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The structural basis for intramembrane assembly of an activating immunoreceptor complex

Abstract

Many receptors that activate cells of the immune system are multisubunit membrane protein complexes in which ligand recognition and signaling functions are contributed by separate protein modules. Receptors and signaling subunits assemble through contacts among basic and acidic residues in their transmembrane domains to form the functional complexes. Here we report the nuclear magnetic resonance (NMR) structure of the membrane-embedded, heterotrimeric assembly formed by association of the DAP12 signaling module with the natural killer (NK) cell–activating receptor NKG2C. The main intramembrane contact site is formed by a complex electrostatic network involving five hydrophilic transmembrane residues. Functional mutagenesis demonstrated that similar polar intramembrane motifs are also important for assembly of the NK cell–activating NKG2D-DAP10 complex and the T cell antigen receptor (TCR)–invariant signaling protein CD3 complex. This structural motif therefore lies at the core of the molecular organization of many activating immunoreceptors.

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Figure 1: Construct design and labeling strategy.
Figure 2: Structure of the DAP12TM-DAP12TM-NKG2CTM complex.
Figure 3: Structural comparison of ζζTM and DAP12TM receptor-binding sites.
Figure 4: Structure of the electrostatic network at the DAP12TM-DAP12TM-NKG2CTM binding site.
Figure 5: A similar electrostatic network governs the assembly of NKG2C-DAP12, NKG2D-DAP10 and TCR-CD3 complexes.
Figure 6: Proposed model for intramembrane immunoreceptor complex assembly.

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Acknowledgements

We thank S.C. Blacklow (Harvard Medical School) for pMM-LR6 vector; M.J. Call and members of the Chou lab for reading the manuscript and discussions; and J. Pyrdol for assistance with initial construct design and expression trials. Supported by the Helen Hay Whitney Foundation (M.E.C.), the Charles A. King Trust (M.E.C.) and the US National Institutes of Health (R01AI054520 to K.W.W., R01HL084329 to J.J.C. and EB002026 to the Center for Magnetic Resonance at the Massachusetts Institute of Technology, where NMR data were collected).

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M.E.C., K.W.W. and J.J.C. conceived of the study; M.E.C. designed and did all biochemical experiments, produced transmembrane peptide constructs and prepared NMR samples; M.E.C. and J.J.C. collected and analyzed NMR data and calculated structures; M.E.C. wrote the paper; and all authors contributed to editing of the manuscript.

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Correspondence to Kai W Wucherpfennig or James J Chou.

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Call, M., Wucherpfennig, K. & Chou, J. The structural basis for intramembrane assembly of an activating immunoreceptor complex. Nat Immunol 11, 1023–1029 (2010). https://doi.org/10.1038/ni.1943

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