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Key residues at the membrane‐distal surface of KACL , but not glycosylation, determine the functional interaction of the keratinocyte‐specific C ‐type lectin‐like receptor KACL with its high‐affinity receptor NK p65
Author(s) -
Bauer Björn,
Spreu Jessica,
Rohe Christina,
Vogler Isabel,
Steinle Alexander
Publication year - 2015
Publication title -
immunology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 2.297
H-Index - 133
eISSN - 1365-2567
pISSN - 0019-2805
DOI - 10.1111/imm.12432
Subject(s) - receptor , cd69 , mutant , siglec , biology , c type lectin , lectin , glycosylation , microbiology and biotechnology , monoclonal antibody , glycoprotein , antibody , gene , biochemistry , il 2 receptor , genetics , cytotoxic t cell , in vitro
Summary Keratinocyte‐associated C ‐type lectin ( KACL ) is a peculiar C‐type lectin‐like receptor ( CTLR ) due to its selective expression by human keratinocytes and cognate interaction with the genetically coupled CTLR NK p65. KACL and NK p65 are members of the CLEC 2 and NKRP 1 subfamilies of natural killer gene complex ( NKC )‐encoded CTLR , respectively. Most NKRP 1 molecules are expressed on NK cells and T cells and act as receptors of CLEC 2 glycoproteins with their genes being intermingled in a certain sub‐region of the mammalian NKC . The reasons for the tight genetic linkage of these dedicated receptor/ligand pairs are unknown, as is the physiological expression of NK p65. Recently, we reported that the CTLR NK p65 and KACL interact with high affinity, resulting in activation of NK p65‐expressing NK ‐92 MI cells. Here, we address the molecular basis of this high‐affinity interaction by analysing KACL mutants with KACL ‐specific monoclonal antibodies ( mA b), soluble NK p65 ( sNK p65) and NK ‐92 MI ‐ NK p65 cells. We find that none of the three N‐linked carbohydrates of KACL glycoproteins significantly contributes to KACL surface expression and NK p65 interaction. However, KACL mutants with non‐conservative amino acid substitutions of arginine 158 or isoleucine 161 abrogated binding of both KACL ‐specific mA b OMA 1 and sNK p65, well in line with the blockade of NK p65– KACL interaction by OMA 1. Accordingly, functional recognition of these KACL mutants by NK ‐92M‐ NK p65 cells was completely abolished. Arginine 158 and isoleucine 161 located at the membrane‐distal surface of KACL were defined as residues, decisively determining functional KACL – NK p65 interaction that is independent of KACL glycosylation.