Definition of five new simian immunodeficiency virus cytotoxic T-lymphocyte epitopes and their restricting major histocompatibility complex class I molecules: evidence for an influence on disease progression.

Simian immunodeficiency virus (SIV) infection of the rhesus macaque is currently the best animal model for AIDS vaccine development. One limitation of this model, however, has been the small number of cytotoxic T-lymphocyte (CTL) epitopes and restricting major histocompatibility complex (MHC) class I molecules available for investigating virus-specific CTL responses. To identify new MHC class I-restricted CTL epitopes, we infected five members of a family of MHC-defined rhesus macaques intravenously with SIV. Five new CTL epitopes bound by four different MHC class I molecules were defined. These included two Env epitopes bound by Mamu-A*11 and -B*03 and three Nef epitopes bound by Mamu-B*03, -B*04, and -B*17. All four restricting MHC class I molecules were encoded on only two haplotypes (b or c). Interestingly, resistance to disease progression within this family appeared to be associated with the inheritance of one or both of these MHC class I haplotypes. Two individuals that inherited haplotypes b and c separately survived for 299 and 511 days, respectively, while another individual that inherited both haplotypes survived for 889 days. In contrast, two MHC class I-identical individuals that did not inherit either haplotype rapidly progressed to disease (survived <80 days). Since all five offspring were identical at their Mamu-DRB loci, MHC class II differences are unlikely to account for their patterns of disease progression. These results double the number of SIV CTL epitopes defined in rhesus macaques and provide evidence that allelic differences at the MHC class I loci may influence rates of disease progression among AIDS virus-infected individuals.

Virus-specific cytotoxic T-lymphocyte responses select for amino-acid variation in simian immunodeficiency virus Env and Nef.

Cytotoxic T-lymphocyte (CTL) responses to human immunodeficiency virus arise early after infection, but ultimately fail to prevent progression to AIDS. Human immunodeficiency virus may evade the CTL response by accumulating amino-acid replacements within CTL epitopes. We studied 10 CTL epitopes during the course of simian immunodeficiency virus disease progression in three related macaques. All 10 of these CTL epitopes accumulated amino-acid replacements and showed evidence of positive selection by the time the macaques died. Many of the amino-acid replacements in these epitopes reduced or eliminated major histocompatibility complex class I binding and/or CTL recognition. These findings strongly support the CTL 'escape' hypothesis.

Chlamydia trachomatis major outer membrane protein (MOMP) epitopes that activate HLA class II-restricted T cells from infected humans.

We localized peptide epitopes within the Chlamydia trachomatis (Ct) major outer membrane protein (MOMP) that activate human T cells. T-MOMP' cells were prepared by culturing PBL from 38 humans who had Ct infections in the presence of Ct serovar E MOMP. Some epitopes were first localized by quantifying proliferative responses of T-MOMP' cells to overlapping MOMP segments (sixths) that were produced in Escherichia coli. Further localization was achieved by using overlapping synthetic 16-22 mers that spanned stimulatory MOMP sixths as Ags. The APCs used were human B cell lines (LCL) that were matched or mismatched with respect to HLA class II alleles of the T-MOMP' cells. T cell epitopes were detected in 18 Ct serovar E MOMP 16-22 mers and were presented in association with HLA-DR1, -7, -13, -17, HLA-DRw52, HLA-DQ3 and at least two from among HLA-DR4, -8, -11, -14, -18, in probable addition to HLA-DP4. Peptide 249-265 stimulated T-MOMP' cells from 83% of the subjects; studies with overlapping 11-13 mers spanning peptide 249-265 revealed at least six epitopes presented with different HLA-class II allotypes. Diverse T-MOMP' cultures responded to between 2 and 7 MOMP epitopes. All but 1 of the 23 epitopes localized to date are distributed among four MOMP constant segments. T-MOMP' cells from subjects infected with serovars other than E also responded.

DMA and DMB are the only genes in the class II region of the human MHC needed for class II-associated antigen processing.

Previous studies have shown that homozygous mutations between the LMP2 and DNA loci in the human MHC cause class II molecules to be abnormally conformed and unstable in the presence of SDS at low temperature, and impede class II-associated Ag processing and presentation. These abnormalities result from impaired ability to form intracellular class II/peptide complexes that predominate in normal cells. We show in this work that this defect results from deficient expression of either the DMA or the DMB gene. Human B-LCL.174 (DR3) cells, which have a deletion of all known expressible genes in the class II region, express transgene-encoded HLA-DR3, but have the abnormalities. Transfer of cosmid HA14, which contains the DMA and DMB genes, into .174 (DR3) cells restored normal DR3 conformation, stability in 0.4% SDS at 0 degree, and ability to process and present tetanus toxoid, but only when both DMA and DMB mRNAs were present. The requirement for both genetic expressions in engendering normal phenotypes was confirmed by transferring the cloned genes into .174 (DR3) cells separately or together. Because normal phenotypes were fully restored in transferent cells expressing DMA plus DMB, other genes in the approximately 1-mb homozygous class II region deletion in .174 (DR3) cells either do not participate in or are dispensable for apparently normal production of intracellular class II/peptide complexes. The properties of DM-deficient EBV-transformed B lymphoblastoid cell lines (LCLs) suggest ways of identifying humans in whom DM deficiency contributes to congenital immunodeficiency and malignancy.

Dissection of the D-region of the human major histocompatibility complex by means of induced mutations in a lymphoblastoid cell line.

This paper describes part of a mutagenic dissection of the human D-region. Twenty-six human lymphoblastoid cell mutants that had lost expressions of HLA-DR were created with a two-step procedure: (i) A mutant from which one entire haplotype had been physically deleted by gamma-rays was isolated by means of immunoselection against cells expressing a specific HLA-B antigen. (ii) This heterozygous deletion mutant was irradiated with gamma-rays or treated with ICR 191, a frameshift mutagen, and mutants that no longer expressed the remaining DR1 antigen were selected with a monoclonal antibody directed against a monomorphic DR determinant. Monoclonal antibody GENOX 3.53 was used to show that four of the gamma-ray induced DR-null mutants did not express the cis-linked MB1/MT1 locus. Since MB1/MT1 was still expressed in the other 16 gamm-ray induced and 6 ICR 191-induced DR-null mutants, the separate loss of expression of MB1/MT1 and DR1 is strong evidence that the DR1 and MB1/MT1 alloantigens are under separate genetic control in the cells we used. Since DR-null mutants bound SB2-specific monoclonal antibody ILR1, whether or not they expressed MB1/MT1, the results mean that gamma-rays resolved the genetic determinants for DR1, MB1/MT1, and SB2. Additional complexity of determinants encoded by D-region genes is indicated by the following results. The amount of MB1/MT1 antigen that was detected with ELISA tests for binding of GENOX 3.53 antibody to cells varied inversely with the number of expressed copies of DR or of a locus near DR. This could result from an increased amount of MB1/MT1 antigen or from increased binding accessibility of GENOZ 3.53-reactive antigen in DR-null mutants. Monoclonal antibodies CC 11.23 and CC 6.4 displayed patterns of binding to parental and diverse mutant cells that differed from that of GENOX 3.53, suggesting the existence of at least one additional D-region antigen that is neither SB, DR, nor MB/MT.