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* Unidad de Inmunología Viral, Centro Nacional de Microbiología, Instituto de Salud Carlos III, Majadahonda (Madrid), Spain; and
Centro de Biología Molecular Severo Ochoa, Consejo Superior de Investigaciones Cientificas/Universidad Autónoma de Madrid, Madrid, Spain
Most pathogen-derived peptides recognized by CD8+ CTL are produced by proteasomes and delivered to the endoplasmic reticulum by the TAP transporters associated with Ag processing. Alternative proteases also produce antigenic peptides, but their actual relevance is unclear. There is a need to quantify the contribution of these supplementary pathways in vitro and in vivo. A well-defined TAP-independent secretory route of Ag processing involves the trans-Golgi network protease furin. Quantitation of this route by using OVA constructs encoded by vaccinia viruses indicates that it provides approximately one-third of all surface complexes of peptide and MHC class I molecules. Generation of the epitope carboxyl terminus is a dramatic rate-limiting step, since bypassing it increased efficiency by at least 1000-fold. Notably, the secretory construct activated a similar percentage of Ag-specific CD8+ T cells in wild type as in TAP1-deficient mice, which allow only secretory routes but which have a 10- to 20-fold smaller CD8 compartment. Moreover, these TAP1–/– OVA-specific CD8+ T lymphocytes accomplished elimination of epitope-bearing cells in vivo. The results obtained with this experimental system underscore the potential of secretory pathways of MHC class I Ag presentation to elicit functional CD8+ T lymphocytes in vivo and support the hypothesis that noncytosolic processing mechanisms may compensate in vivo for the lack of proteasome participation in Ag processing in persons genetically deficient in TAP and thus contribute to pathogen control.
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1 This work was supported by grants from the European Union, the Spanish Ministerio de Ciencia e Innovación, the Instituto de Salud Carlos III, and the Comunidad de Madrid. F.M. was supported by the European Union, the Comunidad de Madrid, and the Instituto de Salud Carlos III, M.R. by the Juan de la Cierva program, S.I. by the Fondo de Investigaciones Sanitarias, M.R.-C. by the Instituto de Salud Carlos III, and P.d.L. by the European Union.
2 Both authors contributed equally.
3 Current address: Unidad de Investigación, Hospital Universitario Puerta del Mar, Cádiz, Spain.
4 Address correspondence and reprint requests to Dr. Margarita Del Val, Unidad de Inmunología Viral, Centro Nacional de Microbiología, Instituto de Salud Carlos III, Ctra. Pozuelo km 2, E-28220 Majadahonda (Madrid), Spain. E-mail address: mdval{at}isciii.es
5 Abbreviations used in this paper: ER, endoplasmic reticulum; BFA, brefeldin A; DC, dendritic cell; decRVKR-cmk, decanoyl-RVKR-chloro-methyl-ketone; HBc, hepatitis B virus core cytosolic protein; HBe, hepatitis B virus secretory pre-core protein; LC, lactacystin; LLnL, leucyl-leucyl-norleucinal; rVACV, recombinant vaccinia virus.
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