Recognition of viral peptides by inhibitory and activating killer immunoglobulin-like receptors

Bastidas Legarda, Leidy Yamile (2020) Recognition of viral peptides by inhibitory and activating killer immunoglobulin-like receptors. University of Southampton, Doctoral Thesis, 205pp.

Record type: Thesis (Doctoral)

Abstract

Background and Aim: Natural killer (NK) cells are lymphocytes from the innate immunity
critical in the control of viral infections. NK cell functions against viruses are largely regulated
by the interaction between Killer Immunoglobulin-like receptors (KIR) and Major
Histocompatibility Complex class I (MHC-I):peptide ligands. Recently, our group reported
that highly conserved peptides derived from viruses of the family Flaviviridae bind the HLAC*
0102 allele and together induce strong KIR2DS2-mediated NK cell functions. Given the
enormous allelic diversity of the MHC-I locus, this thesis aimed to investigate the ability of
Flaviviridae-derived peptides to bind different Human Leukocyte Antigen C (HLA-C) alleles
and to activate NK cells through engagement with KIR2DS2.
Methods: An in vitro co-culture system using different combinations of target and effector
cell lines was optimized. As target cells, we used Transporter-associated with Antigen
Processing (TAP)-deficient or TAP-competent cells expressing different HLA-C alleles and
presenting exogenous or endogenous viral peptides containing the AT motif and derived
from the non-structural 3 (NS3) proteins of Hepatitis C virus (HCV; LNPSVAATL),
Dengue/Zika virus (DENV/ZIKV; IVDLMCHATF), Yellow Fever virus (YFV; VIDAMCHATL)
and West Nile/Japanese Encephalitis virus (WNV/JEV; IVDVMCHATL). As effector cells,
NKL cell lines or primary NK cells expressing inhibitory KIR2DL2 or activating KIR2DS2
receptors, were used. Activation of NK cells to different targets was determined by
assessing NK cell cytotoxicity, degranulation and inhibition of viral replication.
Results: Huh-7 cells expressing the HCV replicon and presenting endogenous viral
peptides in the context of HLA-C*0304 or HLA-C*0802 were strongly recognized by the
Natural Killer Cell Leukaemia (NKL) cell line expressing KIR2DS2; this effect was
independent of LNPSVAATL peptide. TAP-deficient 721.221 cells expressing HLA-C*0802
or HLA-C*0501 and loaded with IVDLMCHATF peptide strongly bound KIR2DS2-PE
tetramers; however only TAP-competent 721.221 cells expressing HLA-C*0802 and
endogenously presenting DENV/NS3-derived peptides induced strong NK cell effector
functions through canonical interaction with KIR2DS2. Finally, 721.221 cells co-expressing
HLA-C*0802 and IVDLMCHATF peptide generated robust NK cell degranulation and
cytotoxic activity. Unlike LNPSVAATL, IVDLMCHATF peptide is predicted to have a broad
HLA-C specificity.
Conclusion: This research identified a Flaviviral peptide carrying the highly conserved
“MCHAT” motif (IVDLMCHATF) that in complex with HLA-C*0802 specifically interacted
with KIR2DS2 leading to robust NK cell effector functions. This is the first viral peptide
defined that binds an activating KIR and more than one HLA-C type. These data imply that
Flaviviruses are credible drivers of the evolution of KIR2DS2.