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Title:
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Direct Interrogation of Viral Peptides Presented by the Class I HLA of HIV-Infected T Cells
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Author:
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Yaciuk, Jane C.; Skaley, Matthew; Bardet, Wilfried; Schafer, Danijela Mojsilovic; Cate, Steven; Stewart, Christopher J.; McMurtrey, Curtis; Jackson, Rico Buchli; Olvera, Alex; Cedeño, Samandhy; Plana, Montserrat; Mothe, B.; Brander, Christian; West, John T.; Hildebrand, William
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Other authors:
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Universitat de Vic - Universitat Central de Catalunya. Càtedra de la Sida i Malalties Relacionades |
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Notes:
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Identification of CD8+ cytotoxic T lymphocyte (CTL) epitopes has traditionally relied upon testing of overlapping peptide libraries for their reactivity with T cells in vitro. Here, we pursued deep ligand sequencing (DLS) as an alternative method of directly identifying those ligands that are epitopes presented to CTLs by the class I human leukocyte antigens (HLA) of infected cells. Soluble class I HLA-A*11:01 (sHLA) was gathered from HIV-1 NL4-3-infected human CD4+ SUP-T1 cells. HLA-A*11:01 harvested from infected cells was immunoaffinity purified and acid boiled to release heavy and light chains from peptide ligands that were then recovered by size-exclusion filtration. The ligands were first fractionated by high-pH high-pressure liquid chromatography and then subjected to separation by nano-liquid chromatography (nano-LC)–mass spectrometry (MS) at low pH. Approximately 10 million ions were selected for sequencing by tandem mass spectrometry (MS/MS). HLA-A*11:01 ligand sequences were determined with PEAKS software and confirmed by comparison to spectra generated from synthetic peptides. DLS identified 42 viral ligands presented by HLA-A*11:01, and 37 of these were previously undetected. These data demonstrate that (i) HIV-1 Gag and Nef are extensively sampled, (ii) ligand length variants are prevalent, particularly within Gag and Nef hot spots where ligand sequences overlap, (iii) noncanonical ligands are T cell reactive, and (iv) HIV-1 ligands are derived from de novo synthesis rather than endocytic sampling. Next-generation immunotherapies must factor these nascent HIV-1 ligand length variants and the finding that CTL-reactive epitopes may be absent during infection of CD4+ T cells into strategies designed to enhance T cell immunity. |
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Subject(s):
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-Sida -- Tractament |
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Rights:
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(c) American Society for Microbiology
Tots els drets reservats
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Document type:
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Article
info:eu-repo/publishedVersion |
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Published by:
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American Socity for Microbiology
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