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. 2000 Feb;99(2):235–242. doi: 10.1046/j.1365-2567.2000.00968.x

Increasing the frequency of T-cell precursors specific for a cryptic epitope of hen-egg lysozyme converts it to an immunodominant epitope

T H Thatcher 1, D P O’brien 1, S Altuwaijri 1, R K Barth 1
PMCID: PMC2327154  PMID: 10692042

Abstract

Efforts to understand the mechanisms that govern how immunodominant T-cell epitopes are selected from protein antigens have focused mostly on differences in the efficiency of processing and presentation of peptide/major histocompatibility complex (MHC) complexes by antigen-presenting cells, while little attention has been directed at the role of the T-cell repertoire. In this report, the influence of the T-cell repertoire on immunodominance was investigated using transgenic mice that express the β chain from a T-cell receptor specific for a cryptic Ek restricted epitope of hen-egg lysozyme, HEL85-96. In these mice, the frequency of HEL85-96-specific T-cell precursors is increased 10–20-fold over non-transgenic mice. Transgenic mice respond as well as non-transgenic controls to intact HEL, even though they respond poorly or not at all to a variety of other antigens, including the dominant H-2k restricted epitopes of HEL. Following immunization with native HEL, the only HEL peptide that could recall a response in vitro in the transgenic mice was HEL85-96. Therefore, this normally cryptic epitope is the sole immunodominant epitope in the transgenic mice, and this alteration in immune response is due solely to an increase in the frequency of specific T-cell precursors. An analysis of four additional H-2k restricted cryptic epitopes of HEL suggests that three are similarly limited by T-cell frequency, and that only one is consistent with a defect in efficient antigen presentation. This indicates that there are at least two different types of cryptic epitopes, one in which crypticity is caused by inefficient processing or presentation, and another in which the frequency of specific T-cell progenitors is limiting.

Introduction

Recognition of antigens by T cells occurs through interaction between the T-cell receptor (TCR) and discrete peptide fragments, which are processed from the antigen and presented by major histocompatibility complex (MHC) molecules of an antigen-presenting cell (APC). Although typical protein antigens possess numerous immunogenic peptide sequences, only a few of these, called dominant epitopes, contribute to the development of the T-cell response against the whole protein. Epitopes that are immunogenic in peptide form, but which do not participate in the immune response against the whole protein, are termed cryptic epitopes (reviewed in 1).

Cryptic epitopes have proven important in understanding self–non-self discrimination and autoimmunity.2,3 Several studies using transgene-derived neo-self antigens have shown that immune tolerance extends only to dominant epitopes, and that autologous T cells specific for cryptic epitopes of self proteins may be immune competent.4–7 Some autoimmune diseases are characterized by determinant spreading, in which the early stages are characterized by T cells reactive against one or a few dominant epitopes, while T-cell populations reactive against cryptic epitopes appear in the later stages of disease.8,9 Additionally, several groups have proposed using cryptic epitopes of tumour antigens as a means of targeting immunological destruction of tumours.10–12 Thus, the mechanisms that determine dominant and cryptic epitopes are of vital interest.

Most efforts to understand the generation of immunodominant and cryptic epitopes have focused on antigen processing and presentation.13,14 Some epitopes may not be presented efficiently by APCs because flanking amino acids hinder processing.15–17 When processing results in multiple peptides that can bind MHC, intermolecular competition for MHC binding may result in display of only some peptide/MHC complexes.18 These variations in antigen presentation result in a T-cell response directed against a subset of the possible immunogenic peptides.

