Systematic Search Fails to Detect Immunogenic MHC Class-I-Restricted Determinants Encoded by Influenza A Virus Noncoding Sequences

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Virology 305, 50–54 (2003) doi:10.1006/viro.2002.1744

Systematic Search Fails to Detect Immunogenic MHC Class-I-Restricted Determinants Encoded by Influenza A Virus Noncoding Sequences Weisan Chen, 1 Jack R. Bennink, 2 and Jonathan W. Yewdell 2 Laboratory of Viral Diseases, National Institute of Allergy and Infectious Diseases, Bethesda, Maryland 20892-0440 Received February 18, 2002; returned to author for revision April 12, 2002; accepted August 23, 2002 It has been demonstrated in a number of systems that CD8 ⫹ T cells (T CD8⫹) can be induced by peptides encoded in alternative reading frames (ARFs) that do not appear to code for bona fide proteins. The biological relevance of ARF peptides remains to be firmly established, however. With this as a goal, we systematically searched for ARF determinants recognized by mouse T CD8⫹ induced by influenza A virus infection. Of 35 candidate ARF peptides that matched H-2 D b, K b, or K d binding motifs, we found that 13 bind to their respective class I molecules at or above the minimal affinity associated with immunogenicity established by past studies. Nine of these peptides were able to induce T CD8⫹ capable of recognizing peptide-coated target cells. Of these, only a lone determinant is antigenic and immunogenic in the context of influenza A virus infections. Ironically, this peptide is derived from a reading frame that encodes a previously unknown influenza virus protein. These findings suggest that alternative reading frames are not a significant source of antigenic peptides in influenza virus infections and raise doubts regarding the general biological significance of ARF determinants. © 2002 Elsevier Science (USA)

proteins. Although mistakenly frame-shifted gene products were not included in the original DRiP hypothesis (which was posited to account the generation of determinants in standard reading frames), they must account for some of the short-lived proteins detected biochemically. Moreover, it has been clearly demonstrated in viral, tumor, and transfection systems that antigenic and immunogenic peptides can be derived from frame-shifted proteins (reviewed in (Mayrand and Green, 1998)). It remains an important question, however, to what extent frame-shifted determinants contribute to T CD8⫹ responses to different antigens in general and viral antigens in particular. The answer to this question likely varies among different antigens depending on intrinsic properties of mRNA sequences, alterations in ribosomal accuracy associated with different virus infections, and events related to the transformation of tumor cells. In the present study we systematically searched for immunogenic determinants encoded by alterative reading frames of the influenza virus A/Puerto Rico/8/34 (PR8).

INTRODUCTION CD8 T cells (T CD8⫹) recognize MHC molecules bound to short peptides, usually composed of 8 to 10 residues (Ro¨tzschke et al., 1990). Peptides are derived largely from a cytosolic pool of substrates principally through the action of proteasomes, though other proteases contribute to the processing of many determinants (Pamer and Cresswell, 1998; Rock and Goldberg, 1999). Most of the antigenic peptides defined in viral, tumor, allogeneic, and minor H systems as well as the cellular peptides biochemically recovered from MHC class I molecules in sufficient quantities for identification are derived from proteins that are apparently stable metabolically (Rammensee et al., 1997; Engelhard, 1994). Recent results suggest that many of these peptides are derived from proteins that never achieve their intended function and are rapidly destroyed by proteasomes following their synthesis (Schubert et al., 2000; Reits et al., 2000). When such short-lived proteins were originally posited (Yewdell et al., 1996), they were termed defective ribosomal products (DRiPs), to emphasize that their rapid destruction could result from any of myriad errors encountered in converting genetic information to functional

RESULTS AND DISCUSSION To identify potential alterative reading frame determinants in the PR8 influenza A virus we used the predictive motifs of Rammensee and colleagues (1997) to identify candidate peptides that bind to H2-D b, -K b, or -K d as described under Materials and Methods. We limited the selection to open reading frames present in the ⫹1 and ⫹2 registers of the eight coding strands that were downstream from a Met that could potentially serve as an initiation codon, regardless of the potential for the up-

1 Current address: Cancer Vaccine, Ludwig Institute for Cancer Research, Austin and Repatriation Medical Centre, Studley Rd., Heidelberg VIC 3084 Australia. 2 To whom correspondence and reprint requests should be addressed at Room 211, Bldg. 4, 4 Center Dr., MSC 0440, NIH, Bethesda, MD 20892-0440. Fax: 301-402-7362. E-mail: [email protected], [email protected].

