The human perforin gene is a direct target of STAT4 activated by IL-12 in NK cells

Biochem Biophys Res Commun. 2002 Oct 11;297(5):1245-52. doi: 10.1016/s0006-291x(02)02378-1.

Abstract

IL-12 activates STAT4 by inducing tyrosine phosphorylation, homo-dimerization, and nuclear translocation in NK cells and thereby stimulates proliferation and activation of these cells. The pore-forming protein perforin is a key effector protein for NK cell- and cytotoxic T lymphocyte-mediated cytolysis. Here we demonstrate that IL-12 induces the expression of the perforin gene in human NK cell line, NKL. Electrophoretic mobility shift assays using a probe containing two putative STAT-binding sequences located at -1085 and -1059 in the human perforin gene showed that STAT4 or STAT5 activated by IL-12 or IL-2, respectively, in NKL cells binds this region. Further analyses using various probes with or without mutated STAT-binding sequences showed that, although either of the two tandem STAT-binding sequences binds STAT4 weakly, the presence of both is required for significant binding of activated STAT4 and for formation of the STAT4-DNA-binding complex with lower electrophoretic mobility. Furthermore, mutation of either of the tandem STAT-binding sequences abolished the IL-12-induced activation of the perforin gene promoter in reporter gene assays. These results indicate that the IL-12-induced expression of the perforin gene in NK cells is directly regulated by STAT4, which binds, most likely as a homo-tetramer, to the tandem STAT-binding sequences in the perforin gene promoter.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Base Sequence
  • Binding, Competitive
  • Blotting, Northern
  • COS Cells
  • Cytokines / metabolism
  • DNA, Complementary / metabolism
  • DNA-Binding Proteins / metabolism*
  • Genes, Reporter
  • Humans
  • Interleukin-12 / metabolism*
  • Killer Cells, Natural / metabolism*
  • Membrane Glycoproteins / genetics*
  • Membrane Glycoproteins / metabolism*
  • Milk Proteins*
  • Molecular Sequence Data
  • Mutation
  • Perforin
  • Phenotype
  • Pore Forming Cytotoxic Proteins
  • Promoter Regions, Genetic
  • Protein Binding
  • Recombinant Proteins / metabolism
  • STAT1 Transcription Factor
  • STAT3 Transcription Factor
  • STAT4 Transcription Factor
  • STAT5 Transcription Factor
  • STAT6 Transcription Factor
  • Time Factors
  • Trans-Activators / metabolism*
  • Ultraviolet Rays
  • Up-Regulation*

Substances

  • Cytokines
  • DNA, Complementary
  • DNA-Binding Proteins
  • Membrane Glycoproteins
  • Milk Proteins
  • Pore Forming Cytotoxic Proteins
  • Recombinant Proteins
  • STAT1 Transcription Factor
  • STAT1 protein, human
  • STAT3 Transcription Factor
  • STAT3 protein, human
  • STAT4 Transcription Factor
  • STAT4 protein, human
  • STAT5 Transcription Factor
  • STAT6 Transcription Factor
  • STAT6 protein, human
  • Trans-Activators
  • Perforin
  • Interleukin-12