T-cell responses to antigen depend not only on presentation of peptide/MHC complexes but also on the availability of specific T-cell precursors. The influence of the T-cell repertoire on immunodominance and crypticity of T-cell epitopes was investigated using transgenic mice that express the TCR β chain from a TH hybridoma which is specific for an I-Ek restricted cryptic epitope of hen-egg lysozyme (HEL), HEL85-96.19 The transgenic β chain is expressed on greater than 95% of T cells with tight allelic exclusion, and associates with a variety of TCR α chains produced through rearrangement of the endogenous α chain loci.20 Limiting dilution analysis revealed that the frequency of HEL85-96 specific T cells in naive transgenic mice is 1 in 28 000, compared with fewer than 1 in 300 000 in non-transgenic controls.21 Consequently, transgenic (Tg) mice had a greater response to immunization with HEL85-96 than non-transgenic (NTg) controls. Interestingly, Tg mice were found to respond poorly or not at all to all other antigens tested, including a dominant epitope of HEL, HEL46-61. This impaired response is correlated with a measured decrease in the frequency of T-cell precursors specific for other antigens.21 These results suggest that these transgenic mice provide a useful system in which to study the influence of the frequency of epitope-specific T cells on immunodominance and crypticity.

This report demonstrates that the increased frequency of HEL85-96 specific T cells in the Tg mice converts HEL85-96 from a cryptic to a dominant epitope, providing the first direct evidence that crypticity can be determined solely by the T-cell repertoire. Further, the ability of APCs incubated with whole HEL to present cryptic epitopes was investigated. Of five cryptic epitopes of HEL in H-2k mice, only one is not effectively presented as a peptide/MHC complex. The other four, including HEL85-96, are efficiently processed and presented, and must be cryptic due to factors in the T cell compartment. The suggestion that many cryptic epitopes are actually presented by APCs has important implications for autoimmunity and tolerance.

Materials and methods

Mice

Transgenic line TB57 carries the rearranged TCR β chain gene from T hybridoma AO1.T13.1, which is specific for an I-Ek restricted epitope of HEL, HEL85-96.19–21 TB57 mice were backcrossed for two generations onto the parental strain C57BR/cdJ (I-Ak, I-Ek). Hemizygous Tg offspring were distinguished from NTg litter mates by DNA dot blot hybridization of genomic DNA to a cDNA probe specific for the TCR Vβ8.3 gene segment used in the transgene.22 Tg and NTg litter mates were used as experimental and control animals, respectively. C57BR/cdJ mice were purchased from Jackson Laboratories (Bar Harbor, ME). Mice were bred and housed in the University of Rochester Cancer Center Transgenic Mouse Facility.

Synthetic peptides

Synthetic peptides corresponding to portions of HEL were synthesized by Macromolecular Resources (Colorado State University, Fort Collins, CO) except for HEL33-53 and HEL116-129, which were prepared by Bio-Synthesis, Inc. (Lewisville, TX). The sequences are, HEL1-18, KVFGRCELAAAMKRHGLD; HEL13-35, KRHGLDNYRGYSLG-NWVCAAKFE; HEL25-43, LGNWVCAAKFESNFNTQAT;HEL33-53, KFESNFNTQATNRNTDGSTDY; HEL46-61, NTDGSTDYGILQINSR; HEL71-85, GSRNLCNIPCSALLS; HEL85-96, SSDITASVNCAK; HEL87-96, DITASVNCAK; HEL96-118, KKIVSDGDGMNAWVAWRNRCKGT; HEL116-129, KGTDVQAWIRGCRL.

Immunizations and T-cell proliferation assays

Groups of three to eight mice were immunized with 3·5 nmol synthetic HEL peptide or whole HEL (Sigma, St. Louis, MO) per hind foot pad. Antigens were dissolved in saline and emulsified in an equal volume of complete Freund’s adjuvant (Sigma). After 8 days, mice were killed and the popliteal lymph nodes were removed. Lymph node cell cultures were performed as described.6 Cells were cultured for 4 days with 1 µCi [methyl-3H]thymidine (Dupont NEN Products, Boston, MA) in 25 µl HL-1 medium added for the last 18 hr. Background proliferation was measured in wells without added antigen. Cells were harvested onto glass fibre filters using a Packard Micromate 196 cell harvester (Packard Co., Meriden, CT) and 3H incorporation was counted with a Packard Matrix 96 direct beta counter. Results shown are the total number of counts registered over a 3-min counting window and are expressed as the mean of quadruplicate cultures (± SEM) after subtraction of background counts.