0042-6822/02 $35.00 © 2002 Elsevier Science (USA) All rights reserved.

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ARF DETERMINANTS IN VIRUS INFECTIONS

stream codon sequences to initiate translation according to the rules of Kozak (1989). Thirty-five peptides meeting these criteria were synthesized and tested for binding to class I molecules and for their abilities to elicit peptideor virus-specific T CD8⫹ responses. Class I binding was determined by the capacity of peptides to prevent the loss of low-temperature stabilized class I molecules on the surface of TAP-deficient cells. Representative binding curves are shown in Fig. 1. The peptide concentration required for protection of half of the induced class I molecules provides a measure of peptide affinity. Ten of the peptides bound to the respective class I molecules with relatively high affinity (K a of ⬎10 7 M ⫺1), and 3 with intermediate (K a between 10 6 and 10 7 M ⫺1) affinity, using the criteria of Sette and colleagues (1994) (summarized in Table 1). Each of the 35 peptides was screened for its ability to elicit peptide-specific T CD8⫹ by immunizing mice with peptide in incomplete Freund’s adjuvant and restimulating splenocytes in vitro with peptide-sensitized APCs or with PR8-infected APCs. Additionally, splenocytes from PR8-immunized mice were restimulated with peptidesensitized APCs. All effectors were tested for their capacity to lyse PR8-infected or peptide-sensitized 51Crlabeled target cells. Representative data are shown in Fig. 2, and results of all the peptides are summarized in Table 1. Nine of thirty-five peptides were able to induce peptide-specific T CD8⫹ upon two in vitro restimulations with peptide-sensitized APCs. All but one of these peptides bound to class I with intermediate or high affinity. The sensitivities of responding T CD8⫹ were determined by the peptide concentration required to sensitize target cells for lysis; 6 of the 9 immunogenic peptides elicited high-affinity T CD8⫹, i.e., T CD8⫹ capable of lysing cells sensitized with less than 10 ⫺11 M peptide (Fig. 2B). The critical part of this experiment was to test the ability of the 9 different peptide-specific T CD8⫹ popula-

FIG. 1. Characterization of peptide binding. Peptide binding to RMA/ S-K d cells was determined by incubating peptides as the concentrations indicated with low temperature-induced RMA/S-K d cells for 60 min. Cells were then shifted to 37°C for 2 h and peptide binding is inferred from protection of K d molecules against unfolding as determined by the binding of a conformationally sensitive mAb detected by flow cytometry. Data shown are for the immunodominant peptide NP 147–155 and three representative peptides.

51 TABLE 1

Summary of Peptide Binding, Immunogenicity, and Antigenicity

Peptide

Sequence

1 (K d)

NP147–155 NP.F2(239) NP.F2(239) NA.F1(246) NA.F2(114) PB1.F2(95)* PB1.F2(702) PB1.F2(448) PB1.F3(28) PB1.F3(331) PB2.F2(519) PB2.F2(445) NP366–374 PB1.F2(513) PA.F2(306) PA.F2(358) HA.F2(168) HA.F1(441) HA.F2(117) NP.F2(241) NS1.F1(41) PB2.F2(450) PA.F2(704) PB1.F2(3) PB2.F2(457) PB2.F2(583) PB1.F2(69)* PB1.F2(101)* HA.F1(498) HA.F2(156) NP.F2(240) NA.F2(301) PA.F3(23) HA.F2(26) PB1.F2(83)* PA.F3(41) NP.F2(386) Total

TYQRTRALV TFSKGNFKLL TFSKGNFKL KFSRSKRGRL MFSLGMQDL VYWKQWLSL IYLKNSSPAV VFNPLTILL NFPLYRRPSL IYDQKSARMV YYCLPRRSV FFKIGELNL ASNENMETM SAWSFPVL KEREYRYM MRRKFQRL GKAVFTEI DFWTFGHI LSRRFHRL SKGNFKLL SLIGFAEI MRKCFFKI MIPGFCLM AGKPFEWM IGELNLSTM KWNLNHFSL LEHRNSTRL LSLRNPILV SVTMNAWKV NQRSNGSML FSKGNFKLL PGLSNRIHL ACGKNNERV RCRHNMYRL QKTMNQVVM QICSNMHSL WRLWNQVHL 35

⫹⫹ ⫺ ⫺ ⫺ ⫺ ⫹⫹ ⫹⫹ ⫹ ⫺ ⫺ ⫹⫹ ⫺

2 (K b)

