title: Elevated SP-1 transcription factor expression and activity drives basal and hypoxia-induced vascular endothelial growth factor (VEGF) expression in non-small cell lung cancer.
abstract: VEGF plays a central role in angiogenesis in cancer. Non-small cell lung cancer (NSCLC) tumors have increased microvascular density, localized hypoxia, and high VEGF expression levels; however, there is a lack of understanding of how oncogenic and tumor microenvironment changes such as hypoxia lead to greater VEGF expression in lung and other cancers. We show that NSCLC cells secreted higher levels of VEGF than normal airway epithelial cells. Actinomycin D inhibited all NSCLC VEGF secretion, and VEGF minimal promoter-luciferase reporter constructs were constitutively active until the last 85 base pairs before the transcription start site containing three SP-1 transcription factor-binding sites; mutation of these VEGF promoter SP-1-binding sites eliminated VEGF promoter activity. Furthermore, dominant negative SP-1, mithramycin A, and SP-1 shRNA decreased VEGF promoter activity, whereas overexpression of SP-1 increased VEGF promoter activity. Chromatin immunoprecipitation assays demonstrated SP-1, p300, and PCA/F histone acetyltransferase binding and histone H4 hyperacetylation at the VEGF promoter in NSCLC cells. Cultured NSCLC cells expressed higher levels of SP-1 protein than normal airway epithelial cells, and double-fluorescence immunohistochemistry showed a strong correlation between SP-1 and VEGF in human NSCLC tumors. In addition, hypoxia-driven VEGF expression in NSCLC cells was SP-1-dependent, with hypoxia increasing SP-1 activity and binding to the VEGF promoter. These studies are the first to demonstrate that overexpression of SP-1 plays a central role in hypoxia-induced VEGF secretion.
authors.affiliation: Centre for Respiratory Research, University of Nottingham, Nottingham, NG5 1PB, United Kingdom. karl.deacon@nottingham.ac.uk
authors.forename: Karl
authors.initials: K
authors.lastname: Deacon
authors.forename: David
authors.initials: D
authors.lastname: Onion
authors.forename: Rajendra
authors.initials: R
authors.lastname: Kumari
authors.forename: Susan A
authors.initials: SA
authors.lastname: Watson
authors.forename: Alan J
authors.initials: AJ
authors.lastname: Knox
mesh_terms: D002289:Carcinoma, Non-Small-Cell Lung / Q000098:blood supply* / Q000235:genetics / Q000378:metabolism* / Q000473:pathology; D015687:Cell Hypoxia; D045744:Cell Line, Tumor; D003609:Dactinomycin / Q000494:pharmacology; D005260:Female; D015972:Gene Expression Regulation, Neoplastic; D006657:Histones / Q000235:genetics / Q000378:metabolism; D006801:Humans; D000860:Hypoxia / Q000235:genetics / Q000378:metabolism* / Q000473:pathology; D008175:Lung Neoplasms / Q000098:blood supply* / Q000235:genetics / Q000378:metabolism* / Q000473:pathology; D008297:Male; D009363:Neoplasm Proteins / Q000096:biosynthesis* / Q000235:genetics; D009389:Neovascularization, Pathologic / Q000235:genetics / Q000378:metabolism / Q000473:pathology; D019384:Nucleic Acid Synthesis Inhibitors / Q000494:pharmacology; D011401:Promoter Regions, Genetic / Q000235:genetics; D011485:Protein Binding; D016329:Sp1 Transcription Factor / Q000096:biosynthesis* / Q000235:genetics; D042461:Vascular Endothelial Growth Factor A / Q000096:biosynthesis* / Q000235:genetics; D050880:p300-CBP Transcription Factors / Q000235:genetics / Q000378:metabolism; D000099224:p300-CBP-Associated Factor
chemical_list: D003609:Dactinomycin; D006657:Histones; D009363:Neoplasm Proteins; D019384:Nucleic Acid Synthesis Inhibitors; D016329:Sp1 Transcription Factor; D042461:Vascular Endothelial Growth Factor A; D050880:p300-CBP Transcription Factors; D000099224:p300-CBP-Associated Factor; C467484:VEGFA protein, human