Results

Whole HEL is immunogenic in Tg mice

HEL85-96 was previously identified as a cryptic epitope based on its lack of participation in the response to intact HEL,23,24 and the observation that it escapes tolerance induction in HEL-tolerant mice.25 Subsequently, however, it was reported that this epitope can be recovered at relatively high yield from the surface of H-2k APCs that were fed intact HEL. In addition, T-cell hybrids specific for HEL85-96/Ek could be isolated from the spleens of mice immunized with intact HEL, although at a much lower frequency than that of T-cell hybrids that recognize the immunodominant, Ak restricted epitope, HEL46-61.26 One interpretation of these data is that the poor response of HEL-immunized mice to HEL85-96 is caused by a relatively low frequency of T-cell precursors specific for HEL85-96/Ek. In order to test this hypothesis, we examined the intact HEL response of mice transgenic for the TCR β-chain gene from an HEL85-96/Ek-restricted T-cell hybrid.20 These mice were shown previously to display an elevated immune response to HEL85-96 peptide and a 10–20-fold increased frequency of HEL85-96 specific T cells, relative to non-transgenic mice. Tg mice also failed to respond to a panel of other antigens, which is correlated with an at least 10-fold reduced frequency of antigen-specific T cells.21 Lymph nodes from Tg and NTg mice were harvested after immunization with whole HEL and tested for the ability to proliferate in response to whole HEL in vitro. As shown in Fig. 1, the Tg mice respond well to priming with whole HEL. As these Tg mice respond poorly to all other antigens tested, including a dominant epitope of HEL, HEL46-61, this result suggests that HEL85-96 is the sole immunodominant epitope of HEL in the Tg mice. However, the possibility that the strong response to whole HEL seen in the Tg mice is due to some combination of dominant and subdominant epitopes other than HEL85-96 or HEL46-61 was not ruled out. To address this issue, the dominant and cryptic epitopes of HEL in the Tg mice were mapped.

Figure 1.

Figure 1

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Transgenic mice respond to immunization with whole HEL. Groups of three Tg and three NTg mice were immunized with whole HEL. Lymph node cells were harvested and challenged in vitro with whole HEL, and proliferation was measured by [3H]thymidine incorporation (see Materials and methods). Filled triangles, NTg; open triangles, Tg. Background counts for this experiment were 3143 for NTg and 2907 for Tg and were subtracted from the results shown. Results are representative of six independent experiments.

Only peptides containing the cryptic epitope HEL85-96 are strongly immunogenic in the TCR β Tg mice

Before mapping the dominant and cryptic epitopes of HEL in the Tg mice, it was necessary to determine which epitopes, if any, were immunogenic. A set of peptides spanning HEL was synthesized (Fig. 2), corresponding to the previously described dominant, subdominant and cryptic HEL epitopes in H-2k mice.23,27 The immune response to the HEL85-96 epitope was evaluated using two synthetic peptides; p85-96, which corresponds to the entire epitope, and p87-96, the core determinant recognized by HEL85-96 epitope-specific T cells.24 Each HEL peptide was immunogenic in NTg control mice (Fig. 3). In contrast, Tg mice responded to only three of eight HEL epitopes (HEL85-96, HEL13-35, and HEL46-61), with the response to HEL13-35 and HEL46-61 consistently lower than in NTg controls. Thus, only T cells specific for these three epitopes could contribute to the response seen against whole HEL in the Tg mice.

Figure 2.

Figure 2

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Schematic showing the H-2k restricted epitopes of HEL studied in this report. For peptide sequences, see Materials and methods.

Figure 3.

