Transcriptional Activation of the Human Inducible Nitric-oxide Synthase Promoter by Krüppel-like Factor 6

Abstract

Nitric oxide is a ubiquitous free radical that plays a key role in a broad spectrum of signaling pathways in physiological and pathophysiological processes. We have explored the transcriptional regulation of inducible nitric-oxide synthase (iNOS) by Krüppel-like factor 6 (KLF6), an Sp1-like zinc finger transcription factor. Study of serial deletion constructs of the iNOS promoter revealed that the proximal 0.63-kb region can support a 3–6-fold reporter activity similar to that of the full-length 16-kb promoter. Within the 0.63-kb region, we identified two CACCC sites (−164 to −168 and −261 to −265) that bound KLF6 in both electrophoretic mobility shift and chromatin immunoprecipitation assays. Mutation of both these sites abrogated the KLF6-induced enhancement of the 0.63-kb iNOS promoter activity. The binding of KLF6 to the iNOS promoter was significantly increased in Jurkat cells, primary T lymphocytes, and COS-7 cells subjected to NaCN-induced hypoxia, heat shock, serum starvation, and phorbol 12-myristate 13-acetate/A23187 ionophore stimulation. Furthermore, in KLF6-transfected and NaCN-treated COS-7 cells, there was a 3–4-fold increase in the expression of the endogenous iNOS mRNA and protein that correlated with increased production of nitric oxide. These findings indicate that KLF6 is a potential transactivator of the human iNOS promoter in diverse pathophysiological conditions. Nitric oxide is a ubiquitous free radical that plays a key role in a broad spectrum of signaling pathways in physiological and pathophysiological processes. We have explored the transcriptional regulation of inducible nitric-oxide synthase (iNOS) by Krüppel-like factor 6 (KLF6), an Sp1-like zinc finger transcription factor. Study of serial deletion constructs of the iNOS promoter revealed that the proximal 0.63-kb region can support a 3–6-fold reporter activity similar to that of the full-length 16-kb promoter. Within the 0.63-kb region, we identified two CACCC sites (−164 to −168 and −261 to −265) that bound KLF6 in both electrophoretic mobility shift and chromatin immunoprecipitation assays. Mutation of both these sites abrogated the KLF6-induced enhancement of the 0.63-kb iNOS promoter activity. The binding of KLF6 to the iNOS promoter was significantly increased in Jurkat cells, primary T lymphocytes, and COS-7 cells subjected to NaCN-induced hypoxia, heat shock, serum starvation, and phorbol 12-myristate 13-acetate/A23187 ionophore stimulation. Furthermore, in KLF6-transfected and NaCN-treated COS-7 cells, there was a 3–4-fold increase in the expression of the endogenous iNOS mRNA and protein that correlated with increased production of nitric oxide. These findings indicate that KLF6 is a potential transactivator of the human iNOS promoter in diverse pathophysiological conditions. inducible nitric-oxide synthase Krüppel-like factor transforming growth factor electrophoretic mobility shift assay chromatin immunoprecipitation phorbol 12-myristate 13-acetate glyceraldehyde-3-phosphate dehydrogenase 2-[bis(2-hydroxyethyl)amino]-2-(hydroxymethyl)propane-1,3-diol signal transducers and activators of transcription Nitric-oxide synthases are key proteins that produce NO and thereby regulate many important biological processes. NO is generated during the oxidation of l-arginine tol-citrulline by at least three different isoforms of nitric-oxide synthase. Endothelial and neuronal nitric-oxide synthases are constitutively expressed, and their activity is Ca2+- and calmodulin-dependent, whereas the third isoform is transcriptionally inducible (iNOS),1 and its activity is independent of Ca2+ and calmodulin and can produce very high levels of nitric oxide over a sustained period of time (1Taylor B.S. Geller D.A. Shock. 