Strategy for Identifying Dendritic Cell-Processed CD4 T Cell Epitopes from the HIV Gag p24 ProteinReport as inadecuate




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Mass Spectrometry MS is becoming a preferred method to identify class I and class II peptides presented on major histocompability complexes MHC on antigen presenting cells APC. We describe a combined computational and MS approach to identify exogenous MHC II peptides presented on mouse spleen dendritic cells DCs. This approach enables rapid, effective screening of a large number of possible peptides by a computer-assisted strategy that utilizes the extraordinary human ability for pattern recognition. To test the efficacy of the approach, a mixture of epitope peptide mimics mimetopes from HIV gag p24 sequence were added exogenously to Fms-like tyrosine kinase 3 ligand Flt3L-mobilized splenic DCs. We identified the exogenously added peptide, VDRFYKTLRAEQASQ, and a second peptide, DRFYKLTRAEQASQ, derived from the original exogenously added 15-mer peptide. Furthermore, we demonstrated that our strategy works efficiently with HIV gag p24 protein when delivered, as vaccine protein, to Flt3L expanded mouse splenic DCs in vitro through the DEC-205 receptor. We found that the same MHC II-bound HIV gag p24 peptides, VDRFYKTLRAEQASQ and DRFYKLTRAEQASQ, were naturally processed from anti-DEC-205 HIV gag p24 protein and presented on DCs. The two identified VDRFYKTLRAEQASQ and DRFYKLTRAEQASQ MHC II-bound HIV gag p24 peptides elicited CD4+ T-cell mediated responses in vitro. Their presentation by DCs to antigen-specific T cells was inhibited by chloroquine CQ, indicating that optimal presentation of these exogenously added peptides required uptake and vesicular trafficking in mature DCs. These results support the application of our strategy to identify and characterize peptide epitopes derived from vaccine proteins processed by DCs and thus has the potential to greatly accelerate DC-based vaccine development.



Author: Leonia Bozzacco , Haiqiang Yu, Jörn Dengjel, Christine Trumpfheller, Henry A. Zebroski III, Nawei Zhang, Victoria Küttner, Beat

Source: http://plos.srce.hr/



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