Figure 3

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Analysis of immunogenic HEL peptides in Tg and NTg mice. Groups of two to three Tg and NTg mice were immunized with the indicated HEL peptide. Lymph node cells were restimulated with the same peptide at a concentration of 14 µ m and proliferation was measured as in Fig. 1. Solid bars, NTg; striped bars, Tg. Background counts were 3741 or less, and were subtracted before graphing. Results are representative of two to four independent experiments.

The immunodominance hierarchy of HEL epitopes is altered in Tg mice

To identify the immunodominant epitopes of HEL in the Tg mice, Tg and NTg control mice were immunized with whole HEL, and T cells from the draining lymph nodes were isolated and stimulated with individual HEL peptides (Fig. 4). T cells from NTg mice responded strongly to HEL13-35 and HEL46-61, and weakly to HEL33-53 and HEL116-129, which defines them as dominant and subdominant epitopes, respectively. Because HEL1-18, HEL71-85, p85-96, p87-96, and HEL96-118 are immunogenic peptides which did not restimulate T cells primed with whole HEL, these define cryptic epitopes in the NTg mice, as does HEL25-43 (data not shown). Note that these results, observed in NTg mice derived from the C57BR/cdJ (Ak, Ek) strain, agree with results observed in B10.A (Ak, Ek) mice.23,24,27

Figure 4.

Figure 4

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Lymph node cells from Tg and NTg mice immunized with whole HEL are recalled in vitro by different peptides. Groups of eight Tg and NTg mice were immunized with whole HEL. Lymph node cells were harvested and restimulated in vitro with individual HEL peptides, and proliferation was measured by [3H]thymidine incorporation. (a) Peptides recalling a strong response in NTg T cells are classified as dominant epitopes. (b) Peptides recalling weak responses in the NTg T cells are classified as subdominant epitopes. (c) Peptides stimulating proliferation less than twice background in the NTg mice are classified as cryptic epitopes. Background counts were 2684–3900 for NTg and 2426–3264 for Tg, and were subtracted prior to graphing. Results shown are representative of three independent experiments.

T cells from Tg mice primed with whole HEL could be recalled in vitro only by p85-96 and p87-96 (Fig. 4c). HEL13-35 and HEL46-61, which were moderately immunogenic in the Tg mice as peptides, did not restimulate Tg T cells primed with whole HEL (Fig. 4a). Therefore, HEL85-96 is the sole immunodominant epitope of HEL in the Tg mice, and HEL13-35 and HEL46-61 are cryptic epitopes. It was previously demonstrated that the frequency of HEL85-96 specific T cells is increased 10–20-fold in the Tg mice,21 and the results of the peptide immunizations (Fig. 3) suggest that the Tg mice have reduced numbers of T-cell precursors specific for HEL13-35 and HEL46-61. Thus, altering the frequency of T-cell precursors in the TCR β chain Tg mice reverses the immunodominance hierarchy of HEL epitopes.

HEL85-96 specific T cells from Tg and NTg mice have the same average affinity

An alternative explanation for our results showing that HEL85-96 is a dominant epitope in Tg mice is that the presence of the transgene-derived TCR β chain gives rise to a population of HEL85-96 specific T cells with an extremely high affinity for their ligand. A high affinity TCR could possibly drive a vigorous immune response in the Tg mice even if HEL85-96 is presented by MHC at such a low level as to cause it to be cryptic in NTg mice. If this is the case, T cells from Tg mice should proliferate more vigorously than NTg T cells when challenged with very low concentrations of peptide. At concentrations of less than 2 µ m peptide, however, T cells from Tg mice immunized with p87-96 proliferate less than NTg T cells (Fig. 5a). Additionally, the EC50 for the Tg T cells was 4 µ m while the EC50 for NTg T cells was 3 µ m. It is clear that the average affinity of HEL85-96-responsive T cells from the Tg mice is equal to or even somewhat less than that of the NTg mice, despite the presence of the Tg TCR β chain. This in turn supports the conclusion that HEL85-96 is presented efficiently in both the Tg and NTg mice, and that the change in immunodominance hierarchy is due solely to a change in the frequency of specific T-cell precursors.

