a)Ribbon diagram of the NKR-P1 CTLD. Secondary structure elements are labeled in different colors: helix α1 is red, helix α2 is yellow, and β-strands and loops are cyan. b)Comparison between NKR-P1 dimers formed by the glycosylated (cyan), deglycosylated free (green), and LLT1-bound (blue) forms of NKR-P1. c) Comparison between helices α1- and α2-centered dimerization of murine dectin-1 (magenta) and human LLT1 (green), respectively; helices α1 and α2 are shown in red and yellow. Structural alignments of dectin-1 and NKR-P1 homodimers and LLT1 and NKR-P1 homodimers, prepared by aligning only one monomer from each dimer, are shown on the right-hand side. Although the CTLD fold is conserved in each pair of the aligned monomers, the helix α1- and helix α2-centered dimers show inverse arrangement.
Signaling by the human C-type lectin-like receptor, natural killer (NK) cell inhibitory receptor NKR-P1, has a critical role in many immune-related diseases and cancer. C-type lectin-like receptors have weak affinities to their ligands; therefore, setting up a comprehensive model of NKR-P1-LLT1 interactions that considers the natural state of the receptor on the cell surface is necessary to understand its functions. Here we report the crystal structures of the NKR-P1 and NKR-P1:LLT1 complexes, which provides evidence that NKR-P1 forms homodimers in an unexpected arrangement to enable LLT1 binding in two modes, bridging two LLT1 molecules. These interaction clusters are suggestive of an inhibitory immune synapse. By observing the formation of these clusters in solution using SEC-SAXS analysis, by dSTORM super-resolution microscopy on the cell surface, and by following their role in receptor signaling with freshly isolated NK cells, we show that only the ligation of both LLT1 binding interfaces leads to effective NKR-P1 inhibitory signaling. In summary, our findings collectively support a model of NKR-P1:LLT1 clustering, which allows the interacting proteins to overcome weak ligand-receptor affinity and to trigger signal transduction upon cellular contact in the immune synapse.
Blaha, J., Skalova, T. Kalouskova, B., Skorepa, O., Cmunt, D., Grobarova, V.,Pazicky, S., Polachova, E., Abreu, C., Stransky, J., Koval, T., Duskova, J.,Zhao, Y.,Harlos, K., Hasek, J. Dohnalek, J., and Vanek, O.:
Structure of the human NK cell NKR-P1:LLT1 receptor: ligand complex reveals clustering in the immune synapse, Nature Comm. (2022)13:5022, https://doi.org/10.1038/s41467-022-32577-6