Walters, L. C. | bioRxiv Science

bioRxiv2020

IgM Natural Antibodies Bind HLA-E-Leader Peptide Complexes and Modulate NK Cell Cytotoxicity

HLA-E is a non-classical class Ib molecule that has limited polymorphism and binds HLA class Ia leader peptides (VL9). HLA-E-VL9 complexes interact with the natural killer (NK) cell receptors NKG2A-C/CD94 and regulate NK cell-mediated cytotoxicity. Here we isolated a murine IgM antibody 3H4, that specifically recognized HLA-E-VL9 bound complexes and enhanced killing of HLA-E-VL9-expressing cells by an NKG2A+ NK cell line. Structural analysis revealed how 3H4 prevents CD94/NKG2A docking on HLA-E-VL9 by binding with an overlapping footprint. Upon in vitro maturation, an affinity-optimized 3H4 IgG showed enhanced NK killing of HLA-E-VL9-expressing cells. Remarkably, HLA-E-VL9-specific IgM autoantibodies with similar specificity and functions to 3H4 were subsequently isolated from naive B cells of cytomegalovirus (CMV)-negative, healthy male human donors. Thus, a repertoire of germline low affinity HLA-E-VL9-reactive antibodies are present in both naive human and murine B cell repertoires. These antibodies can enhance NK cell cytotoxicity and therefore have potential for therapeutic modulation of NK cell function.

immunology↗

bioRxiv2020

Detailed and atypical HLA-E peptide binding motifs revealed by a novel peptide exchange binding assay

Diverse SIV epitopes that bind the rhesus homologue of HLA-E, Mamu-E, have recently been identified in SIV-vaccine studies using a recombinant Rhesus cytomegalovirus (RhCMV 68-1) vector, where unprecedented protection against SIV challenge was achieved. Additionally, several Mycobacterial peptides identified both algorithmically and following elution from infected cells, are presented to CD8+ T cells by HLA-E in humans. Yet, a comparative and comprehensive analysis of relative HLA-E peptide binding strength via a reliable, high throughput in vitro assay is currently lacking. To address this we developed and optimised a novel, highly sensitive peptide exchange ELISA-based assay that relatively quantitates peptide binding to HLA-E. Using this approach we screened multiple peptides, including peptide panels derived from HIV, SIV and Mtb predicted to bind HLA-E. Our results indicate that although HLA-E preferentially accommodates canonical MHC class I leader peptides, many non-canonical, sequence diverse, pathogen-derived peptides also bind HLA-E, albeit generally with lower relative binding strength. Additionally, our screens demonstrate that the majority of peptides tested, including some key Mtb and SIV epitopes which have been shown to elicit strong Mamu-E-restricted T cell responses, either bind HLA-E extremely weakly or give signals that are indistinguishable from the negative, peptide-free controls.

immunology↗