Peptide Repertoire Changes Caused by Defects in Antigen Processing

Peptide Repertoire Changes Caused by Defects in Antigen ProcessingByKristin Camfield LindDoctor of Philosophy in Molecular and Cell BiologyUniversity of California, BerkeleyProfessor Nilabh Shastri, ChairMHC I molecules display peptides derived from intracellular proteins on the cell surface for recognition by CD8+ T cells. Presentation of these peptide:MHC I complexes (pMHC I) is important for elimination of viruses and transformed cells. In order to generate an effective CD8 T cell response, a high-quality set of pMHC I must be presented. However, the editing mechanisms that select these optimal pMHC I are not clear. In these studies, we used biochemical and immunological methods to examine the role of two ER-resident pMHC I editors, tapasin and ERAAP in shaping the pMHC I repertoire. Analysis of tapasin-deficient mice revealed a qualitative loss as well as gain of some pMHC I. However, these changes did not overlap with the ERAAP-unedited pMHC I repertoire, indicating that tapasin edits peptides in a distinct way. Sequencing of peptides unique to tapasin-deficient cells revealed altered C-termini, suggesting that tapasin determines the peptide C-terminus while ERAAP shapes the peptide N-terminus. We separately examined the allele-specific effects of ERAAP-deficiency on the H-2d pMHC I repertoire. In addition to Kd and Dd, these mice express Ld, an MHC I molecule which is particularly affected by loss of ERAAP. We found that ERAAP-deficiency has distinct effects on the peptide repertoire presented by each MHC I molecule. Analysis of the peptide sequences revealed unique peptides bound to all three MHC I, although long, structurally unique peptides were only found on Dd and Kd. Furthermore, using a T cell hybridoma specific for loss of ERAAP function, we identified and characterized an ERAAP-unedited peptide presented by Dd. Together, these findings highlight the importance of both tapasin and ERAAP in determining an optimal pMHC I repertoire