Intracellular antigens must be processed before presentation to CD8+ T cells by major histocompatibility complex (MHC) class I molecules. Using a recombinant vaccinia virus (Vac) to transiently express the Kd molecule, we studied the antigen processing efficiency of 26 different human tumor lines. Three cell lines, all human small cell lung carcinoma, consistently failed to process endogenously synthesized proteins for presentation to Kd-restricted, Vac-specific T cells. Pulse-chase experiments showed that MHC class I molecules were not transported by these cell lines from the endoplasmic reticulum to the cell surface. This finding suggested that peptides were not available for binding to nascent MHC molecules in the endoplasmic reticulum. Northern blot analysis of these cells revealed low to nondetectable levels of mRNAs for MHC-encoded proteasome components LMP-7 and LMP-2, as well as the putative peptide transporters TAP-1 and TAP-2. Treatment of cells with interferon gamma enhanced expression of these mRNAs and reversed the observed functional and biochemical deficits. Our findings suggest that downregulation of antigen processing may be one of the strategies used by tumors to escape immune surveillance. Potential therapeutic applications of these findings include enhancing antigen processing at the level of the transcription of MHC-encoded proteasome and transporter genes.

RMA/S is a mutant cell line with decreased cell surface expression of major histocompatibility complex class I molecules that has been reported to be deficient in presenting endogenously synthesized influenza virus nucleoprotein (NP) to cytotoxic T lymphocytes (CTL). In the present study we show that RMA/S cells can present vesicular stomatitis virus nucleocapsid protein, and, under some conditions, NP, to Kb-and Db-restricted CTL, respectively. Antigen presentation results from processing of cytosolic pools of endogenously synthesized proteins, and not the binding to cell surface class I molecules of antigenic peptides present in the virus inoculum or released from infected cells. Antigen processing of RMA/S differs, however, from processing by wild-type cells in requiring greater amounts of antigen, longer times to assemble or transport class I-peptide complexes, and in being more sensitive to blocking by anti-CD8 antibody. Thus, the antigen processing deficit in RMA/S cells is of a partial rather than absolute nature.

The influenza A virus infection continues to be a threat to the human population. The seasonal variation of the virus and the likelihood of periodical pandemics caused by completely new virus strains make it difficult to produce vaccines that efficiently protect against this infection. Antibodies (Abs) are very important in preventing the infection and in blocking virus propagation once the infection has taken place. However, the precise protection mechanism provided by these Abs still needs to be established. Furthermore, most research has focused on Abs directed to the globular head domain of hemagglutinin (HA). However, other domains of HA (like the stem) and other proteins are also able to elicit protective Ab responses. In this article, we review the current knowledge about the role of both neutralizing and non-neutralizing anti-influenza proteins Abs that play a protective role during infection or vaccination.

The transporter associated with antigen processing (TAP) transports short peptides from the cytosol to the endoplasmic reticulum, where peptides assemble with class I molecules of the major histocompatibility complex. TAP is comprised of two subunits, termed TAP1 and TAP2. We produced recombinant vaccinia viruses that direct synthesis of the TAP subunits, either individually or together. Virus-encoded TAP is rapidly and efficiently assembled (t1/2 of 5 min or less) by cells and does not spontaneously assemble in detergent extracts. By confocal immunofluorescence microscopy, TAP1 when expressed alone or with TAP2 is largely, if not exclusively, localized to the endoplasmic reticulum. Metabolic labeling with [2-3H]mannose demonstrates that TAP1 (but not TAP2) possesses Asn-linked oligosaccharides, but the lack of binding of [35S]methionine-labeled TAP to concanavalin A-agarose suggests that the glycosylated form represents a minor population of TAP1. The two subunits of the assembled complex present in detergent extracts photolabeled equally with 8-azido-[alpha-32P]ATP. Photolabeling of the two subunits was inhibited in parallel by various di- and trinucleotides, suggesting that their nucleotide binding sites function in a highly similar manner. Incubation of detergent extracts at 37 degrees C results in the rapid loss of TAP1 immunoreactivity, indicating either an unusual sensitivity to proteases or an irreversible conformation alteration.

MHC class I-restricted CTL play a central role in the immune response against methylcholanthrene (MCA)-induced sarcomas in mice. We, therefore, hypothesized that MCA-induced tumors may evade immune recognition by failing to present Ag to CD8+ CTL. Of a number of previously described MCA-induced sarcomas, one, MCA 101, fails to induce CTL, is nonimmunogenic, and grows rapidly and lethally in nonimmunosuppressed recipients. To better understand the nonimmunogenicity of MCA 101 we examined its ability to present foreign Ag to CTL. Unlike immunogenic sarcomas, MCA 101 failed to present endogenously synthesized influenza virus Ag to influenza virus-specific CTL. The deficiency in presentation of endogenous Ag by MCA 101 was attributed to a markedly reduced rate of synthesis of class I molecules because up-regulation of class I synthesis by IFN-gamma greatly increased the presentation of influenza A virus Ag. Despite low levels of cell surface class I expression, MCA 101 presented exogenous peptide Ag to anti-influenza CTL with efficiency similar to immunogenic MCA sarcoma cell lines. These findings could not be attributed to deficiencies in class I assembly or transport, as has been suggested by others who have studied mutant cells with defective Ag presentation. Furthermore, our studies suggest that some tumor cells can escape recognition by CTL and subsequent immune eradication by suppressing presentation of endogenous Ag.