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OR1 The bound peptides are the determinants for the potential of HLA-G mediated immune regulation
Human Immunology 78 (2017) 1–50
Contents lists available at ScienceDirect
www.ashi-hla.org
journal homepage: www.elsevier.com/locate/humimm
Tuesday, September 12, 2017 2:00 PM–3:30 PM Scholar Award Abstract Session
OR1
THE BOUND PEPTIDES ARE THE DETERMINANTS FOR THE POTENTIAL OF HLA-G MEDIATED IMMUNE REGULATION. Alexander A. Celik a, Gwendolin Simper a, Trevor Huyton b, Rainer Blasczyk a, Christina Bade-Doeding a. a Hannover Medical School, Hannover, Germany; bMax Planck Institute for Biophysical Chemistry, Goettingen, Germany. Aim: The essential role of HLA-G in immune tolerance and its expression frequency in immune privileged tissues is well known; however the specific importance of allelic subtypes in immune responses remains unclear. 14 allelic variants exist of which HLA-G*01:01, *01:03 and *01:04 are the most prevalent with these variants differing at residues 31 and 110, respectively. Systematic analysis of these subtypes was the aim of this study. Methods: In cytotoxicity assays using artificial K562 cells expressing the HLA-G variants and NK cells as effector cells the differential protective potential of HLA-G variants was analyzed. The inhibitory receptors ILT-2 and KIR2DL4 are known HLA-G ligands. To determine binding of HLA-G to these molecules, recombinant sHLA-G molecules were used to block inhibitory NK surface receptors. These experiments were repeated reciprocally by blocking or silencing the respective receptor and analyze the sHLA-G blocking sites. HLAG*01:01, G*01:03 and G*01:04 restricted peptide profiles were determined utilizing soluble HLA technology and mass spectrometry. Results: An increased protective potential of HLA-G*01:04 could be observed, G*01:04 demonstrated a stronger affinity to the inhibitory receptors on NK cells than G*01:01 or G*01:03 which we attribute to their altered peptide motifs. All variants exhibit a unique repertoire with marginal overlap, while G*01:04 differs in its peptide anchor profile substantially and appears more invariant than G*01:01 or G*01:03. Using the identified peptides bound to the G*01:04 binding groove, structural analysis explains the impact of the L110I mismatch on the HLA-G conformation and its differential interaction with NK cell receptors. Conclusions: For the first time a contribution of structural alterations within the HLA-G heavy chain for peptide specificity and NK cell engagement could be observed. These results will be a step towards understanding immune tolerance and guide towards immune therapeutic strategies.
Contents lists available at ScienceDirect
www.ashi-hla.org
journal homepage: www.elsevier.com/locate/humimm
Tuesday, September 12, 2017 2:00 PM–3:30 PM Scholar Award Abstract Session
OR1
THE BOUND PEPTIDES ARE THE DETERMINANTS FOR THE POTENTIAL OF HLA-G MEDIATED IMMUNE REGULATION. Alexander A. Celik a, Gwendolin Simper a, Trevor Huyton b, Rainer Blasczyk a, Christina Bade-Doeding a. a Hannover Medical School, Hannover, Germany; bMax Planck Institute for Biophysical Chemistry, Goettingen, Germany. Aim: The essential role of HLA-G in immune tolerance and its expression frequency in immune privileged tissues is well known; however the specific importance of allelic subtypes in immune responses remains unclear. 14 allelic variants exist of which HLA-G*01:01, *01:03 and *01:04 are the most prevalent with these variants differing at residues 31 and 110, respectively. Systematic analysis of these subtypes was the aim of this study. Methods: In cytotoxicity assays using artificial K562 cells expressing the HLA-G variants and NK cells as effector cells the differential protective potential of HLA-G variants was analyzed. The inhibitory receptors ILT-2 and KIR2DL4 are known HLA-G ligands. To determine binding of HLA-G to these molecules, recombinant sHLA-G molecules were used to block inhibitory NK surface receptors. These experiments were repeated reciprocally by blocking or silencing the respective receptor and analyze the sHLA-G blocking sites. HLAG*01:01, G*01:03 and G*01:04 restricted peptide profiles were determined utilizing soluble HLA technology and mass spectrometry. Results: An increased protective potential of HLA-G*01:04 could be observed, G*01:04 demonstrated a stronger affinity to the inhibitory receptors on NK cells than G*01:01 or G*01:03 which we attribute to their altered peptide motifs. All variants exhibit a unique repertoire with marginal overlap, while G*01:04 differs in its peptide anchor profile substantially and appears more invariant than G*01:01 or G*01:03. Using the identified peptides bound to the G*01:04 binding groove, structural analysis explains the impact of the L110I mismatch on the HLA-G conformation and its differential interaction with NK cell receptors. Conclusions: For the first time a contribution of structural alterations within the HLA-G heavy chain for peptide specificity and NK cell engagement could be observed. These results will be a step towards understanding immune tolerance and guide towards immune therapeutic strategies.