Figure 5.

Figure 5

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T cells from Tg and NTg mice have the same affinity for the HEL85-96 epitope. Tg and NTg mice were immunized withp87-96. Lymph node cells were harvested and restimulated with the same peptide, and proliferation was measured as in Fig. 1. Filled triangles, NTg; open triangles, Tg. (a) Tg T cells do not proliferate more than NTg T cells at low antigen concentrations. (b) The entire antigen dilution range, showing the maximum proliferation values. The EC50 for Tg cells is 4 µ m; the EC50 for NTg cells is 3 µ m.

APCs efficiently process and present four of five cryptic epitopes of HEL

The results presented above show that HEL85-96 is well presented by APCs and its dominance or crypticity is determined by the T-cell precursor frequency. To determine which epitopes of HEL were cryptic due to inefficient antigen presentation and which cryptic due to the T-cell population, specific T cells were elicited in NTg mice by immunization with peptide, and lymph node cells were harvested and stimulated in vitro with peptide or with whole HEL. Since the T cells are primed in vivo with peptide, this assay tests whether APCs in the lymph node cell cultures can present the same epitope from intact HEL. T cells specific for the dominant and subdominant epitopes can be stimulated in vitro by whole HEL, as might be expected (Fig. 6a). HEL85-96-specific T cells are also recalled by whole HEL, consistent with the results reported above. Of the remaining cryptic epitopes, T cells specific for HEL1-18, HEL25-43, and HEL71-85 can also be recalled in vitro with whole HEL (Fig. 6b). The efficiency of presentation, measured as the ratio of stimulation by intact antigen to stimulation by peptide, is in the same range for the dominant and subdominant epitopes (0·57–0·88) as these cryptic epitopes (0·51–1·20), indicating that these epitopes are efficiently presented from intact HEL and suggesting that their crypticity is determined by the T-cell population. Only HEL96-118-specific T cells can not be recalled by whole HEL, suggesting that HEL96-118 is cryptic because of failure to be efficiently presented to specific T cells. Thus, of five identified cryptic HEL epitopes, only one behaves in a manner consistent with poor processing and presentation from intact HEL.

Figure 6.

Figure 6

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Recall of peptide-elicited T cells by whole HEL discriminates between two types of cryptic epitopes. Groups of 3 NTg mice were immunized with the indicated HEL peptides. Lymph node cells were harvested and [3H]thymidine incorporation was measured after stimulation in vitro by the same peptide or by whole HEL at a concentration of 14 µ m. Solid bars, peptide; striped bars, whole HEL. (a) Dominant and subdominant epitopes as defined in Fig. 4. (b) Cryptic epitopes as defined in Fig. 4. Background counts have been subtracted. Results shown are representative of two independent experiments.

Discussion

Immunization with native antigen gives rise to a T-cell response focused on a few so-called dominant determinants while ignoring other, potentially immunogenic determinants. Because these cryptic epitopes figure prominently in autoimmunity and may play a role in antitumour immune responses, a thorough understanding of the mechanisms behind dominance and crypticity is needed. Broadly speaking, the T-cell response is determined by two factors; antigen processing and presentation, and the T-cell repertoire.14 Several studies have reported various mechanisms that influence the efficiency of peptide display by MHC, thereby giving rise to either dominant or cryptic epitopes.13,16,17,28,29 Ours is the first study to directly demonstrate that the frequency of epitope-specific T cells in a naive animal can determine whether an epitope is dominant or cryptic. Unlike a conventional TCR α/β double transgenic model, in which 90–95% of T cells share a single specificity, our model uses a single TCR β chain transgene. This results in a bias towards the HEL85-96 epitope, reflected in a 10–20-fold increase in the number of HEL85-96-specific precursors. Other specificities remain present, however, demonstrated by the weak but significant response to HEL13-35 and HEL46-61 (Fig. 2) and the full-strength response to pigeon cytochrome C seen with repeated antigen boosting.21 Although we21 and others26,30 have previously reported that certain cryptic epitopes are associated with a T-cell frequency lower than that for other, dominant, epitopes, here we have demonstrated that increasing the frequency of HEL85-96-specific T cells converts this epitope from cryptic to dominant, and decreasing the frequency of T cells specific for other epitopes renders HEL13-35 and HEL46-61 cryptic. Therefore, the frequency of epitope-specific T cells in a naive animal can be the sole determining factor in whether an epitope is immunodominant or cryptic.

