Identification of a novel nuclear factor-kappaB sequence involved in expression of urokinase-type plasminogen activator receptor

Eur J Biochem. 2000 Jun;267(11):3248-54. doi: 10.1046/j.1432-1327.2000.01350.x.

Abstract

We have previously defined the promoter of human urokinase-type plasminogen activator receptor (uPAR) gene in a 188-bp fragment between bases -141 and +47 relative to the translation start site. Here, we report that a novel nuclear factor-kappaB (NF-kappaB)-like sequence (5'-GGGAGGAGTC-3') at -45 is located in the uPAR promoter and one of the two DNase I-protected regions, region I between bases -51 and -30. This NF-kappaB-like motif differs at positions 7-9 from the decameric consensus sequences of NF-kappaB (5'-GGGRNNYYCC-3' where R indicates A or G, Y indicates C or T, and N indicates any nucleotide) and at positions 1 and 7-9 from the kappaB-like motifs (5'-HGGARNYYCC-3' where H indicates A, C, or T, R indicates A or G, Y indicates C or T, and N indicates any nucleotide). Nuclear extracts from HCT116 cells contain proteins that specifically bind to the NF-kappaB-like site at position -45. Mutation of the NF-kappaB-like motif decreased the binding of transcription factor NF-kappaB and reduced the uPAR promoter activity in comparison with the wild-type sequences. Co-transfection with a dominant negative I-kappaB kinase-2 expression vector reduced uPAR promoter activity by 65-75%. These results demonstrate that a previously uncharacterized NF-kappaB motif is required for uPAR promoter activity.

Publication types

  • Comparative Study
  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • 5' Untranslated Regions / genetics*
  • Base Sequence
  • Binding Sites / genetics
  • Colonic Neoplasms / pathology
  • Consensus Sequence
  • DNA Footprinting
  • Gene Expression Regulation*
  • Gene Expression Regulation, Neoplastic
  • Genes, Reporter
  • Humans
  • I-kappa B Kinase
  • Luciferases / biosynthesis
  • Molecular Sequence Data
  • NF-kappa B / physiology*
  • Promoter Regions, Genetic / genetics*
  • Protein-Serine-Threonine Kinases / metabolism
  • Receptors, Cell Surface / biosynthesis
  • Receptors, Cell Surface / genetics*
  • Receptors, Urokinase Plasminogen Activator
  • Recombinant Fusion Proteins / biosynthesis
  • Recombinant Fusion Proteins / genetics
  • Regulatory Sequences, Nucleic Acid*
  • Sequence Alignment
  • Sequence Homology, Nucleic Acid
  • Transcription, Genetic*
  • Transfection
  • Tumor Cells, Cultured / metabolism

Substances

  • 5' Untranslated Regions
  • NF-kappa B
  • PLAUR protein, human
  • Receptors, Cell Surface
  • Receptors, Urokinase Plasminogen Activator
  • Recombinant Fusion Proteins
  • Luciferases
  • Protein-Serine-Threonine Kinases
  • CHUK protein, human
  • I-kappa B Kinase
  • IKBKB protein, human
  • IKBKE protein, human