title: Differential regulation of MMPs by E2F1, Sp1 and NF-kappa B controls the small cell lung cancer invasive phenotype.
abstract: BACKGROUND
  E2F1 transcription factor plays a vital role in the regulation of diverse cellular processes including cell proliferation, apoptosis, invasion and metastasis. E2F1 overexpression has been demonstrated in small cell lung cancer (SCLC), and extensive metastasis in early phase is the most important feature of SCLC. In this study, we investigated the involvement of E2F1 in the process of invasion and metastasis in SCLC by regulating the expression of matrix metalloproteinases (MMPs).
  
  
  METHODS
  Immunohistochemistry was performed to evaluate the expression of E2F1 and MMPs in SCLC samples in a Chinese Han population. The impact of E2F1 on invasion and metastasis was observed by transwell and wound healing experiments with depletion of E2F1 by specific siRNA. The target genes regulated by E2F1 were identified by chromatin immunoprecipitation (ChIP)-to-sequence, and the expressions of target genes were detected by real time PCR and western blotting. The dual luciferase reporter system was performed to analyze the regulatory relationship between E2F1 and MMPs.
  
  
  RESULTS
  E2F1 is an independent and adverse prognosis factor that is highly expressed in SCLC in a Chinese Han population. Knockdown of E2F1 by specific siRNA resulted in the downregulation of migration and invasion in SCLC. The expressions of MMP-9 and -16 in SCLC were higher than other MMPs, and their expressions were most significantly reduced after silencing E2F1. ChIP-to-sequence and promoter-based luciferase analysis demonstrated that E2F1 directly controlled MMP-16 expression via an E2F1 binding motif in the promoter. Although one E2F1 binding site was predicted in the MMP-9 promoter, luciferase analysis indicated that this binding site was not functionally required. Further study demonstrated that E2F1 transcriptionally controlled the expression of Sp1 and p65, which in turn enhanced the MMP-9 promoter activity in SCLC cells. The associations between E2F1, Sp1, p65, and MMP-9 were validated by immunohistochemistry staining in SCLC tumors.
  
