Francisco A. Chaves

The relationship between immunodominance, DM editing, and the kinetic stability of MHC class II:peptide complexes

Summary:  Immunodominance refers to the restricted antigen specificity of T cells detected in the immune response after immunization with complex antigens. Despite the presence of many potential peptide epitopes within these immunogens, the elicited T‐cell response apparently focuses on a very limited number of peptides. Over the last two decades, a number of distinct explanations have been put forth to explain this very restricted specificity of T cells, many of which suggest that endosomal antigen processing restricts the array of peptides available to recruit CD4 T cells. In this review, we present evidence from our laboratory that suggest that immunodominance in CD4 T‐cell responses is primarily due to an intrinsic property of the peptide:class II complexes. The intrinsic kinetic stability of peptide:class II complexes controls DM editing within the antigen‐presenting cells and thus the initial epitope density on priming dendritic cells. Additionally, we hypothesize that peptides that possess high kinetic stability interactions with class II molecules display persistence at the cell surface over time and will more efficiently promote T‐cell signaling and differentiation than competing, lower‐stability peptides contained within the antigen. We discuss this model in the context of the existing data in the field of immunodominance.

Cutting Edge: CD4 T Cells Generated from Encounter with Seasonal Influenza Viruses and Vaccines Have Broad Protein Specificity and Can Directly Recognize Naturally Generated Epitopes Derived from the Live Pandemic H1N1 Virus

The unexpected emergence of pandemic H1N1 influenza has generated significant interest in understanding immunological memory to influenza and how previous encounters with seasonal strains influence our ability to respond to novel strains. In this study, we evaluate the memory T cell repertoire in healthy adults to determine the abundance and protein specificity of influenza-reactive CD4 T cells, using an unbiased and empirical approach, and assess the ability of CD4 T cells to recognize epitopes naturally generated by infection with pandemic H1N1 virus. Our studies revealed that most individuals have abundant circulating CD4 T cells that recognize influenza-encoded proteins and that a strikingly large number of CD4 T cells can recognize autologous cells infected with live H1N1 virus. Collectively, our results indicate that a significant fraction of CD4 T cells generated from priming with seasonal virus and vaccines can be immediately mobilized upon infection with pandemic influenza strains derived from antigenic shift.

The impact of DM on MHC class II–restricted antigen presentation can be altered by manipulation of MHC–peptide kinetic stability

DM edits the peptide repertoire presented by major histocompatibility complex class II molecules by professional antigen-presenting cells (APCs), favoring presentation of some peptides over others. Despite considerable research by many laboratories, there is still significant uncertainty regarding the biochemical attributes of class II–peptide complexes that govern their susceptibility to DM editing. Here, using APCs that either do or do not express DM and a set of unrelated antigens, we found that the intrinsic kinetic stability of class II–peptide complexes is tightly correlated with the effects of DM editing within APCs. Furthermore, through the use of kinetic stability variants of three independent peptides, we demonstrate that increasing or decreasing the kinetic stability of class II–peptide complexes causes a corresponding alteration in DM editing. Finally, we show that the spontaneous kinetic stability of class II complexes correlates directly with the efficiency of presentation by DM+ APCs and the immunodominance of that class II–peptide complex during an immune response. Collectively, these results suggest that the pattern of DM editing in APCs can be intentionally changed by modifying class II–peptide interactions, leading to the desired hierarchy of presentation on APCs, thereby promoting recruitment of CD4 T cells specific for the preferred peptides during an immune response.

Direct Ex Vivo Analyses of HLA-DR1 Transgenic Mice Reveal an Exceptionally Broad Pattern of Immunodominance in the Primary HLA-DR1-Restricted CD4 T-Cell Response to Influenza Virus Hemagglutinin