⫹⫹⫹ ⫺ ⫺ ⫹ ⫺ ⫺ ⫺ ⫹⫹ ⫺ ⫺ ⫺

4

3

3 4 5 (D b) (␣-Peptide) (␣-PR8)

⫹⫹⫹

⫹⫹ ⫺ ⫺ ⫹⫹⫹ ⫺ ⫹ ⫺ ⫹⫹ ⫹⫹⫹ ⫺ ⫹⫹⫹ ⫺ ⫺ 6

High ⫺ ⫺ ⫺ Low High High ⫺ ⫺ ⫺ High ⫺ High High ⫺ ⫺ ⫺ ⫺ ⫺ ⫺ High ⫺ ⫺ ⫺ Low ⫺ ⫺ High ⫺ ⫺ ⫺ Low ⫺ ⫺ ⫺ ⫺ ⫺ 9

⫹⫹⫹ ⫺ ⫺ ⫺ ⫺ ⫺ ⫺ ⫺ ⫺ ⫺ ⫺ ⫺ ⫹⫹⫹ ⫺ ⫺ ⫺ ⫺ ⫺ ⫺ ⫺ ⫺ ⫺ ⫺ ⫺ ⫺ ⫺ ⫺ ⫹⫹⫹ ⫺ ⫺ ⫺ ⫺ ⫺ ⫺ ⫺ ⫺ ⫺ 1

Note. Defined representative immunodominant peptides from viral gene products are in bold. The sequence numbers for each ARF peptide designate the AA number from the first codon of the designated reading frame, F1, F2, and F3. * Indicates peptides derived from the newly identified 11th PR8 gene product. Columns 1–3: Binding to the predicted class I molecules ranged from nondetectable (⫺) to strong (⫹⫹⫹). Column 4: peptide immunogenicity was determined by immunizing mice with synthetic peptides and restimulating in vitro with the same peptide. The affinity of responding T CD8⫹ was estimated by the amount of peptide required for lysis. “High” indicates sensitization was achieved using peptide at less than 10 pM. “Low” indicates that 10 pM or more was required. Column 5: This summarizes the activity of antipeptide T CD8⫹ against PR8-infected cells and the activity of anti-PR8 T CD8⫹ against peptide-sensitized cells.

tions to recognize virus-infected APCs. To maximize the sensitivity of the assay we optimized the infection protocol and confirmed that even low abundant defined determinants (from standard reading frames) were efficiently presented to the appropriate T CD8⫹ (not shown).

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CHEN, BENNINK, AND YEWDELL

FIG. 2. Characterization of peptide immunogenicity and antigenicity. Splenocytes from mice primed with PR8 or peptide were restimulated in vitro for 7 days with either peptide-pulsed APC or PR8-infected APCs. After depleting CD4 ⫹ and B220 ⫹ cells, the remaining cells were tested for their capacity to lyse peptide- or PR8-sensitized cells. Shown in A are results for two representative peptides that bind to K d with high affinity and the positive control NP 147–155 immunodominant peptide. In B, the above-mentioned T CD8⫹ were restimulated one more time with peptide-pulsed APC for 7 days and then tested for their dose-dependent peptide recognition. This enabled detection of high-affinity TCD8⫹ specific for PB1F2(702) that were not detected by a single round of stimulation. Under similar conditions, the control immunodominant peptide did not stimulate detectable specific CTL from naı¨ve splenic cells.

Despite this, only 1 of the 9 T CD8⫹ populations was able to lyse PR8-infected cells (Fig. 3). The peptide that elicited these T CD8⫹ cells (PB1.F2101) was also the lone peptide of all 35 tested to effectively restimulate T CD8⫹ derived from PR8-infected mice in vitro.

FIG. 3. T CD8⫹ specific for PB1F2101 lyse PR8-infected cells. Splenocytes from mice primed with PR8 (A) or peptide PB1.F2(101) (B) were restimulated in vitro for 7 days with peptide-pulsed APC. Peptidespecific T CD8⫹ were then tested for their capacity to lyse peptide- or PR8-sensitized cells.

FIG. 4. Recognition of ARF peptides by polyclonal T CD8⫹. Peptide antigenicity was determined by incubating peptide-sensitized target cells with polyclonal in vitro restimulated anti-PR8 T CD8⫹ from BALB/c mice. In A it is shown that polyclonal T CD8⫹ could lyse cells expressing each of the influenza virus gene products shown (negative control provided by M1). In B are shown responses against standard reading frame determinants from NP. Even peptides at the bottom of the immunodominance hierarchy are recognized under these conditions, demonstrating the sensitivity of the method. In C are shown typical results for ARF peptides.