2000; 13: 413-424Crossref PubMed Scopus (151) Google Scholar, 2Alderton W.K. Cooper C.E. Knowles R.G. Biochem. J. 2001; 357: 593-615Crossref PubMed Scopus (3276) Google Scholar). It has been shown that iNOS is transcriptionally up-regulated in pathophysiologic conditions such as hypoxia, ischemia-reperfusion injury, and trauma and by reactive oxygen species (3Pfeilschifter J. Eberhardt W. Beck K.F. Pfluegers Arch. 2001; 442: 479-486Crossref PubMed Scopus (113) Google Scholar, 4Ganster R.W. Taylor B.S. Shao L. Geller D.A. Proc. Natl. Acad. Sci. U. S. A. 2001; 98: 8638-8643Crossref PubMed Scopus (281) Google Scholar). NO is a key central molecule in cellular biochemical processes, as it is freely diffusible and traverses cell membranes to reach different targets, alters signaling networks by redox-sensitive modifications, and transcriptionally regulates multiple gene families (5Bouchie J.L. Hansen H. Feener E.P. Arterioscler. Thromb. Vasc. Biol. 1998; 18: 1771-1779Crossref PubMed Scopus (105) Google Scholar, 6Eberhardt W. Huwiler A. Beck K.F. Walpen S. Pfeilschifter J. J. Immunol. 2000; 165: 5788-5797Crossref PubMed Scopus (198) Google Scholar, 7Eberhardt W. Beeg T. Beck K.F. Walpen S. Gauer S. Bohles H. Pfeilschifter J. Kidney Int. 2000; 57: 59-69Abstract Full Text Full Text PDF PubMed Scopus (131) Google Scholar, 8Eberhardt W. Beck K.F. Pfeilschifter J. Kidney Int. 2002; 61: 20-30Abstract Full Text Full Text PDF PubMed Scopus (34) Google Scholar, 9Grisham M.B. Jourd'heuil D. Wink D.A. Aliment. Pharmacol. Ther. 2000; 14 Suppl. 1: 3-9Crossref PubMed Google Scholar, 10Efron D.T. Most D. Barbul A. Curr. Opin. Clin. Nutr. Metab. Care. 2000; 3: 197-204Crossref PubMed Scopus (73) Google Scholar). NO production following iNOS up-regulation is associated with increased wound healing and repair in tissue injury (11Efron D.T. Thornton F.J. Steulten C. Tantry U.S. Witte M.B. Kiyama T. Barbul A. J. Gastrointest. Surg. 1999; 3: 592-601Crossref PubMed Scopus (42) Google Scholar, 12Yamasaki K. Edington H.D. McClosky C. Tzeng E. Lizonova A. Kovesdi I. Steed D.L. Billiar T.R. J. Clin. Invest. 1998; 101: 967-971Crossref PubMed Scopus (376) Google Scholar). NO is also known to activate multiple gene and cell signaling pathways through processes such as nitrosation and cGMP production (3Pfeilschifter J. Eberhardt W. Beck K.F. Pfluegers Arch. 2001; 442: 479-486Crossref PubMed Scopus (113) Google Scholar, 9Grisham M.B. Jourd'heuil D. Wink D.A. Aliment. Pharmacol. Ther. 2000; 14 Suppl. 1: 3-9Crossref PubMed Google Scholar). Furthermore, numerous studies have shown that NO has antitumor effects and that forced expression of iNOS causes regression of tumors (13Juang S.H. Xie K. Xu L. Shi Q. Wang Y. Yoneda J. Fidler I.J. Hum. Gene Ther. 1998; 9: 845-854Crossref PubMed Scopus (84) Google Scholar, 14Juang S.H. Xie K. Xu L. Wang Y. Yoneda J. Fidler I.J. Cancer Biother. Radiopharm. 