PEPTIDE-MHC-STABILITY DETERMINES THE SIZE OF THE CD8+ T CELL RESPONSE TOWARD AN IMMUNORECESSIVE TUMOR ANTIGEN DETERMINANT
Open Access
- Author:
- Watson, Alan Michael
- Graduate Program:
- Microbiology and Immunology
- Degree:
- Doctor of Philosophy
- Document Type:
- Dissertation
- Date of Defense:
- February 15, 2011
- Committee Members:
- Todd Schell, Dissertation Advisor/Co-Advisor
Todd Schell, Committee Chair/Co-Chair
Neil David Christensen, Committee Member
David Joseph Spector, Committee Member
Mary E Truckenmiller, Committee Member
Ronald Paul Wilson, Committee Member - Keywords:
- Antigen Presentation
Cross-Priming
Peptide MHC Stability
Peptide
T cell
SV40 T Antigen
Tumor
Cancer
Mouse - Abstract:
- CD8+ T cells recognize peptide-determinants bound to major histocompatibility complex class-I (MHC-I) molecules on the surface of antigen presenting cells. Differences in the number of T cells responding to various peptide-determinants often results in the establishment of an immunodominance hierarchy in which T cells specific for one or more determinants predominate the overall response. T cells specific for determinants that mount a strong response are termed immunodominant whereas weaker responses are termed subdominant, cryptic or immunorecessive. A relationship between the affinity of peptide-determinants for MHC molecules and the size of both CD8+ and CD4+ T cell responses has been well established since the early 1990’s. Specifically, a few studies have established that the dissociation rate of peptides from their MHC complex (pMHC-stability) correlates strongly with immunodominance; those determinants that have a slow dissociation rate (high pMHC-stability) are generally dominant while rapid dissociation rates (low pMHC-stability) are often subdominant. However, the mechanism(s) that connect pMHC-stability and immunodominance is largely unknown. Using the virus-derived oncoprotein SV40 Large Tumor-Antigen (TAg) as a model we have addressed the relationship between pMHC-stability and immunodominance. TAg encodes four H-2b-restricted CD8+ T cell determinants, sites I, II/III, IV and V. Upon immunization with TAg, a well characterized CD8+ T cell immunodominance hierarchy is established such that IV>I>II/III; responses to site V TAg (489-497) are only detected following the deletion or inactivation of the other three determinants. The availability of T cell antigen receptor (TCR) transgenic mice that produce T cells specific for sites I and V make these determinants well suited for characterizing the differences between immunodominant and immunorecessive determinants. The immunorecessive nature of site V is due in part to inefficient cross-priming of naïve site V-specific T cells such that only a fraction of naïve site V-specific T cells undergo priming. In contrast, nearly all site I-specific T cells undergo cross-priming in response to the same immunization. The cause of this discrepancy is unknown. However, site V forms low-stability pMHC whereas site I forms high-stability pMHC. Thus, it may be the low-stability of site V that results in inefficient cross-priming. The contribution of pMHC-stability to the regulation of naïve T cell cross-priming has never been directly addressed in the TAg system or others. Thus, in this study we have investigated whether the pMHC-stability of site V affects site V-specific T cell cross-priming. In order to address this question, we identified two point mutants, Q489A and G490A, which enhanced the pMHC-stability of site V and conserved epitope recognition by site V-specific T cells. We incorporated the mutations into TAg and produced immortalized cell lines. Immunization with Q489A or G490A cell lines resulted in a detectable endogenous site V-specific T cell response. This is the first time that an endogenous site V-specific T cell response has been observed following immunization with cell lines expressing the three dominant TAg determinants. Thus Q489A and G490A overcame the immunorecessive nature of site V. Our data suggest that this novel phenotype results from two distinct mechanisms: 1) As pMHC-stability increases, so does the efficiency of naïve T cell cross-priming such that a greater fraction of naïve site V-specific T cells undergo activation. 2) As pMHC-stability increases, so does the duration of site V-specific T cell cross-priming, driving enhanced accumulation of responding T cells. Our results indicate that following immunization with TAg immortalized cell lines approximately one-third of TCR transgenic T cells specific for site V (TCR-V cells) undergo cross-priming. In contrast, immunization with cell lines expressing Q489A or G490A TAg results in a two fold increase in the number of TCR-V cells that undergo cross-priming. Using a highly sensitive T cell magnetic-enrichment protocol, we demonstrate for the first time that TAg immortalized cells prime previously undetectable populations of endogenous site V-specific T cells. Although technical limitations hinder direct determination of the efficiency of endogenous site V-specific T cell priming, extrapolation of our data suggests that endogenous site V-specific T cells also experience inefficient cross-priming. Altogether, our results suggest that pMHC-stability determines the fraction of site V-specific T cells that is cross-primed. Our results also suggest that the size of the site V-specific T cell response increases as the duration of detectable cross-priming increases. Our results indicate that the window for cross-priming of TCR-V cells is limited to approximately 24-48 hours. In contrast, cross-priming of TCR transgenic T cells specific for site I (TCR-I) is detected for at least seven days. We demonstrate that the Q489A and G490A substitutions resulted in an increase in the duration of site V-specific T cell cross-priming. Using two independent methods, our results suggest a direct relationship between the duration of site V-specific T cell cross-priming and the size of the site V-specific T cell response. We find that the duration of cross-priming determines the size of the T cell response via enhanced T cell accumulation of site V-specific T cells. Thus, our results suggest that pMHC-stability determines the size of the site V-specific T cell response by determining the duration of site V-specific T cell cross-priming. The results presented in this study suggest that pMHC-stability determines the size of the site V-specific T cell response by first determining the efficiency of naïve T cell cross-priming and subsequently facilitating the accumulation of activated T cells. Thus, the low peptide-MHC stability of site V contributes to its immunorecessive nature by reducing the fraction of T cells that are cross-primed and limiting their expansion. Our findings have implications for the optimization of T cell responses toward subdominant and immunorecessive determinants in the context of multivalent vaccines. Such optimization is critical for the successful development of tumor immunotherapy strategies and progress toward viral vaccines that limit immune escape.