The MHC class I genes of the rhesus monkey. Different evolutionary histories of MHC class I and II genes in primates

Homologues of the human HLA-A and -B MHC class I loci have been found in great apes and Old World primates suggesting that these two loci have existed for at least 30 million years. The C locus, however, shows some sequence similarity to the B locus and has been found only in gorillas, chimpanzees, and humans. To determine the age of the MHC class I C locus and to examine the evolution of the A and B loci we have cloned, sequenced, and in vitro translated 16 MHC class I cDNAs from two unrelated rhesus monkeys (Macaca mulatta) using both cDNA library screening and PCR amplification. Analyses of these sequences suggest that the C locus is not present in the rhesus monkey, indicating that this locus may be of recent origin in gorillas, chimpanzees, and humans. The rhesus monkey's complement of MHC class I genes includes the products of at least one expressed A locus and at least two expressed B loci, indicating that a duplication of the B locus has taken place in the lineage leading to these Old World primates. Comparison of rhesus monkey MHC class I cDNAs to their primate counterparts reveals fundamental differences between MHC class I and class II evolution in primates. Although MHC class II allelic lineages are shared between humans and Old World primates, no such trans-species sharing of allelic lineages is seen at the MHC class I loci.     

Mutation of RFXAP, a regulator of MHC class II genes, in primary MHC class II deficiency

BACKGROUND: Major-histocompatibility-complex (MHC) class II deficiency is an autosomal recessive primary immunodeficiency disease in which MHC class II molecules are absent. It is a genetically heterogeneous disease of gene regulation resulting from defects in several transactivating genes that regulate the expression of MHC class II genes. The mutations responsible for MHC class II deficiency are classified according to complementation group (a group in which the phenotype remains uncorrected in pairwise fusions of cells). There are three known complementation groups (A, B, and C). METHODS: To elucidate the genetic defect in patients with MHC class II deficiency that was not classified genetically, we performed direct complementation assays with the three genes known to regulate the expression of MHC class II genes, CIITA, RFX5, and RFXAP, and the relevant mutations were identified in each patient. RESULTS: Mutations in the RFXAP gene were found in three patients from unrelated families, and the resulting defect was classified as belonging to a novel complementation group (D). Transfection with the wild-type RFXAP gene restored the expression of MHC class II molecules in the patients' cells. CONCLUSIONS: Mutations in a novel MHC class II transactivating factor, RFXAP, can cause MHC class II deficiency. These mutations abolish the expression of MHC class II genes and lead to the same clinical picture of immunodeficiency as in patients with mutations in the other two MHC class II regulatory genes.     

Proto-oncogene PML controls genes devoted to MHC class I antigen presentation

Fragments of foreign antigens associated with class I molecules of the major histocompatibility complex (MHC) are presented at the cell surface to elicit an immune response. This presentation requires the coordinated expression of several genes contained in the MHC, including those encoding the MHC class I heavy chain, the proteins LMP-2 and LMP-7, which are involved in the proteasomal degradation of cytosolic antigens into peptide fragments that are destined for association with MHC class I molecules, and TAP-1 and TAP-2, which transport these fragments across the membrane of the endoplasmic reticulum at the start of their journey to the cell surface. In many virus-transformed cell lines and spontaneous tumours, these genes are simultaneously repressed. However, the key factor(s) that are essential for their expression and repression have not been identified. Here we report that the proto-oncogene product PML induces expression of LMP-2, LMP-7, TAP-1 and TAP-2 in an MHC-class I-negative, recurrent tumour, leading to the re-expression of cell-surface MHC in tumours and to rejection of the tumours. PML also regulates MHC expression in untransformed fibroblasts. We conclude that malfunction of PML may enable a tumour to evade the immune defence of its host.     

Role of MHC class I molecules in autoimmune disease

The MHC class I molecules play a pivotal role in triggering cellular immune responses, binding and presenting intracellularly derived peptide antigens. Studies of MHC class I expression revealed a complex regulatory mechanism that integrates tissue-specific and hormonal modulation. Dynamic regulation occurs in the thyroid, in response to hormonal repression by TSH and stimulation by thyroid hormone. This dynamic cycle provides the basis for proposing the model that such regulation is important to maintain tolerance to self-antigens in tissues synthesizing large amounts of secretory proteins. Failure to appropriately regulate class I levels is predicted to result in autoimmunity. In support of this model, we found that class I-deficient mice are resistant to the experimentally induced autoimmune diseases, SLE, and blepharitis. Furthermore, pharmacological treatment with an agent that reduces class I expression also reduces the incidence and severity of both experimental and spontaneous autoimmune SLE.     