1997; 12: 167-175Crossref PubMed Scopus (29) Google Scholar, 15Xie K. Huang S. Dong Z. Juang S.H. Gutman M. Xie Q.W. Nathan C. Fidler I.J. J. Exp. Med. 1995; 181: 1333-1343Crossref PubMed Scopus (324) Google Scholar, 16Xu W. Liu L. Charles I.G. FASEB J. 2002; 16: 213-215Crossref PubMed Scopus (105) Google Scholar). Krüppel-like factor 6 (KLF6) is a ubiquitously expressed member of the Krüppel-like family of transcription factors, which have characteristic Cys2/His2 zinc finger motifs and bind very similar “GC box” or “CACCC element” sites on DNA (17Bieker J.J. J. Biol. 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Nephrol. 2001; 12: 726-735Crossref PubMed Google Scholar, 23Inuzuka H. Nanbu-Wakao R. Masuho Y. Muramatsu M. Tojo H. Wakao H. Biochem. Biophys. Res. Commun. 1999; 265: 664-668Crossref PubMed Scopus (75) Google Scholar). KLF6 is rapidly induced in cells after acute injury and directly activates collagen α1 and TGF-β along with TGF-β receptor I and II genes, thereby mediating wound-healing mechanisms of fibrogenesis and extracellular matrix formation (18Ratziu V. Lalazar A. Wong L. Dang Q. Collins C. Shaulian E. Jensen S. Friedman S.L. Proc. Natl. Acad. Sci. U. S. A. 1998; 95: 9500-9505Crossref PubMed Scopus (237) Google Scholar, 24Kim Y. Ratziu V. Choi S.G. Lalazar A. Theiss G. Dang Q. Kim S.J. Friedman S.L. J. Biol. Chem. 1998; 273: 33750-33758Abstract Full Text Full Text PDF PubMed Scopus (229) Google Scholar). Recently, KLF6 has been shown to function as a tumor suppressor gene that was mutated in prostate cancer (25Narla G. Heath K.E. Reeves H.L. Li D. J. F.J. A.M. J.A. Friedman S.L. 2001; PubMed Scopus Google Scholar). The iNOS promoter a of and sites through the 16-kb of iNOS cell injury, heat shock, and have been to their by or (1Taylor B.S. Geller D.A. Shock. 2000; 13: 413-424Crossref PubMed Scopus (151) Google Scholar, B.S. R.W. Wang Q. Billiar T.R. Geller D.A. J. Biol. Chem. 1998; 273: Full Text Full Text PDF PubMed Scopus Google Scholar, Biochem. Biophys. Res. Commun. PubMed Scopus Google Scholar, J. J. J. Biol. Chem. 1998; 273: Full Text Full Text PDF PubMed Scopus Google Scholar). KLF6 and iNOS are involved in processes such as cell injury, wound tissue and of and the iNOS promoter multiple CACCC sites we that KLF6 to the iNOS gene and regulates its we have identified a transcriptional of we that KLF6 to CACCC sites the proximal 0.63-kb region and regulates the expression of the iNOS promoter in cell We also that in cells to the expression of KLF6 causes a increase in expression of the endogenous iNOS gene and Jurkat cells in with COS-7 cells in with cells T cells a T cell to the The human reporter constructs and have been Billiar T.R. Geller D.A. Proc. Natl. Acad. Sci. U. S. A. PubMed Scopus Google Scholar). The two CACCC sites in the 0.63-kb iNOS promoter mutated to the The by The gene and as The iNOS constructs have been Billiar T.R. Geller D.A. Proc. Natl. Acad. Sci. U. S. A. PubMed Scopus Google Scholar). The KLF6 expression and the have been Bocco J.L. A. J. Biol. Chem. 1997; Full Text Full Text PDF PubMed Scopus Google Scholar). The and a and and DNA of Jurkat T cells and COS-7 cells in following the The 6 COS-7 cells or Jurkat T cells in of was with DNA in to to The to and by the and The cells in a at and with of reporter activity was with of and of assay in a was in by with of the and the activity was to the activity. COS-7 cells with the KLF6 expression the cells and with and as E. P. W. Res. PubMed Scopus Google Scholar). The of the are as and its and and its The by with and by and as in as of at with the in of and of of and and at to the of the The was on a in The the and COS-7 cells with the KLF6 expression or the and cells two was and the was with both the Jurkat cells and T also to COS-7 the