Two independent calcineurin-binding regions in the N-terminal domain of murine NF-ATx1 recruit calcineurin to murine NF-ATx1

J Immunol. 1999 Apr 15;162(8):4755-61.

Abstract

Intracellular calcium regulates events controlling nuclear translocation of nuclear factor of activated T cells (NF-AT). Calcium-dependent phosphatase calcineurin (CN) plays a central role in this process. Structural and functional analyses of the N-terminal domain of murine NF-ATx1, a member of the NF-AT family, have defined two distinct CN binding regions (CNBRs), CNBR1 and CNBR2, which are located in the region preceding the SP boxes of serine/proline-rich sequences and the region between the SP boxes and Rel similarity domain, respectively. The binding of murine NF-ATx1 (mNF-ATx1) to CN was abolished by deletion of these two regions, yet was unaffected by the individual deletion. In contrast, the nuclear translocation of mNF-ATx1 was much reduced when only CNBR2 was removed. Luciferase assay revealed that both regions are required for mNF-ATx1-dependent activation of the murine IL-2 promoter. Most importantly, recombinant CNBR2 bound CN with a higher affinity, and when expressed in Jurkat cells, it functioned as a dominant negative mutant that prevented the transcription driven by exogenous mNF-ATx1, probably by interfering with the function of CN. We propose that activation of mNF-ATx1 can be modulated through two distinct CN target regions. Our findings provide a new opportunity for pharmacological intervention with Ca2+-dependent signaling events.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Binding Sites / genetics
  • Binding Sites / immunology
  • Biological Transport / genetics
  • Biological Transport / immunology
  • Calcineurin / metabolism*
  • Cell Nucleus / genetics
  • Cell Nucleus / immunology
  • DNA-Binding Proteins / genetics
  • DNA-Binding Proteins / metabolism*
  • DNA-Binding Proteins / physiology
  • Interleukin-2 / genetics
  • Mice
  • NFATC Transcription Factors
  • Nuclear Proteins*
  • Peptide Fragments / genetics
  • Peptide Fragments / immunology*
  • Peptide Fragments / metabolism*
  • Protein Binding / genetics
  • Protein Binding / immunology
  • Protein Structure, Tertiary
  • Recombinant Fusion Proteins / genetics
  • Recombinant Fusion Proteins / metabolism
  • Recombinant Fusion Proteins / pharmacology
  • Sequence Deletion
  • Transcription Factors / genetics
  • Transcription Factors / metabolism*
  • Transcription Factors / physiology
  • Transcriptional Activation / immunology

Substances

  • DNA-Binding Proteins
  • Interleukin-2
  • NFATC Transcription Factors
  • Nuclear Proteins
  • Peptide Fragments
  • Recombinant Fusion Proteins
  • Transcription Factors
  • Calcineurin