MHC AFFINITY, PEPTIDE LIBERATION, T-CELL REPERTOIRE, AND IMMUNODOMINANCE ALL CONTRIBUTE TO THE PAUCITY OF MHC CLASS I-RESTRICTED PEPTIDES RECOGNIZED BY ANTIVIRAL CTL

MHC class I-restricted T cell responses to viral proteins focus on a l imited set of peptides. To better understand this phenomenon, we exami ned all of the 26 nonameric peptides encoded by the influenza virus A/ Puerto Rico/8/34 (PR8) conforming to the canonical K-d binding motif. Ten peptides bound strongly to K-d as assessed by a cell surface stabi lization assay, Five of these 10 induced in vitro secondary CD8(+) T c ell responses from splenocytes derived from PR8-immunized mice. The st rongest responses were induced by the two previously defined antigenic peptides, which ranked only second and fifth in relative binding affi nity. To examine the limiting factors in the immunogenicity of K-d-bin ding peptides, we produced recombinant vaccinia viruses (rVVs) express ing cytosolic or endoplasmic reticulum (ER)-targeted peptides, rVVs ex pressing ER-targeted versions of the 7 peptides with the highest relat ive affinities for K-d rescued K-d cell surface expression in T2 cells , while those expressing the 3 lowest affinity peptides did not. The i mmunogenicity of several, but not all, of the highest affinity peptide s was greatly enhanced when expressed as VV-encoded cytosolic or ER-ta rgeted peptides as compared with full length proteins. We conclude tha t limitations in the immunogenicity of class I binding peptides reflec ts, in order of decreasing importance, peptide liberation by cellular proteases, T cell repertoire, and TAP-mediated peptide transport. We a lso observed an additional important contributing factor: suppression of T cell responses to nondominant peptides by an immunodominant pepti de located in the same protein.