The results presented here and elsewhere26 demonstrate that HEL85-96 is processed and presented from native HEL as efficiently as the dominant epitopes HEL13-35 and HEL46-61. Not only can APCs incubated with whole HEL stimulate epitope-specific T cells (Fig. 5), but immunization with whole HEL gives rise to an in vivo response against HEL85-96 in animals which only differ from the controls by the presence of the TCR β transgene (Fig. 3). But if HEL85-96 is efficiently presented in the control mice, why are these T cells not activated in vivo in normal mice in spite of their low frequency? The explanation we favour is that the early T-cell response to immunization with whole HEL is dominated by T-cell specificities that occur at high frequencies, which will encounter the antigen more often. These higher-frequency T-cell precursors then have a competitive advantage for access to cytokines, APCs, and other features of the lymph-node microenvironment, limiting the potential of low-frequency T-cell populations to expand.

To determine if any of the other cryptic epitopes of HEL were efficiently processed and presented, we elicited epitope-specific T cells via immunization with peptides, and then tested for the ability of APCs plus native HEL to stimulate proliferation. T cells specific for HEL1-18, HEL25-43 and HEL71-85 were recalled by whole HEL at least as efficiently as the dominant and subdominant epitopes, indicating that these epitopes are efficiently processed and presented from intact antigen. Only HEL96-118 specific T cells can not be recalled by whole HEL, suggesting that HEL96-118 is not efficiently presented by APCs. Unanue and coworkers have reported that immunization with synthetic peptides may give rise to T cells which are unable to recognize the same epitope when processed and presented from intact antigen.26,31 Therefore, the negative result with HEL96-118 is not conclusive proof that this epitope is not processed and presented. However, it is clear that T cells elicited by HEL1-18, HEL25-43, HEL71-85 and HEL85-96 can be restimulated by intact HEL, proving both that the majority of these T cells do not discriminate between the synthetic and naturally processed forms of the epitope, and that these epitopes are efficiently presented.

A review of the scientific literature suggests a certain ambiguity in the definition of cryptic epitopes. A broad definition, that cryptic epitopes are peptides that are immunogenic by themselves but which do not participate in the immune response against intact antigens, has been used by many investigators.32–36 Some epitopes that are cryptic according to these criteria have been shown to escape tolerance when the corresponding intact antigen is introduced as a toleragen.5–7,10,25 Based on results from these tolerance studies, the term cryptic epitope has been used by some investigators to describe any immunogenic peptide derived from a self-antigen.4,35,37–39 In contrast, a narrow definition has also been used, which stipulates that in addition to the above criteria, T cells primed by immunization with cryptic epitopes can not be restimulated by intact antigen.23 However, by focusing only on epitopes that are not presented, this additional criterion incorporates mechanistic constraints into the definition of cryptic epitopes. This narrower definition of cryptic epitopes excludes epitopes of self proteins that escape tolerance, but which appear to be efficiently presented.4,35,37–39 Furthermore, the narrower definition also excludes a number of epitopes that have been described as cryptic based on polyclonal activation studies, but which are efficiently presented by autologous APCs,4,33,36,39 or for which the presentation status is undetermined.32,34,40

