A single amino acid change in inhibitory killer cell Ig-like receptor results in constitutive receptor self-association and phosphorylation

J Immunol. 2015 Jan 15;194(2):817-26. doi: 10.4049/jimmunol.1401830. Epub 2014 Dec 12.

Abstract

Signaling by immunoreceptors is often initiated by phosphorylation of cytosolic tyrosines, which then recruit effector molecules. In the case of MHC class I-specific inhibitory receptors, phosphorylation of cytosolic tyrosine residues within ITIMs results in recruitment of a protein tyrosine phosphatase that blocks activation signals. Recent work showed that signaling by an HLA-C-specific killer cell Ig-like receptor (KIR) is independent of signaling by activation receptors. It is not known how ITIM phosphorylation is initiated and regulated. In this article, we show that substitution of His-36 in the first Ig domain of KIR2DL1 with alanine (KIR2DL1-H36A) resulted in constitutive KIR2DL1 self-association and phosphorylation, as well as recruitment of tyrosine phosphatase SHP-1. Furthermore, substitution of His-36 with a similar bulky amino acid, phenylalanine, maintained the receptor in its unphosphorylated state, suggesting that steric hindrance by the His-36 side chain prevents constitutive KIR2DL1 self-association and ITIM phosphorylation. The equally strong phosphorylation of KIR2DL1 and KIR2DL1-H36A after inhibition of tyrosine phosphatase by pervanadate suggested that KIR2DL1-H36A is selectively protected from dephosphorylation. We propose that KIR phosphorylation is controlled by the accessibility of ITIM to tyrosine phosphatases and that KIR binding to HLA-C must override the hindrance that His-36 puts on KIR2DL1 self-association. Expression of KIR2DL1-H36A on NK cells led to stronger inhibition of lysis of HLA-C(+) target cells than did expression of wild-type KIR2DL1. These results revealed that ITIM phosphorylation is controlled by self-association of KIR and that His-36 serves as a gatekeeper to prevent unregulated signaling through KIR2DL1.

Publication types

  • Research Support, N.I.H., Intramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Substitution / immunology*
  • Cell Line
  • HLA-C Antigens* / genetics
  • HLA-C Antigens* / immunology
  • Humans
  • Killer Cells, Natural / cytology
  • Killer Cells, Natural / immunology*
  • Phosphorylation / genetics
  • Phosphorylation / immunology
  • Protein Tyrosine Phosphatase, Non-Receptor Type 6* / genetics
  • Protein Tyrosine Phosphatase, Non-Receptor Type 6* / immunology
  • Receptors, KIR2DL1* / genetics
  • Receptors, KIR2DL1* / immunology
  • Signal Transduction* / genetics
  • Signal Transduction* / immunology

Substances

  • HLA-C Antigens
  • KIR2DL1 protein, human
  • Receptors, KIR2DL1
  • PTPN6 protein, human
  • Protein Tyrosine Phosphatase, Non-Receptor Type 6