COS-7 cells subjected to KLF6 the cells also with or and subjected to The assay was following the of and 1998; Full Text Full Text PDF PubMed Scopus Google Scholar, R. 1998; Full Text Full Text PDF PubMed Scopus (198) Google Scholar, V. Biochem. Sci. 2000; Full Text Full Text PDF PubMed Scopus Google Scholar). KLF6 was to DNA by to a of The chromatin KLF6 with and at and with serum by with The the as as at with by with The with and and DNA was a DNA was with the proximal as as sites in the proximal 0.63-kb iNOS promoter and on a by with The immunoprecipitation as was cells an with and and The by as and and and and was by in time the increase in of the on a to the The expression of the gene in KLF6-transfected and NaCN-treated cells was as with and cells, gene expression was assay was KLF6-transfected and COS-7 cells in with as J. Clin. Invest. 1998; 101: PubMed Scopus Google and protein was a protein assay of the protein by and to The was with and The membranes and with the by The by of cell with in the of to to D. Biochem. PubMed Scopus Google Scholar, S.L. J. Pharmacol. 1995; 273: PubMed Scopus Google Scholar). the of which with conditions to a was with a with at and at and was by with a of of the was with 14 and as motifs in the region of the human iNOS gene is shown in the of KLF6 on the expression of human we reporter in COS-7 cells with the full-length 16-kb iNOS promoter and the KLF6 expression shown in of COS-7 cells with KLF6 induced the activity by with the the of iNOS with a we the of transcription factor and on the iNOS promoter. has been to bind multiple sites on the iNOS promoter and to its activity by in and by in cells following (1Taylor B.S. Geller D.A. Shock. 2000; 13: 413-424Crossref PubMed Scopus (151) Google Scholar). of COS-7 cells with and with the full-length iNOS promoter induced promoter activity by and These indicate that KLF6 can the transcription of the iNOS promoter at levels to induced by R.W. Taylor B.S. Shao L. Geller D.A. Proc. Natl. Acad. Sci. U. S. A. 2001; 98: 8638-8643Crossref PubMed Scopus (281) Google Scholar, B.S. R.W. Wang Q. Billiar T.R. Geller D.A. J. Biol. Chem. 1998; 273: Full Text Full Text PDF PubMed Scopus Google Scholar, Billiar T.R. Geller D.A. Proc. Natl. Acad. Sci. U. S. A. PubMed Scopus Google Scholar, Q.W. Y. Nathan C. J. Biol. Chem. Full Text PDF PubMed Google Scholar, Kim Biochem. Biophys. Res. Commun. 1999; PubMed Scopus Google Scholar). that KLF6 can activate the full-length 16-kb iNOS we to the region of the iNOS promoter that is the of transcription by KLF6 Billiar T.R. Geller D.A. Proc. Natl. Acad. Sci. U. S. A. PubMed Scopus Google Scholar). COS-7 and Jurkat cells with serial deletion constructs of the 16-kb iNOS reporter with cells with the that the of iNOS promoter activity similar in constructs with the full-length 16-kb iNOS promoter and studies have shown that KLF6 to the CACCC motifs of DNA J.L. K.F. J. Biol. Chem. 2000; Full Text Full Text PDF PubMed Scopus Google Scholar). The 0.63-kb iNOS has two CACCC and there are a of CACCC binding sites in the of the 16-kb iNOS promoter. of the 0.63-kb iNOS promoter by KLF6 that sites the iNOS promoter activity in the of CACCC sites in the proximal 0.63-kb the iNOS promoter with the CACCC sites was with the a increase in the activity with the KLF6 expression with with the was with the and whereas there was with the iNOS promoter region to The of is the of KLF6 to the of the 0.63-kb iNOS promoter by the two CACCC we constructs in which we mutated the proximal