Therefore, we have chosen to adopt the broad definition of cryptic epitopes that includes any immunogenic region of a protein that does not contribute to the immune response against native antigen, without reference to the underlying mechanism. Because T-cell responses arise from an interaction between APCs and the T-cell repertoire,14 we propose, where evidence of mechanism is available, to divide cryptic epitopes into two subclasses. Type I cryptic epitopes arise from any of several mechanisms that result in inefficient antigen processing and presentation.13,14,16,17,29 Type II cryptic epitopes are efficiently processed and presented, and are therefore probably cryptic due to limitations of the T-cell repertoire. Based on the evidence presented here, HEL96-118 is a Type I cryptic epitope of HEL in H-2k mice, and HEL1-18, HEL25-43, HEL71-85 and HEL85-96 are Type II cryptic epitopes.

The differences between Type I and Type II cryptic epitopes need to be considered when investigating the use of cryptic epitopes as vaccines against pathogens or tumour antigens.11,12,33 Immunization with a Type I cryptic epitope may be able to activate specific T cells in vivo. However, unless some way can be found to intentionally modify the host APCs to efficiently present the epitope, the Type I cryptic epitope will not be presented at the site of the tumour or infection, and the immune response will not be self-perpetuating. On the other hand, immunization with Type II cryptic peptides may be able to activate low-frequency T-cell precursors and expand them to the point at which the response becomes dominant and self-sustaining. It should be noted that two cryptic epitopes identified in the Plasmodium chabaudi adami apical membrane antigen which are efficiently processed and presented (Type II), were protective against viral challenge.33 Thus, the behaviour of particular epitopes should be evaluated prior to undertaking these types of experiments.

Type II cryptic epitopes offer an alternate explanation for determinant spreading. Determinant spreading is a phenomenon of autoimmune disease, in which later stages of the disease are characterized by autoreactive T cells specific for cryptic epitopes, which were not present in the early stages.8,41 Sercarz and others have proposed that over the course of chronic disease, changes occur in the APC population that result in changes in antigen-processing mechanisms, or recruitment of APCs with different processing mechanisms, which leads to efficient presentation of cryptic epitopes that were not presented earlier.3,13,14,42 An alternative explanation is that at least some of these epitopes are Type II cryptic epitopes, which are efficiently presented but represented by few specific T-cell precursors. Continued antigenic stimulation during the course of the disease eventually elevates the frequency of these T cells to the point where they participate in the immune response at a detectable level.

Type II cryptic epitopes also create a serious problem in self–non-self discrimination and tolerance. It has been reported that the degree of tolerance to individual epitopes of self proteins varies with the amount of epitope/MHC complexes presented,5,7,14 leading to the ‘cryptic self’ hypothesis, that tolerance is induced to dominant epitopes of self proteins but not to cryptic epitopes.6,10 One consequence of this hypothesis is that immunogenic peptides derived from self-proteins are assumed to correspond to inefficiently processed (Type I) cryptic epitopes.1 However, as with immunization studies, crypticity in self antigens is not always associated with inefficient or absent antigen presentation. There is growing evidence in both autologous and transgenic neo-self antigens that some epitopes that escape tolerance are efficiently processed and presented.4,35,37–39 It appears that these epitopes are analogous to Type II cryptic epitopes, in that efficient presentation fails to lead to tolerance. If Type II cryptic epitopes are as common among self proteins as they are in HEL (four of five cryptic H-2k epitopes), then an understanding of how such epitopes can be efficiently processed and presented without either raising an immune response or inducing tolerance will be critical to an understanding of tolerance and autoimmunity.

Acknowledgments

This work was supported by grants from the American Cancer Society (IM-569) and the National Institutes of Health (Hl-48170) to R.K.B. D.P.O’B. was supported by an NIH Training Grant (T32-AI07285) awarded to the University of Rochester Department of Microbiology and Immunology. T.H.T. is supported by an NSF Predoctoral Fellowship.

Glossary

Abbreviations

APC

antigen presenting cell

HEL

hen-egg lysozyme

Tg

transgenic

NTg

non-transgenic

TCR

T-cell receptor

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