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Processing of the DRB1*1103-restricted measles virus nucleoprotein determinant 185-199 in the endosomal compartment
S Demotz1, W Ammerlaan, P Fournier
1Institute of Biochemistry, University of Lausanne, Epalinges, Switzerland.
Abstract:
MHC class II molecules present to CD4+ T cells protein fragments which mostly derive from the extracellular and from the endosomal compartments. Determinants of cytosolic proteins are, however, also displayed by MHC class II molecules following pathways which are still not yet fully characterized. Here we describe the isolation of DRB1*1103-restricted T cell clones specific for the measles virus (MV) nucleoprotein peptide 185-199 (N185). Experiments were then conducted to delineate how this determinant is assembled with DR molecules. In vitro binding analyses indicated that complexes between the N185 peptide and DRB1*1103 protein are optimally constituted at pH 4-4.5. In cellular experiments it was observed that chloroquine, leupeptin and emetine, which are classical inhibitors of presentation of MHC class II-restricted antigens, when added during infection of B cells with MV, prevent presentation of the N185 determinant. In addition, it was found that the N185 determinant is efficiently presented when the nucleoprotein is exogenously provided to B cells, either by blocking MV fusion with the peptide FFG or by the use of purified nucleoprotein. In contrast, it was observed that nucleoprotein recombinant vaccinia virus (vv-N)-infected B cells weakly stimulated N185-specific T cells, indicating that the restricted localization of the nucleoprotein in the cytosol resulted in a poor presentation of the N185 determinant. Taken together, these findings suggest that it is prior to delivery of the nucleoprotein into the cytosol that the N185 determinant is efficiently assembled with newly synthesized DR molecules in the acidic environment of the endosomal compartment.
Insights
This study reveals how measles virus nucleoprotein peptides are presented by MHC class II molecules. Findings suggest peptide assembly occurs in the endosomal compartment before cytosolic delivery, impacting T cell responses.
Area of Science:
- Immunology
- Molecular Biology
- Virology
Background:
- MHC class II molecules typically present peptides from extracellular and endosomal compartments to CD4+ T cells.
- The presentation of cytosolic protein fragments by MHC class II molecules involves less understood pathways.
Purpose of the Study:
- To investigate the presentation pathway of a measles virus nucleoprotein (MV N) peptide (185-199) restricted by DRB1*1103.
- To elucidate how this determinant is assembled with DR molecules.
Main Methods:
- Isolation of DRB1*1103-restricted T cell clones specific for the MV N185-199 peptide.
- In vitro binding assays to determine optimal complex formation pH.
- Cellular experiments using MV-infected B cells and antigen presentation inhibitors (chloroquine, leupeptin, emetine).
- Exogenous delivery of nucleoprotein and recombinant vaccinia virus infection to assess presentation efficiency.
Main Results:
- Optimal binding of the N185 peptide with DRB1*1103 occurred at pH 4-4.5.
- Antigen presentation inhibitors blocked N185 determinant presentation in MV-infected B cells.
- Exogenous nucleoprotein facilitated efficient N185 determinant presentation.
- Intracellularly produced nucleoprotein via vaccinia virus resulted in weak N185-specific T cell stimulation.
Conclusions:
- The N185 determinant is efficiently assembled with DR molecules in the acidic endosomal compartment prior to nucleoprotein delivery into the cytosol.
- This pathway is crucial for effective presentation of cytosolic viral antigens by MHC class II molecules.