ABSTRACT The recent threat of an avian influenza pandemic has generated significant interest in enhancing our understanding of the events that dictate protective immunity to influenza and in generating vaccines that can induce heterosubtypic immunity. Although antigen-specific CD4 T cells are known to play a key role in protective immunity to influenza through the provision of help to B cells and CD8 T cells, little is known about the specificity and diversity of CD4 T cells elicited after infection, particularly those elicited in humans. In this study, we used HLA-DR transgenic mice to directly and comprehensively identify the specificities of hemagglutinin (HA)-specific CD4 T cells restricted to a human class II molecule that were elicited following intranasal infection with a strain of influenza virus that has been endemic in U.S. human populations for the last decade. Our results reveal a surprising degree of diversity among influenza virus-specific CD4 T cells. As many as 30 different peptides, spanning the entire HA protein, were recognized by CD4 T cells, including epitopes genetically conserved among H1, H2, and H5 influenza A viruses. We also compared three widely used major histocompatibility class II algorithms to predict HLA-DR binding peptides and found these as yet inadequate for identifying influenza virus-derived epitopes. The results of these studies offer key insights into the spectrum of peptides recognized by HLA-DR-restricted CD4 T cells that may be the focus of immune responses to infection or to experimental or clinical vaccines in humans.

Immunodominance in CD4 T-cell responses: implications for immune responses to influenza virus and for vaccine design.

CD4 T cells play a primary role in regulating immune responses to pathogenic organisms and to vaccines. Antigen-specific CD4 T cells provide cognate help to B cells, a requisite event for immunoglobulin switch and affinity maturation of B cells that produce neutralizing antibodies and also provide help to cytotoxic CD8 T cells, critical for their expansion and persistence as memory cells. Finally, CD4 T cells may participate directly in pathogen clearance via cell-mediated cytotoxicity or through production of cytokines. Understanding the role of CD4 T-cell immunity to viruses and other pathogens, as well as evaluation of the efficacy of vaccines, requires insight into the specificity of CD4 T cells. This review focuses on the events within antigen-presenting cells that focus CD4 T cells toward a limited number of peptide antigens within the pathogen or vaccine. The molecular events are discussed in light of the special challenges that the influenza virus poses, owing to the high degree of genetic variability, unpredictable pathogenicity and the repeated encounters that human populations face with this highly infectious pathogenic organism.

Infection of HLA-DR1 Transgenic Mice with a Human Isolate of Influenza A Virus (H1N1) Primes a Diverse CD4 T-Cell Repertoire That Includes CD4 T Cells with Heterosubtypic Cross-Reactivity to Avian (H5N1) Influenza Virus

ABSTRACT The specificity of the CD4 T-cell immune response to influenza virus is influenced by the genetic complexity of the virus and periodic encounters with variant subtypes and strains. In order to understand what controls CD4 T-cell reactivity to influenza virus proteins and how the influenza virus-specific memory compartment is shaped over time, it is first necessary to understand the diversity of the primary CD4 T-cell response. In the study reported here, we have used an unbiased approach to evaluate the peptide specificity of CD4 T cells elicited after live influenza virus infection. We have focused on four viral proteins that have distinct intracellular distributions in infected cells, hemagglutinin (HA), neuraminidase (NA), nucleoprotein, and the NS1 protein, which is expressed in infected cells but excluded from virion particles. Our studies revealed an extensive diversity of influenza virus-specific CD4 T cells that includes T cells for each viral protein and for the unexpected immunogenicity of the NS1 protein. Due to the recent concern about pandemic avian influenza virus and because CD4 T cells specific for HA and NA may be particularly useful for promoting the production of neutralizing antibody to influenza virus, we have also evaluated the ability of HA- and NA-specific CD4 T cells elicited by a circulating H1N1 strain to cross-react with related sequences found in an avian H5N1 virus and find substantial cross-reactivity, suggesting that seasonal vaccines may help promote protection against avian influenza virus.

Analyses of the Specificity of CD4 T Cells During the Primary Immune Response to Influenza Virus Reveals Dramatic MHC-Linked Asymmetries in Reactivity to Individual Viral Proteins

