Lipopolysaccharide induction of THP-1 cells activates binding of c-Jun, Ets, and Egr-1 to the tissue factor promoter.
These studies examine the molecular basis for increased transcription of tissue factor (TF) in THP-1 cells stimulated with lipopolysaccharide (LPS). DNase I footprinting identified six sites of protein-DNA interaction between -383 and the cap site that varied between control and induced extracts. Four footprints show qualitative differences in nuclease sensitivity. Footprints I (-85 to -52) and V (-197 to -175) are induction-specific and localize to regions of the promoter that mediate serum, phorbol ester, partial LPS response (-111 to +14), and the major LPS-inducible element (-231 to -172). Electrophoretic mobility shift assays with the -231 to -172 probe demonstrate JunD and Fos binding in both control and induced nuclear extracts; however, binding of c-Jun is only detected following LPS stimulation. Antibody inhibition studies implicate binding of Ets-1 or Ets-2 to the consensus site between -192 and -177, a region that contains an induction-specific footprint. The proximal region (-85 to -52), containing the second inducible footprint, binds Egr-1 following induction. These data suggest that LPS stimulation of THP-1 cells activates binding of c-Jun, Ets, and Egr-1 to the TF promoter and implicates these factors in the transcriptional activation of TF mRNA synthesis.
Groupp
E R
ER
Department of Pathology, University of Pittsburgh School of Medicine, Pennsylvania 15261, USA.
Donovan-Peluso
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D001483:Base Sequence; D002460:Cell Line; D018983:DNA Footprinting; D017931:DNA Primers; D004268:DNA-Binding Proteins / Q000378:metabolism*; D008070:Lipopolysaccharides / Q000378:metabolism*; D008969:Molecular Sequence Data; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D050784:Proto-Oncogene Protein c-ets-1; D011518:Proto-Oncogene Proteins / Q000378:metabolism*; D050783:Proto-Oncogene Proteins c-ets; D016755:Proto-Oncogene Proteins c-jun / Q000378:metabolism*; D013925:Thromboplastin / Q000235:genetics* / Q000378:metabolism; D014157:Transcription Factors / Q000378:metabolism*
D017931:DNA Primers; D004268:DNA-Binding Proteins; D008070:Lipopolysaccharides; D050784:Proto-Oncogene Protein c-ets-1; D011518:Proto-Oncogene Proteins; D050783:Proto-Oncogene Proteins c-ets; D016755:Proto-Oncogene Proteins c-jun; D014157:Transcription Factors; D013925:Thromboplastin