriptional; D015854:Up-Regulation / Q000187:drug effects*
C493956:CDKN1A protein, human; D050759:Cyclin-Dependent Kinase Inhibitor p21; D017931:DNA Primers; D056604:Grape Seed Extract; C535150:MAPK1 protein, human; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3
Exp Cell Res. 1999 Feb 1;246(2):280-9
9925742
Am J Pathol. 1997 Jan;150(1):167-75
9006333
Antioxid Redox Signal. 2005 Jan-Feb;7(1-2):108-18
15650400
Cancer Res. 2000 Sep 1;60(17):4785-91
10987287
Cell Cycle. 2008 Jun 15;7(12):1828-35
18583941
Cell Cycle. 2004 Feb;3(2):138-40
14712075
J Biol Chem. 2009 Nov 27;284(48):33006-18
19805545
Int J Oncol. 2000 May;16(5):871-86
10762622
Cancer Res. 1998 Nov 1;58(21):4851-7
9809990
Mol Cell Biol. 2005 Oct;25(20):8938-47
16199872
Nature. 1993 Dec 16;366(6456):701-4
8259214
Cancer Cell. 2003 Dec;4(6):425-9
14706334
Mol Cell Biol. 1998 Mar;18(3):1400-7
9488455
Mutat Res. 2010 Apr-Jun;704(1-3):12-20
20096807
Pathol Biol (Paris). 2000 Apr;48(3):190-202
10858953
Apoptosis. 2007 Jul;12(7):1339-47
17431791
Blood. 1994 Dec 1;84(11):3781-4
7949134
Cancer Cell. 2005 Feb;7(2):113-4
15710322
J Biol Chem. 2003 Aug 29;278(35):32507-16
12748190
Genes Dev. 1999 Jun 15;13(12):1501-12
10385618
Cancer Chemother Pharmacol. 2006 Feb;57(3):317-27
16170570
Gen Physiol Biophys. 2008 Jun;27(2):71-84
18645221
Growth Factors. 2006 Mar;24(1):21-44
16393692
Crit Rev Oral Biol Med. 2002;13(6):453-64
12499239
Am J Physiol. 1997 Mar;272(3 Pt 1):L558-65
9124614
World J Gastroenterol. 2002 Jun;8(3):400-5
12046058
J Biol Chem. 2003 Jan 31;278(5):2937-46
12431987
EMBO J. 2000 Nov 1;19(21):5762-71
11060027
Hepatogastroenterology. 2005 Sep-Oct;52(65):1421-6
16201087
Oncogene. 2007 Jul 19;26(33):4806-16
17297446
Food Chem Toxicol. 2002 May;40(5):599-607
11955665
Clin Cancer Res. 2006 Oct 15;12(20 Pt 1):6194-202
17062697
Cancer Res. 1999 Oct 15;59(20):5341-8
10537318
J Biol Chem. 2002 Mar 29;277(13):11352-61
11756412
J Cell Biochem. 1999 Jun 1;73(3):291-302
10321829
Dev Cell. 2008 Feb;14(2):159-69
18267085
Toxicol Lett. 2010 May 19;195(1):23-30
20206247
Mol Carcinog. 1999 Sep;26(1):44-52
10487521
Biochem Soc Trans. 2009 Oct;37(Pt 5):937-53
19754430
Mol Cancer Ther. 2006 May;5(5):1265-74
16731759
Clin Cancer Res. 2000 Jun;6(6):2440-7
10873097
EMBO J. 1999 Mar 15;18(6):1571-83
10075928
Hum Pathol. 2003 Jun;34(6):580-8
12827612
Protoplasma. 2001;216(3-4):119-42
11732181
Eur J Biochem. 1997 May 1;245(3):730-7
9183012
Proc Natl Acad Sci U S A. 1997 Jan 21;94(2):448-52
9012803
Int J Biochem Cell Biol. 2007;39(7-8):1367-74
17412634
Carcinogenesis. 2006 Jul;27(7):1445-53
16474170