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The activating Ly49W and inhibitory Ly49G NK cell receptors display similar affinities for identical MHC class I ligands

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Abstract

The Ly49 receptor family plays an important role in the regulation of murine natural killer (NK) cell effector function. They recognize cell surface-expressed class I MHC (MHC-I) and are functionally equivalent to the killer Ig-related receptors (KIRs) in human NK cells. Ly49s exist in activating and inhibitory forms with highly homologous extracellular domains, displaying greater variability in the stalk regions. Inhibitory Ly49s can recognize self-MHC-I and therefore mediate tolerance to self. The role of activating Ly49 receptors is less clear. Some activating Ly49 receptors have been shown to recognize MHC-I molecules. The binding affinity of activating Ly49 receptors with MHC-I is currently unknown, and we sought to examine the affinities of two highly related receptors, an activating and an inhibitory Ly49 receptor, for their shared MHC-I ligands. The ectodomain of inhibitory Ly49G of the BALB/c mouse strain is highly similar to the Ly49W activating receptor in the nonobese diabetic (NOD) mouse. Recombinant soluble Ly49G and W were expressed, refolded, and analyzed for binding affinity with MHC-I by surface plasmon resonance. We found that Ly49G and Ly49W bound with similar affinity to the same MHC-I molecules. These results are a first determination of an activating Ly49 receptor affinity for MHC-I and show that, unlike prior results obtained with activating and inhibitory KIR receptors, functional homologues to Ly49 receptors, activating and inhibitory Ly49, can recognize common MHC-I ligands, with similar affinities.

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Abbreviations

CTLD:

C-type lectin-like domain

FACS:

Fluorescence-activated cell sorting

FITC:

Fluorescein isothiocyanate

IgG:

Immunoglobulin G

IPTG:

Isopropyl-1-thio-β-d-galactopyranoside

ITAM:

Immunomodulatory tyrosine-based activating motif

ITIM:

Immunomodulatory tyrosine-based inhibitory motif

KIR:

Killer cell immunoglobulin-like receptor

MCMV:

Murine cytomegalovirus

MHC:

Major histocompatibility complex

MHC-I:

MHC class I

NK:

Natural killer

PE:

Phycoerythrin

SPR:

Surface plasmon resonance

β2m:

β2-Microglobulin

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Acknowledgments

This work was supported by an operating grant from the Canadian Institutes for Health Research (to K.P.K.). B.J.M. was supported by the Canadian Institutes for Health Research and Alberta Heritage Foundation for Medical Research studentships.

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The authors declare that they have no conflicts of interest.

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Authors and Affiliations

  1. Department of Medical Microbiology and Immunology, University of Alberta, Edmonton, Alberta, T6G 2E1, Canada

    Brian J. Ma, Carla M. Craveiro Salvado & Kevin P. Kane

  2. Department of Medical Microbiology and Immunology, University of Alberta, 6-020 Katz Group Centre, Edmonton, Alberta, T6G 2E1, Canada

    Kevin P. Kane

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  1. Brian J. Ma
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  2. Carla M. Craveiro Salvado
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  3. Kevin P. Kane
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Correspondence to Kevin P. Kane.

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The manuscript does not contain clinical studies or patient data nor were animals used in this study.

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Ma, B.J., Craveiro Salvado, C.M. & Kane, K.P. The activating Ly49W and inhibitory Ly49G NK cell receptors display similar affinities for identical MHC class I ligands. Immunogenetics 66, 467–477 (2014). https://doi.org/10.1007/s00251-014-0777-2

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