Influenza is a contagious, acute respiratory disease that is a major cause of morbidity and mortality throughout the world. CD4 T cells play an important role in the immune response to this pathogen through the secretion of antiviral cytokines, and by providing help to CD8 T cells and B cells to promote the development of immunological memory and neutralizing antibody responses. Despite these well-defined roles in the anti-influenza response, our understanding of CD4 T-cell diversity and specificity remains limited. In the study reported here, overlapping peptides representing 5 different influenza viral proteins were used in EliSpot assays to enumerate and identify the specificity of anti-influenza CD4 T cells directly ex vivo following infection of mice with influenza virus, using two strains that express unrelated MHC class II molecules. These experiments evaluated whether the reactivity of CD4 T cells generally tracked with particular influenza proteins, or whether MHC preferences were the predominant factor dictating anti-CD4 T-cell specificity in the primary immune response. We made the unexpected discovery that the distribution of CD4 T-cell specificities for different influenza proteins varied significantly depending on the single class II molecule expressed in vivo. In SJL mice, the majority of epitopes were specific for the HA protein, while the NP protein dominated the response in C57BL/10 mice. Given the diversity of human MHC class II molecules, these findings have important implications for the ability to rationally design a vaccine that will generate a specific CD4 T-cell immune response that is effective across diverse human populations.

The utility and limitations of current Web-available algorithms to predict peptides recognized by CD4 T cells in response to pathogen infection.

The ability to track CD4 T cells elicited in response to pathogen infection or vaccination is critical because of the role these cells play in protective immunity. Coupled with advances in genome sequencing of pathogenic organisms, there is considerable appeal for implementation of computer-based algorithms to predict peptides that bind to the class II molecules, forming the complex recognized by CD4 T cells. Despite recent progress in this area, there is a paucity of data regarding the success of these algorithms in identifying actual pathogen-derived epitopes. In this study, we sought to rigorously evaluate the performance of multiple Web-available algorithms by comparing their predictions with our results—obtained by purely empirical methods for epitope discovery in influenza that used overlapping peptides and cytokine ELISPOTs—for three independent class II molecules. We analyzed the data in different ways, trying to anticipate how an investigator might use these computational tools for epitope discovery. We come to the conclusion that currently available algorithms can indeed facilitate epitope discovery, but all shared a high degree of false-positive and false-negative predictions. Therefore, efficiencies were low. We also found dramatic disparities among algorithms and between predicted IC50 values and true dissociation rates of peptide–MHC class II complexes. We suggest that improved success of predictive algorithms will depend less on changes in computational methods or increased data sets and more on changes in parameters used to “train” the algorithms that factor in elements of T cell repertoire and peptide acquisition by class II molecules.

Immunodominance of CD4 T Cells to Foreign Antigens Is Peptide Intrinsic and Independent of Molecular Context: Implications for Vaccine Design

Immunodominance refers to the restricted peptide specificity of T cells that are detectable after an adaptive immune response. For CD4 T cells, many of the mechanisms used to explain this selectivity suggest that events related to Ag processing play a major role in determining a peptide’s ability to recruit CD4 T cells. Implicit in these models is the prediction that the molecular context in which an antigenic peptide is contained will impact significantly on its immunodominance. In this study, we present evidence that the selectivity of CD4 T cell responses to peptides contained within protein Ags is not detectably influenced by the location of the peptide in a given protein or the primary sequence of the protein that bears the test peptide. We have used molecular approaches to change the location of peptides within complex protein Ags and to change the flanking sequences that border the peptide epitope to now include a protease site, and find that immunodominance or crypticity of a peptide observed in its native protein context is preserved. Collectively, these results suggest immunodominance of peptides contained in complex Ags is due to an intrinsic factor of the peptide, based upon the affinity of that peptide for MHC class II molecules. These findings are discussed with regard to implications for vaccine design.

Abortive activation of CD4 T cell responses during competitive priming in vivo

Immunodominance refers to the highly selective peptide reactivity of T cells during an immune response. In this study, we tested the hypothesis that persistence of peptide:class II complexes is one key parameter that selects the final specificity of CD4 T cells. We found that low-stability peptide:class II complexes support the initial priming and expansion of CD4 T cells, but the expansion becomes strikingly aborted in the presence of competitive T cell responses to unrelated peptides. Our experiments revealed that for inhibition to occur, the competitive responses must be initiated by the same antigen presenting cell, and it is not because of competition for MHC binding. These studies not only provide an insight into the events that regulate competitive CD4 T cell priming in vivo, but also provide a previously undescribed conceptual framework to understand the parameters that select the final specificity of the T cell repertoire during pathogen or vaccine-induced immune responses.