As an independent measure of the immunogenicity of alterative reading frame (ARF) peptides, we examined the ability of potent in vitro restimulated polyclonal antiPR8 T CD8⫹ to lyse target cells sensitized with the various ARF peptides. As shown in Fig. 4A, T CD8⫹ in the BALB/cderived culture consisted of multispecificities against antigenic determinants derived from 7 PR8 gene products expressed by vaccinia viruses (M1 serves as a negative control). Each of the 3 previously K d-restricted defined determinants in NP (NP 39–47, NP 147–155, NP 218–226) was recognized by these T CD8⫹ (Fig. 4B). None of the 11 K d motif-containing peptides was able to sensitize P815 cells for lysis by these T CD8⫹ (Fig. 4C shows results for 3 of the 4 immunogenic peptides). We performed a similar analysis with B6 polyclonal T CD8⫹ (not shown) that were capable of specifically recognizing cells sensitized with any of the previously defined K b- or D b-restricted peptides. In this case, only the PB1.F2 62–70 peptide described above was capable of sensitizing cells for lysis.

ARF DETERMINANTS IN VIRUS INFECTIONS

PB1.F2101 derives from a suspiciously long open reading frame (87 residues). In fact, this reading frame encodes a bona fide viral protein that results probably from ribosomal scanning past three upstream AUG codons with poor Kozak consensus sequences for initiation (Chen et al., 2001). This peptide corresponds to residues 62–70 in the new protein, tentatively designated PB1-F2 until its functions are more firmly established. Thus, using a tried and true approach for identifying immunogenic determinants, our search for determinants among ARF peptides encoded by PR8 revealed, ironically, only a novel influenza virus gene product. Of course, these findings do not eliminate the possibility that such ARF determinants exist in influenza virus or, for than matter, play an important role in T CD8⫹ responses, even in H-2 b of H-2 d mice. Our search was limited to only a subset of potential ARF peptides; we excluded from analysis peptides with noncanonical class I binding motifs and peptides whose translation does not depend on a proximal Met. In the end, only 6 of 35 potential determinants were capable of eliciting highaffinity T CD8⫹. Similar studies indicate that only ⬃20% of class I binding peptides are generated in sufficient amounts to be immunogenic by the antigen processing machinery acting on standard open reading frames in viral proteins (Yewdell and Bennink, 1999). This fraction might be expected to be higher for ARFs due to their anticipated rapid degradation. On the other hand, this may be more than compensated for by lower levels of expression than typical viral proteins. While the sample size is low, the present findings indicate that ARFs are not a common source of immunogenic peptides in influenza virus infections in mice. With this in mind, it is worth a critical (if brief) reappraisal of the current evidence supporting the biological relevance of determents encoded by nonstandard reading frames. It has been demonstrated that determinants may be derived from surprising locations in cellular genes (Wang et al., 1996), but the underlying mechanisms have not been carefully examined, and it is uncertain whether the expression of these determinants is simply based on the generation of unexpected but otherwise normal mRNA (Le Fur et al., 1997). Using a transfection system, it has been elegantly demonstrated that alternative start codons may result in the generation of unexpected peptides, but the system used probably generates abnormally high levels of peptide expression (Malarkannan et al., 1999). These problems are minimized in viral systems. A highly immunogenic peptide is present in what appears to be an ARF in a mouse retrovirus (Mayrand et al., 1998). This ARF, however, could potentially encode a 193-residue peptide. The odds of randomly encoding such a long open reading frame are quite low, shifting the burden of proof to demonstrating that a viral protein is not produced from such a reading frame. Studies in which