and CACCC sites and a with both CACCC sites studies with these revealed that the two which are by an on the of the 0.63-kb iNOS promoter. The CACCC to and of activity in COS-7 cells and Jurkat cells the proximal CACCC to and in COS-7 cells and Jurkat cells Mutation of both CACCC sites the KLF6-induced iNOS promoter that the CACCC motifs are the of KLF6 with the 0.63-kb iNOS promoter. These studies that the proximal and CACCC sites in the 0.63-kb iNOS promoter are promoter activity. that KLF6 to the human iNOS we two the CACCC at to −168 and −261 to in the 0.63-kb iNOS H. Mol. Med. PubMed Google Scholar). The and was KLF6-transfected COS-7 these cells bound to both by the 0.63-kb iNOS promoter and the of the an to shown in and the in was by that KLF6 directly with the iNOS promoter in The of a with and the of KLF6 the CACCC binding to KLF6 with iNOS in we primary and COS-7 The with by of and the proximal and CACCC binding sites in the 0.63-kb iNOS promoter COS-7 cells, the of the was significantly with the cells that increased expression of KLF6 its binding to the iNOS promoter. It has been in independent studies that iNOS and KLF6 are up-regulated during cell (3Pfeilschifter J. Eberhardt W. Beck K.F. Pfluegers Arch. 2001; 442: 479-486Crossref PubMed Scopus (113) Google Scholar, 4Ganster R.W. Taylor B.S. Shao L. Geller D.A. Proc. Natl. Acad. Sci. U. S. A. 2001; 98: 8638-8643Crossref PubMed Scopus (281) Google Scholar, V. Lalazar A. Wong L. Dang Q. Collins C. Shaulian E. Jensen S. Friedman S.L. Proc. Natl. Acad. Sci. U. S. A. 1998; 95: 9500-9505Crossref PubMed Scopus (237) Google Scholar, 24Kim Y. Ratziu V. Choi S.G. Lalazar A. Theiss G. Dang Q. Kim S.J. Friedman S.L. J. Biol. Chem. 1998; 273: 33750-33758Abstract Full Text Full Text PDF PubMed Scopus (229) Google Scholar). up-regulation of iNOS is by KLF6 conditions of we COS-7 and Jurkat cells and primary T with and cellular Sci. 2000; PubMed Scopus Google Scholar). shown in and with increased the binding of KLF6 to the iNOS promoter as by the increased of the We also subjected and primary T cells to heat serum starvation, and and the binding of KLF6 to the iNOS promoter by shown in with the cells, KLF6 binding to the iNOS promoter was increased in a similar in cell subjected to these conditions. These that the of KLF6 with the iNOS promoter is increased in cells subjected to conditions of and stimulation. we the of KLF6 a role in its binding to the iNOS promoter. and of KLF6 plays a role in binding to the iNOS we a immunoprecipitation or by with the protein the bound and immunoprecipitation with The that the KLF6 that bound to the iNOS promoter was in T cells, which was increased in and NaCN-treated cells we that KLF6 bound to the iNOS promoter in NaCN-treated The of the and immunoprecipitation the of findings of KLF6 binding to iNOS in conditions. the KLF6 and iNOS we the iNOS mRNA and protein and NO production in KLF6-transfected COS-7 We to the of mRNA on the time with shown in the production of KLF6 mRNA was induced by to and by with the KLF6 there was a increase in the iNOS which a increase to was increase in the to the of iNOS protein by KLF6 following KLF6 expression we at time and the of KLF6 and iNOS expression over KLF6 protein levels increased at 6 whereas iNOS protein levels increased at that KLF6 is to transcription and to produce iNOS protein 6 It was that an increase in KLF6 mRNA in cells is by an increase in KLF6 the of of KLF6 protein is also (18Ratziu V. Lalazar A. Wong L. Dang Q. Collins C. Shaulian