  
  CONCLUSIONS
  E2F1 acts as a transcriptional activator for MMPs and directly enhances MMP transcription by binding to E2F1 binding sequences in the promoter, or indirectly activates MMPs through enhanced Sp1 and NF-kappa B as a consequence of E2F1 activation in SCLC.
authors.forename: Zunling
authors.initials: Z
authors.lastname: Li
authors.forename: Yanxia
authors.initials: Y
authors.lastname: Guo
authors.forename: Hanming
authors.initials: H
authors.lastname: Jiang
authors.forename: Tingguo
authors.initials: T
authors.lastname: Zhang
authors.forename: Changzhu
authors.initials: C
authors.lastname: Jin
authors.forename: Charles Y F
authors.initials: CY
authors.lastname: Young
authors.affiliation: Department of Biochemistry and Molecular Biology, Shandong University School of Medicine, Jinan, China. lyuanhq@sdu.edu.cn.
authors.forename: Huiqing
authors.initials: H
authors.lastname: Yuan
mesh_terms: D000368:Aged; D000369:Aged, 80 and over; D045744:Cell Line, Tumor; D049109:Cell Proliferation; D050687:E2F1 Transcription Factor / Q000235:genetics* / Q000378:metabolism; D015972:Gene Expression Regulation, Neoplastic; D006801:Humans; D008297:Male; D053513:Matrix Metalloproteinase 16 / Q000096:biosynthesis*; D020780:Matrix Metalloproteinase 9 / Q000096:biosynthesis*; D008875:Middle Aged; D009361:Neoplasm Invasiveness / Q000235:genetics; D009362:Neoplasm Metastasis / Q000235:genetics; D011494:Protein Kinases / Q000235:genetics / Q000378:metabolism*; D034741:RNA, Small Interfering; D055752:Small Cell Lung Carcinoma / Q000235:genetics* / Q000473:pathology; D051996:Transcription Factor RelA / Q000235:genetics / Q000378:metabolism*
chemical_list: D050687:E2F1 Transcription Factor; D034741:RNA, Small Interfering; D051996:Transcription Factor RelA; D011494:Protein Kinases; C065999:Sp1 kinase; D053513:Matrix Metalloproteinase 16; D020780:Matrix Metalloproteinase 9
references.citation: Nat Rev Genet. 2010 Mar;11(3):204-20
references.pmid: 20142834
references.citation: PLoS One. 2010 Oct 15;5(10):e13442
references.pmid: 20976175
references.citation: J Biol Chem. 2004 Feb 13;279(7):5110-7
references.pmid: 14645238
references.citation: Carcinogenesis. 2012 Jun;33(6):1178-86
references.pmid: 22467239
references.citation: Cancer. 2006 Dec 1;107(11):2637-46
references.pmid: 17078054
references.citation: Mol Cell Biol. 1996 Jul;16(7):3327-37
references.pmid: 8668148
references.citation: J Pathol. 2004 Jul;203(3):744-53
references.pmid: 15221933
references.citation: Oncol Lett. 2013 Jan;5(1):3-11
references.pmid: 23255884
references.citation: J Clin Oncol. 1999 Jun;17(6):1802-8
references.pmid: 10561218
references.citation: Nucleic Acids Res. 2001 Nov 1;29(21):4361-72
references.pmid: 11691923
references.citation: Proc Natl Acad Sci U S A. 2008 Dec 30;105(52):20734-9
references.pmid: 19088195
references.citation: J Mol Biol. 1999 Nov 12;293(5):1005-15
references.pmid: 10547281
references.citation: Oncogene. 2005 May 12;24(21):3525-34
references.pmid: 15735752
references.citation: Pharmacogenomics. 2011 Apr;12(4):535-46
references.pmid: 21521025
references.citation: Biochem J. 2012 Jul 15;445(2):229-36
references.pmid: 22519702
references.citation: Clin Cancer Res. 2009 Mar 1;15(5):1741-6
references.pmid: 19240177
references.citation: Cancer Lett. 2011 Aug 28;307(2):141-8
references.pmid: 21511395
references.citation: Circulation. 2002 Nov 19;106(21):2707-13
references.pmid: 12438297
references.citation: Mol Cell Biol. 1996 Apr;16(4):1668-75
references.pmid: 8657142
references.citation: PLoS One. 2014 Jan 21;9(1):e85936
references.pmid: 24465799
references.citation: Lung Cancer. 2006 Mar;51(3):313-21
references.pmid: 16423426
references.citation: Cancer Res. 2000 Nov 1;60(21):5972-6
references.pmid: 11085515
references.citation: PLoS One. 2012;7(5):e38000
references.pmid: 22666431
references.citation: Oncogene. 2001 Mar 29;20(14):1678-87
references.pmid: 11313916
references.citation: Am J Pathol. 2012 Dec;181(6):1895-9
references.pmid: 23063657
references.citation: J Thorac Cardiovasc Surg. 2010 Apr;139(4):984-90; discussion 990
references.pmid: 20304142
references.citation: Cancer Res. 2000 May 1;60(9):2365-7
references.pmid: 10811110
references.citation: Am J Respir Cell Mol Biol. 1998 Jul;19(1):18-24
references.pmid: 9651176
references.citation: Neuroimmunomodulation. 1996 Mar-Jun;3(2-3):69-75
references.pmid: 8945720
references.citation: Oncogene. 1996 Dec 5;13(11):2379-86
references.pmid: 8957079
references.citation: Lung Cancer. 2012 Feb;75(2):235-41
references.pmid: 21764478
references.citation: Gene. 2007 Nov 15;403(1-2):89-97
references.pmid: 17881156
references.citation: Clin Cancer Res. 2007 Dec 1;13(23):6938-46
references.pmid: 18056168
references.citation: Lung Cancer. 2002 Mar;35(3):249-55
references.pmid: 11844598
references.citation: Pediatr Cardiol. 2011 Mar;32(3):263-6
references.pmid: 21293853
references.citation: Cancer Res. 2010 Jun 15;70(12):4931-40
references.pmid: 20516113
references.citation: Exp Hematol Oncol. 2012 Jul 20;1(1):18
references.pmid: 23210897
references.citation: Lancet Oncol. 2001 Jul;2(7):437-42
references.pmid: 11905738
references.citation: Int J Cancer. 2012 Feb 1;130(3):593-601
references.pmid: 21469139
references.citation: J Cell Physiol. 2007 Nov;213(2):355-64
references.pmid: 17654499
references.citation: Eur Respir J. 2002 Sep;20(3):703-9
references.pmid: 12358350
references.citation: Cell Mol Life Sci. 2013 Sep;70(17):3109-24
references.pmid: 23184252
references.citation: J Pathol. 2002 Oct;198(2):142-56
references.pmid: 12237873
references.citation: Cancer Res. 2012 Jan 15;72(2):516-26
references.pmid: 22086850