53

defined peptides are placed in artificial ARFs in rVV convincingly demonstrate that such determinants are capable of being generated by VV in vivo (Elliott et al., 1996; Bullock and Eisenlohr, 1996; Hahn et al., 1991). These studies fall a critical step short of demonstrating that such peptides are actually generated from VV proteins. It is plausible, for example, that mechanisms that operate on manipulated genes are not directly applicable to nonmanipulated genes, which must stand the test of evolution in nature. A similar argument holds for otherwise convincing demonstrations of ARFs from transfected genes. Thus, the jury is still out on the biological relevance of ARF peptides, particularly in viral infections. Given the number of viruses and the diversity of molecular mechanisms of gene expression, there ought to be examples of physiological recognition of a true ARF peptide, that is, a peptide from a protein mistakenly expressed with no evolutionary relevance to the transmissibility of the virus. This is particularly true with viruses propagated out of the watchful gaze of T CD8⫹ (like influenza, which is grown in eggs or tissue culture cells). Alas, as yet there is no airtight example of such a thing. MATERIALS AND METHODS Cell culture The TAP2 mutant cell line RMA/S (H-2 b) and its H-2K dtransfectant RMAs-K d were cultured in DMEM containing 10% fetal calf serum, 5 ⫻ 10 ⫺5 M 2-mercaptoethanol, antibiotics, and 2 mM glutamax (Life Technologies, Rockville, MD) (D-10). The thymoma cell line EL-4 (H-2 b) and the mastocytoma cell line P815 (H-2 d) were maintained in RPMI 1640 with the above supplements (RP-10). T CD8⫹ were stimulated and maintained in RP-10 medium supplied with 10 U/ml of recombinant human IL-2 (see below). Monoclonal antibodies Fluorescein-labeled anti-K d (SF1-1.1.1) was purchased from PharMingen (San Diego, CA). Anti-K b (Y3) and anti-D b (22.249) mAbs were used for flow cytometry as hybridoma tissue culture supernatants. Anti-CD4 mAb culture supernatant from hybridoma GK1.4 and unlabeled anti-CD45A/B220 antibody RA3-6B2 (PharMingen) were used to coat M450 Dynal beads (Dynal Biotech, Lake Success, NY) for depleting CD4 ⫹ cells and B220 ⫹ expressing NK cells after the initial T CD8⫹ stimulation. Generation of T CD8⫹ Eight- to ten-week-old female C57BL/J6 (B6, H-2 b) and BALB/c (H-2 d) mice (Taconic, Germantown, NY) were injected ip with 600 hemagglutinating units of influenza A virus Puerto Rico/8/34 (PR8) or sc at the base of the tail with 50 ␮g of peptides emulsified in 25 ␮l of incomplete

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Freund’s adjuvant (IFA). Splenocytes (3 ⫻ 10 7) were cultured in vitro in RP-10 with 10 U/ml of human IL-2 with ⬃1.5 ⫻ 10 5 PR8-infected or peptide-pulsed irradiated syngeneic cell lines (200 Gy). Viable cells were harvested from Ficoll–Hypaque gradients and enriched for T CD8⫹ by depleting B220 ⫹ and CD4 ⫹ cells using mAbcoated M-450 Dynal beads. T CD8⫹ activities were measured via 51Cr release microcytotoxicity assays, as described (Chen et al., 2000). Peptides, binding assays, and flow cytometry Potential immunogenic peptides were chosen based on published motifs (Rammensee et al., 1997): D b all nonamers, Asn 5, Leu/Ile/Met/Val 9; K b, all octamers, Phe/ Try 5, Leu/Ile/Met/Val 9; K d nonamers and decamers, Tyr/ Phe 2, Leu/Ile/Val 9,10. Peptides were synthesized, HPLCpurified, and analyzed by mass spectrometry by, or under the supervision of, the Biologic Resource Branch, NIAID (Rockville, MD). All peptides were of greater than 95% purity. Peptides were dissolved in DMSO (or water for Cys-containing peptides) at 1 mM as stock solutions and maintained at ⫺30°C. Relative peptide binding was assessed by measuring the stabilization of K b or D b molecules on the surface of RMA/S cells or K d on RMA/S-K d cells as described (Chen et al., 1999). FACS data were analyzed using FlowJo software (TreeStar, CA). ACKNOWLEDGMENTS Bethany Buschling provided outstanding technical assistance. W. Chen received support from C. J. Martin Fellowship 967036 from the Australian National Health and Medical Research Council.

REFERENCES Bullock, T. N. J., and Eisenlohr, L. C. (1996). Ribosomal scanning past the primary initiation codon as a mechanism for expression of CTL epitopes encoded in alternative reading frames. J. Exp. Med. 184, 1319–1329. Chen, W., Anton, L. C., Bennink, J. R., and Yewdell, J. W. (2000). Dissecting the multifactorial causes of immunodominance in class I-restricted T cell responses to viruses. Immunity 12, 83–93. Chen, W., Calvo, P. A., Malide, D., Gibbs, J., Schubert, U., Bacik, I., Basta, S., O’Neill, R., Schickli, J., Palese, P., Henklein, P., Bennink, J. R., and Yewdell, J. W. (2001). A novel influenza A virus mitochondrial protein that induces cell death. Nature Med. 7, 1306–1312. Chen, W., Yewdell, J. W., Levine, R. L., and Bennink, J. R. (1999). Modification of cysteine residues in vitro and in vivo affects the immunogenicity and antigenicity of major histocompatibility complex class I-restricted viral determinants. J. Exp. Med. 189, 1757–1764. Elliott, T., Bodmer, H., and Townsend, A. (1996). Recognition of out-offrame major histocompatibility complex class I-restricted epitopes in vivo. Eur. J. Immunol. 26, 1175–1179.