E. Jensen S. Friedman S.L. Proc. Natl. Acad. Sci. U. S. A. 1998; 95: 9500-9505Crossref PubMed Scopus (237) Google Scholar). of these we that the of KLF6 protein to of KLF6 as a of protein Furthermore, NO as by assay NO and a increase in cells with KLF6 was increase in KLF6 or iNOS mRNA or protein or NO in cells and in cells subjected to which similar conditions as the KLF6-transfected These with the that KLF6 as a transactivator of the iNOS increased binding of KLF6 to the iNOS promoter in cells to to a the expression of KLF6 and iNOS in pathophysiological we the KLF6 and iNOS mRNA and protein and NO production in NaCN-treated COS-7 The cells with and in time of cells to the production of KLF6 mRNA was by shown in a of KLF6 mRNA was with levels and over levels by and with the KLF6 mRNA there was an increase in KLF6 protein by and the high sustained over a iNOS protein also induced by and the levels a by Furthermore, NO as by its and also a increase in the levels at 6 with a and sustained increase over we the of transcription in the of iNOS following the with findings and increased iNOS mRNA and protein expression following KLF6 and that KLF6 plays a role in iNOS following of cells with the the regulation of the human iNOS promoter by a member of the Krüppel-like family of transcription reporter gene and we have that a member of the Krüppel-like family of transcription factors, directly with the iNOS promoter in cells and with following cell injury, and stimulation. Furthermore, we have shown that up-regulated KLF6 both iNOS mRNA and protein which with the nitric oxide The that KLF6 is a transcriptional of iNOS conditions of and cell The human iNOS promoter is and is of the known (1Taylor B.S. Geller D.A. Shock. 2000; 13: 413-424Crossref PubMed Scopus (151) Google Scholar). regulation is and at multiple by multiple transcription factors, in to diverse conditions in a transcription such as and factor are known to regulate the iNOS promoter in in B.S. Geller D.A. Shock. 2000; 13: 413-424Crossref PubMed Scopus (151) Google Scholar). is ubiquitously expressed and regulates iNOS in multiple cell and the regulation of iNOS by is R.W. Taylor B.S. Shao L. Geller D.A. Proc. Natl. Acad. Sci. U. S. A. 2001; 98: 8638-8643Crossref PubMed Scopus (281) Google Scholar, B.S. R.W. Wang Q. Billiar T.R. Geller D.A. J. Biol. Chem. 1998; 273: Full Text Full Text PDF PubMed Scopus Google Scholar, Billiar T.R. Geller D.A. Proc. Natl. Acad. Sci. U. S. A. PubMed Scopus Google Scholar, Q.W. Y. Nathan C. J. Biol. Chem. Full Text PDF PubMed Google Scholar, Kim Biochem. Biophys. Res. Commun. 1999; PubMed Scopus Google Scholar). that the of iNOS by KLF6 is to that by the that KLF6 an important of the of at least CACCC sites in the 16-kb iNOS the KLF6-induced transcriptional of the 0.63-kb was similar to that of the 16-kb in COS-7 and Jurkat cells The CACCC sites in the 0.63-kb are in at and −261 with the CACCC the of which is at studies have similar binding of KLF6 to sites or sites in to in and in to the promoter KLF6 is known to bind the synthase promoter at two CACCC sites and to the promoter J.L. K.F. J. Biol. Chem. 2000; Full Text Full Text PDF PubMed Scopus Google Scholar). KLF6 was shown to with and both TGF-β receptor I and II at multiple KLF6 by binding to two sites and with with promoter sites Y. Ratziu V. Choi S.G. Lalazar A. Theiss G. Dang Q. Kim S.J. Friedman S.L. J. Biol. Chem. 1998; 273: 33750-33758Abstract Full Text Full Text PDF PubMed Scopus (229) Google Scholar). The of