urea-mediated cross-presentation of soluble epstein-barr virus bzlf1 proteinurea-mediated cross-presentation可溶性的巴尔病毒bzlf1蛋白质.pdf

Urea-Mediated Cross-Presentation of Soluble Epstein- Barr Virus BZLF1 Protein 1 2 1 1 1 3 Sascha Barabas , Regina Gary , Tanja Bauer , Juha Lindner , Petra Lindner , Birgit Weinberger , 1 1 1 Wolfgang Jilg , Hans Wolf , Ludwig Deml * 1 Institute of Medical Microbiology, University of Regensburg, Germany, 2 Department of Hematology and Oncology, University of Erlangen-Nuernberg, Germany, 3 Institute for Biomedical Aging Research, Austrian Academy of Sciences, Austria Abstract Soluble extracellular proteins usually do not enter the endogenous human leukocyte antigen (HLA) I–dependent presentation pathway of antigen-presenting cells, strictly impeding their applicability for the re-stimulation of protein- specific CD8+ cytotoxic T lymphocytes (CTL). Here we present for the Epstein-Barr virus (EBV) BZLF1 a novel strategy that facilitates protein translocation into antigen-presenting cells by its solubilisation in high molar urea and subsequent pulsing of cells in presence of low molar urea. Stimulation of PBMC from HLA-matched EBV-seropositive individuals with urea- treated BZLF1 but not untreated BZLF1 induces an efficient reactivation of BZLF1-specific CTL. Urea-treated BZLF1 (uBZLF1) enters antigen-presenting cells in a temperature-dependent manner by clathrin-mediated endocytosis and is processed by the proteasome into peptides that are bound to nascent HLA I molecules. Dendritic cells and monocytes but also B cells can cross-present uBZLF1 in vitro. The strategy described here has potential for use in the development of improved technologies for the monitoring