title: GADD45a promoter regulation by a functional genetic variant associated with acute lung injury.
abstract: BACKGROUND
  Growth arrest DNA damage inducible alpha (GADD45a) is a stress-induced gene we have shown to participate in the pathophysiology of ventilator-induced lung injury (VILI) via regulation of mechanical stress-induced Akt ubiquitination and phosphorylation. The regulation of GADD45a expression by mechanical stress and its relationship with acute lung injury (ALI) susceptibility and severity, however, remains unknown.
  
  
  OBJECTIVE
  We examined mechanical stress-dependent regulatory elements (MSRE) in the GADD45a promoter and the contribution of promoter polymorphisms in GADD45a expression and ALI susceptibility.
  
  
  RESULTS
  Initial studies in GADD45a knockout and heterozygous mice confirmed the relationship of GADD45a gene dose to VILI severity. Human lung endothelial cells (EC) transfected with a luciferase vector containing the full length GADD45a promoter sequence (-771 to +223) demonstrated a >4 fold increase in GADD45a expression in response to 18% cyclic stretch (CS, 4 h) compared to static controls while specific promoter regions harboring CS-dependent MSRE were identified using vectors containing serial deletion constructs of the GADD45a promoter. In silico analyses of GADD45a promoter region (-371 to -133) revealed a potential binding site for specificity protein 1 (SP1), a finding supported by confirmed SP1 binding with the GADD45a promoter and by the significant attenuation of CS-dependent GADD45a promoter activity in response to SP1 silencing. Separately, case-control association studies revealed a significant association of a GADD45a promoter SNP at -589 (rs581000, G>C) with reduced ALI susceptibility. Subsequently, we found allelic variation of this SNP is associated with both differential GADD45a expression in mechanically stressed EC (18% CS, 4 h) and differential binding site of interferon regulatory factor 7 (IRF7) at this site.
  
  
  CONCLUSIONS
  These results strongly support a functional role for GADD45a in ALI/VILI and identify a specific gene variant that confers risk for ALI.
authors.affiliation: Division of Pulmonary, Critical Care, Sleep and Allergy, Department of Medicine, University of Illinois at Chicago, Chicago, Illinois, United States of America.
authors.forename: Sumegha
authors.initials: S
authors.lastname: Mitra
authors.affiliation: Division of Pulmonary, Critical Care, Sleep and Allergy, Department of Medicine, University of Illinois at Chicago, Chicago, Illinois, United States of America.
authors.forename: Michael S
authors.initials: MS
authors.lastname: Wade
authors.affiliation: Division of Pulmonary, Critical Care, Sleep and Allergy, Department of Medicine, University of Illinois at Chicago, Chicago, Illinois, United States of America.
authors.forename: Xiaoguang
authors.initials: X
authors.lastname: Sun
authors.affiliation: Division of Pulmonary, Critical Care, Sleep and Allergy, Department of Medicine, University of Illinois at Chicago, Chicago, Illinois, United States of America.
authors.forename: Nurgul
authors.initials: N
authors.lastname: Moldobaeva
authors.affiliation: Centro de Investigacion Biomedica en red Enfermedades Respiratorias, Instituto de Salud Carlos III, Madrid, Spain; Research Unit, Hospital Universitario Nuestra Senora de Candelaria, Tenerife, Spain.
authors.forename: Carlos
authors.initials: C
authors.lastname: Flores
authors.affiliation: Division of Pulmonary, Critical Care and Sleep, University of Chicago, Chicago, Illinois, United States of America.
authors.forename: Shwu-Fan
authors.initials: SF
authors.lastname: Ma
authors.affiliation: Division of Pulmonary, Critical Care, Sleep and Allergy, Department of Medicine, University of Illinois at Chicago, Chicago, Illinois, United States of America; Department of Pediatrics, University of Illinois at Chicago, Chicago, Illinois, United States of America.
authors.forename: Wei
authors.initials: W
authors.lastname: Zhang
authors.affiliation: Arizona Health Sciences Center, University of Arizona, Tucson, Arizona, United States of America.
authors.forename: Joe G N
authors.initials: JG
authors.lastname: Garcia
authors.affiliation: Division of Pulmonary, Critical Care, Sleep and Allergy, Department of Medicine, University of Illinois at Chicago, Chicago, Illinois, United States of America.
authors.forename: Jeffrey R
authors.initials: JR
authors.lastname: Jacobson
mesh_terms: D055371:Acute Lung Injury / Q000235:genetics* / Q000378:metabolism / Q000473:pathology; D000328:Adult; D000368:Aged; D000818:Animals; D001665:Binding Sites; D018797:Cell Cycle Proteins / Q000235:genetics* / Q000378:metabolism; D042783:Endothelial Cells / Q000378:metabolism / Q000473:pathology; D005260:Female; D018628:Gene Dosage; D005786:Gene Expression Regulation; D017930:Genes, Reporter; D006579:Heterozygote; D006801:Humans; D050839:Interferon Regulatory Factor-7 / Q000235:genetics / Q000378:metabolism; D008156:Luciferases / Q000235:genetics / Q000378:metabolism; D008168:Lung / Q000378:metabolism / Q000473:pathology; D008297:Male; D040542:Mechanotransduction, Cellular; D051379:Mice; D018345:Mice, Knockout; D008875:Middle Aged; D009687:Nuclear Proteins / Q000235:genetics* / Q000378:metabolism; D020641:Polymorphism, Single Nucleotide; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D016329:Sp1 Transcription Factor / Q000235:genetics / Q000378:metabolism; D013314:Stress, Mechanical; D055397:Ventilator-Induced Lung Injury / Q000235:genetics* / Q000378:metabolism / Q000473:pathology; D000097642:GADD45 Proteins
chemical_list: D018797:Cell Cycle Proteins; D050839:Interferon Regulatory Factor-7; D008156:Luciferases; D009687:Nuclear Proteins; D016329:Sp1 Transcription Factor; C439070:GADD45A protein, human; D000097642:GADD45 Proteins; C499072:Gadd45a protein, mouse; C494235:Irf7 protein, mouse