the TGF-β receptor II promoter the of to and to Y. Ratziu V. Choi S.G. Lalazar A. Theiss G. Dang Q. Kim S.J. Friedman S.L. J. Biol. Chem. 1998; 273: 33750-33758Abstract Full Text Full Text PDF PubMed Scopus (229) Google Scholar). it is very that the two CACCC in to the promoter in the 0.63-kb iNOS promoter are of the iNOS gene by that CACCC sites and and and in the iNOS promoter a role in regulation of the iNOS gene in the of proximal CACCC The that is very in Bocco J.L. A. J. Biol. Chem. 1997; Full Text Full Text PDF PubMed Scopus Google and that its as as Bocco J.L. A. J. Biol. Chem. 1997; Full Text Full Text PDF PubMed Scopus Google that the activity and binding of KLF6 to the iNOS promoter also by and are of transcription by their the and of are known to on its L. J.J. J. Biol. Chem. 1998; 273: Full Text Full Text PDF PubMed Scopus Google Scholar). studies have that two of and are of by H. J. J. Biol. Chem. 2001; 276: Full Text Full Text PDF PubMed Scopus Google Scholar, T. P. 2000; PubMed Scopus Google Scholar). that KLF6 to the iNOS promoter in and cells and that as as KLF6 to the iNOS promoter in NaCN-treated cells indicate that of KLF6 a very important role in gene The of is of iNOS by KLF6 have important in the of tissue injury and studies have the role of NO in NO by multiple NO J. Billiar T.R. 1999; PubMed Scopus Google by and S. J. M. A.M. J. Exp. Med. 1997; PubMed Scopus Google Scholar). NO the and Y. Y. Kim R. Billiar T.R. N. Y. Acad. Sci. 2002; PubMed Scopus Google Scholar). NO an increase in and the expression of heat proteins such as that have an of cells by by the of the and by of and C.W. J.L. J. Biochem. PubMed Scopus Google Scholar). NaCN-treated cells, we increased binding of KLF6 to the iNOS promoter in to increased levels of iNOS protein and NO production and a similar it has been shown that NO NaCN-treated Sci. 2000; PubMed Scopus Google Scholar). NO plays a role in wound healing and tissue studies on and wound the role of NO through up-regulation of iNOS has been D.T. Most D. Barbul A. Curr. Opin. Clin. Nutr. Metab. Care. 2000; 3: 197-204Crossref PubMed Scopus (73) Google Scholar, S.H. 1998; PubMed Scopus Google Scholar, S.H. PubMed Scopus Google Scholar). KLF6 plays a role in conditions of acute KLF6 was to healing of cells (18Ratziu V. Lalazar A. Wong L. Dang Q. Collins C. Shaulian E. Jensen S. Friedman S.L. Proc. Natl. Acad. Sci. U. S. A. 1998; 95: 9500-9505Crossref PubMed Scopus (237) Google and cells S. S. K. Y. J. Friedman S.L. 2000; 95: Google Scholar). that in and NaCN-treated cells, there is a binding of KLF6 to the iNOS thereby that KLF6 can its and effects by NO through iNOS Recently, KLF6 was to a tumor suppressor gene that was mutated in of prostate cancer and it was shown to in a through the (25Narla G. Heath K.E. Reeves H.L. Li D. J. F.J. A.M. J.A. Friedman S.L. 2001; PubMed Scopus Google Scholar). The role of iNOS is also (13Juang S.H. Xie K. Xu L. Shi Q. Wang Y. Yoneda J. Fidler I.J. Hum. Gene Ther. 1998; 9: 845-854Crossref PubMed Scopus (84) Google K. Huang S. Dong Z. Juang S.H. Gutman M. Xie Q.W. Nathan C. Fidler I.J. J. Exp. Med. 1995; 181: 1333-1343Crossref PubMed Scopus (324) Google Scholar, 16Xu W. Liu L. Charles I.G. FASEB J. 2002; 16: 213-215Crossref PubMed Scopus (105) Google Scholar). NO production by iNOS through its transcriptional up-regulation by KLF6 as the antitumor of findings that KLF6 to the human iNOS promoter and regulates its expression conditions of cell injury, and with in the of injury and We the and