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* Department of Medical Microbiology, Center of Infectious Diseases, Leiden University Medical Center, Leiden, The Netherlands;
Institute of Biochemistry, Biocenter, Goethe-University Frankfurt, Frankfurt/Main, Germany;
Institut National de la Santé et de la Recherche Médicale Unité 725, Etablissement Français du Sang-Alsace, Strasbourg, France;
Department of Medicine, Cambridge Institute of Medical Research, Cambridge, United Kingdom;
¶ Department of Immunohematology and Blood Transfusion, Leiden University Medical Center, Leiden, The Netherlands; and
|| Institute of Immunology, Friedrich-Loeffler-Institute, Tuebingen, Germany
TAP translocates virus-derived peptides from the cytosol into the endoplasmic reticulum, where the peptides are loaded onto MHC class I molecules. This process is crucial for the detection of virus-infected cells by CTL that recognize the MHC class I-peptide complexes at the cell surface. The varicellovirus bovine herpesvirus 1 encodes a protein, UL49.5, that acts as a potent inhibitor of TAP. UL49.5 acts in two ways, as follows: 1) by blocking conformational changes of TAP required for the translocation of peptides into the endoplasmic reticulum, and 2) by targeting TAP1 and TAP2 for proteasomal degradation. At present, it is unknown whether UL49.5 interacts with TAP1, TAP2, or both. The contribution of other members of the peptide-loading complex has not been established. Using TAP-deficient cells reconstituted with wild-type and recombinant forms of TAP1 and TAP2, TAP was defined as the prime target of UL49.5 within the peptide-loading complex. The presence of TAP1 and TAP2 was required for efficient interaction with UL49.5. Using deletion mutants of TAP1 and TAP2, the 6+6 transmembrane core complex of TAP was shown to be sufficient for UL49.5 to interact with TAP and block its function. However, UL49.5-induced inhibition of peptide transport was most efficient in cells expressing full-length TAP1 and TAP2. Inhibition of TAP by UL49.5 appeared to be independent of the presence of other peptide-loading complex components, including tapasin. These results demonstrate that UL49.5 acts directly on the 6+6 transmembrane TAP core complex of TAP by blocking essential conformational transitions required for peptide transport.
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1 This work was supported by grants from the Dutch Diabetes Research Foundation (to D.K.-L.), the Dutch Cancer Society (Grant UL 2005-3259, to M.E.R. and E.J.H.J.W.), the M.W. Beijerinck Virology Fund of the Royal Academy of Arts and Sciences (to M.E.R.), and The Netherlands Organization for Scientific Research (Vidi Grant 917.76.330, to M.E.R.).
2 Current address: Department of Molecular Cell Biology, Leiden University Medical Center, 2300 RC, Leiden, The Netherlands.
3 Current address: Georg-Speyer-Haus, Institute of Biomedical Research, Paul-Ehrlich-Strasse 42-44, D-60596, Frankfurt am Main, Germany.
4 Address correspondence and reprint requests to Dr. Emmanuel J.H.J. Wiertz, Department of Medical Microbiology, Leiden University Medical Center, 2300 RC, Leiden, The Netherlands. E-mail address: Wiertz{at}lumc.nl
5 Abbreviations used in this paper: ER, endoplasmic reticulum; 
N, N-terminally truncated; β2m, β2-microglobulin; huTAP, human TAP; MDR, multiple drug resistance; NGFR, nerve growth factor receptor; PLC, peptide-loading complex; TM, transmembrane.
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