Production of granulocyte/macrophage-colony-stimulating factor by human natural killer cells: Modulation by the p75 subunit of the interleukin 2 receptor and by the CD2 receptor

L. J. Levitt, A. Nagler, F. Lee, J. Abrams, M. Shatsky, D. Thompson

Research output: Contribution to journalArticlepeer-review

Abstract

Resting natural killer (NK) cells express the p75 chain of the IL-2 receptor (IL-2Rβ) and most NK cells express the CD2 (erythrocyte rosette) receptor. The cell adhesion molecule, LFA-3, is a natural co-ligand for CD2. Tac antigen (IL-2Rα), a p55 IL-2R subunit, can be expressed after NK activation and may play a role in IL-2-induced NK proliferation. Little is known of the molecular mechanisms underlying cytokine production in NK cells. We investigated the roles of IL-2Rα, IL-2Rβ, and CD2/LFA-3 in the molecular regulation of NK cell granulocyte/macrophage-colony-stimulating factor (GM-CSF) production. Enriched populations of peripheral blood NK cells were separated into CD16-positive and CD16-negative fractions by flow cytometry; positively selected cells were > 97% positive for CD16 (the FcIII receptor for IgG which is present on almost all NK cells), < 1% positive for the T cell antigen CD3, and did not demonstrate rearrangement of the T cell receptor β chain gene by Southern blot. NK cell supernatants were harvested after 3-4 d of incubation with 0-100 U/ml IL-2, or after incubation with anti-CD2 (T113) MAb and sheep red blood cells (SRBC are a homologue for LFA-3). Parallel cell aliquots were harvested at 3-16 h for transcriptional run-on assays, S1 nuclease assays, and actinomycin D mRNA t( 1/2 ) determinations. IL-2-activated NK supernatants contained large amounts of GM-CSF (178±35 pg/ml) by ELISA as did supernatants from CD2-activated NK cells (T113 MAb + SRBC: 212±42) vs. < 20 pg/ml for NK cells incubated alone or with either SRBC or T113 MAb alone. Sepharose-linked anti-CD3 MAb did not induce GM-CSF release from NK cells. By S1 analysis, both IL-2 and CD2 stimulation markedly augmented GM-CSF mRNA expression but with very different latencies of onset. IL-2Rβ MAb inhibited > 85% of GM-CSF release from IL-2-activated NK cells and markedly suppressed IL-2-induced GM-CSF mRNA expression, whereas IL-2Rα MAb even at 2,000-fold molar excess of IL-2 had little effect (< 10%) on either GM-CSF release or mRNA expression. Run-on assays showed that GM-CSF is constitutively transcribed in NK cells and that IL-2 and CD2-activated cells had a three- to fourfold increased rate of GM-CSF transcription compared to nonstimulated cells. The t( 1/2 ) of GM-CSF mRNA in IL-2-activated NK cells was identical to that of unstimulated NK cells (15 min), whereas GM-CSF mRNA t( 1/2 ) in CD2-activated NK cells was increased 2.5-fold. We conclude that GM-CSF production in NK cells is regulated by both the IL-2Rβ and the CD2 receptor but not by IL-2Rα, that both transcriptional and posttranscriptional signals act together to modulate the level of GM-CSF mRNA in NK cells, and that the molecular mechanisms underlying NK cell GM-CSF production are dependent in part on differential surface receptor activation.

Original languageEnglish
Pages (from-to)67-75
Number of pages9
JournalJournal of Clinical Investigation
Volume88
Issue number1
DOIs
StatePublished - 1991
Externally publishedYes

Keywords

  • Cytotoxicity
  • Granulocyte/macrophage-colony-stimulating factor
  • Interleukin 2
  • Natural killer cells
  • Receptor
  • Transcription

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