Engelhard, V. H. (1994). Structures of peptides associated with class I and class II MHC molecules. Annu. Rev. Immunol. 12, 181–207. Hahn, Y. S., Braciale, V. L., and Braciale, T. J. (1991). Presentation of viral antigen to class I major histocompatibility complex-restricted cytotoxic T lymphocytes. Recognition of an immunodominant influenza hemagglutinin site by cytotoxic T lymphocytes is independent of the position of the site in the hemagglutinin translation product. J. Exp. Med. 174, 733–736. Kozak, M. (1989). The scanning model for translation: An update. J. Cell. Biol. 108, 229–241. Le Fur, N., Kelsall, S. R., Silvers, W. K., and Mintz, B. (1997). Selective increase in specific alternative splice variants of tyrosinase in murine melanomas: a projected basis for immunotherapy. Proc. Natl. Acad. Sci. USA 94, 5332–5337. Malarkannan, S., Horng, T., Shih, P. P., Schwab, S., and Shastri, N. (1999). Presentation of out-of-frame peptide/MHC class I complexes by a novel translation initiation mechanism. Immunity 10, 681–690. Mayrand, S. M., and Green, W. R. (1998). Non-traditionally derived CTL epitopes: Exceptions that prove the rules? Immunol. Today 19, 551– 556. Mayrand, S. M., Schwarz, D. A., and Green, W. R. (1998). An alternative translational reading frame encodes an immunodominant retroviral CTL determinant expressed by an immunodeficiency-causing retrovirus. J. Immunol. 160, 39–50. Pamer, E., and Cresswell, P. (1998). Mechanisms of MHC class I—restricted antigen processing. Annu. Rev. Immunol. 16, 323–358. Rammensee, H.-G., Bachmann, J., and Stevanovic, S. (1997). “MHC Ligands and Peptide Motifs.” Landes Bioscience, Austin, Texas. Reits, E. A., Vos, J. C., Gromme, M., and Neefjes, J. (2000). The major substrates for TAP in vivo are derived from newly synthesized proteins. Nature 404, 774–778. Rock, K. L., and Goldberg, A. L. (1999). Degradation of cell proteins and the generation of MHC class I-presented peptides. Annu. Rev. Immunol. 17, 739–779. Ro¨tzschke, O., Falk, K., Deres, K., Schild, H., Norda, M., Metzger, J., Jung, G., and Rammensee, H.-G. (1990). Isolation and analysis of naturally processed viral peptides as recognized by cytotoxic T cells. Nature 348, 252–254. Schubert, U., Anton, L. C., Gibbs, J., Norbury, C. C., Yewdell, J. W., and Bennink, J. R. (2000). Rapid degradation of a large fraction of newly synthesized proteins by proteasomes. Nature 404, 770–774. Sette, A., Vitiello, A., Reherman, B., Fowler, P., Nayersina, R., Kast, W. A., Melief, C. J. M., Oseroff, C., Yuan, L., Ruppert, J., Sidney, J., Guercio, M. F. D., Southwood, S., Kubo, R. T., Chesnut, R. W., Grey, H. M., and Chisari, F. V. (1994). The relationship between class I binding affinity and immunologenicity of potential cytotoxic T cell epotopes. J. Immunol. 153, 5586–5592. Wang, R.-F., Parkhurst, M. R., Kawakami, Y., Robbins, P. F., and Rosenberg, S. A. (1996). Utilization of an alternative open reading frame of a normal gene in generating a novel human cancer antigen. J. Exp. Med. 183, 1131–1140. Yewdell, J. W., Anto´n, L. C., and Bennink, J. R. (1996). Defective ribosomal products (DRiPs). A major source of antigenic peptides for MHC class I molecules? J. Immunol. 157, 1823–1826. Yewdell, J. W., and Bennink, J. R. (1999). Immunodominance in major histocompatibility complex class I-restricted T lymphocyte responses. Annu. Rev. Immunol. 17, 51–88.

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