Recessive inheritance of obesity in familial non-insulin-dependent diabetes mellitus, and lack of linkage to nine candidate genes

Segregation analysis of body-mass index (BMI) supported recessive inheritance of obesity, in pedigrees ascertained through siblings with non-insulin dependent diabetes mellitus (NIDDM). BMI was estimated as 39 kg/m2 for those subjects homozygous at the inferred locus. Two-locus segregation analysis provided weak support for a second recessive locus, with BMI estimated as 32 kg/m2 for homozygotes. NIDDM prevalence was increased among those subjects presumed to be homozygous at either locus. Using both parametric and nonparametric methods, we found no evidence of linkage of obesity to any of nine candidate genes/regions, including the Prader-Willi chromosomal region (PWS), the human homologue of the mouse agouti gene (ASP), and the genes for leptin (OB), the leptin receptor (OBR/DB), the beta3-adrenergic receptor (ADRB3), lipoprotein lipase (LPL), hepatic lipase (LIPC), glycogen synthase (GYS), and tumor necrosis factor alpha (TNFA).     

Interpreting MHC class I expression and class I/class II reciprocity in the CNS: reconciling divergent findings

MHC-restricted T cells are thought to contribute to clinical demyelination in MS and other circumstances. The step-by-step mechanisms involved and ways of controlling them are still being defined. Identification of the MHC+ cells in the CNS in situ has been controversial. This chapter reviews MHC expression in neural tissue, including normal, pathological, experimental, and developing tissue in situ and isolated cells in vitro. A basic pattern is defined, in which MHC expression is limited to nonneural cells and strongest class I and II expression are on different cell types. Variations from the basic pattern are reviewed. Ways of reconciling divergent findings are discussed, including the use of "mock tissue" to help choose between technical and biological bases for divergent findings, the potential contribution of internal antigen to the in situ staining patterns, and the possibility that class I upregulation is actively suppressed in situ. Functional implications of the observed patterns of MHC expression and ways of confirming the function of each MHC+ cell type in situ are described. It is suggested that modulating MHC expression in different cell types at different times or in different directions might be desirable.     

Immunosuppressive peptides corresponding to MHC class I sequences

The induction of tolerance is a long-standing goal in transplantation. Intact MHC molecules, or fragments of them, are being used to render T cells unresponsive both in vitro and in vivo. Elucidation of the mechanisms underlying the effects of these treatments should aid the design of novel therapies for transplantation.     

Identification of a novel MHC class I gene, Mamu-AG, expressed in the placenta of a primate with an inactivated G locus

Maternal tolerance of the fetal allograft remains poorly understood. In humans, expression of the highly polymorphic classical HLA-A and HLA-B loci is suppressed, while expression of the nonclassical HLA-G locus is up-regulated at the maternal-fetal interface. Like other nonclassical MHC class I molecules, HLA-G exhibits limited diversity, but certain characteristics of HLA-G distinguish it from other nonclassical MHC class I molecules: it has a truncated cytoplasmic domain, it is the product of alternatively spliced mRNAs, and it is expressed primarily in the placenta. We have examined MHC class I expression in the placenta of the rhesus monkey to determine whether this animal is a suitable model in which to study the function of HLA-G. Although the rhesus monkey possesses orthologs of many MHC class I and II loci found in humans, the HLA-G ortholog is a pseudogene in this nonhuman primate species. In this study, we report the identification of a novel nonclassical MHC class I locus expressed in the placenta of the rhesus monkey, Mamu-AG (Macaca mulatta-AG). Although unrelated to HLA-G, Mamu-AG encodes glycoproteins with all of the characteristics of HLA-G. These Mamu-AG glycoproteins are limited in their diversity, possess truncated cytoplasmic domains, are the products of alternatively spliced mRNAs, and their expression is restricted to the placenta. Taken together, these data suggest that convergent evolution may have resulted in the expression of a unique nonclassical MHC class I molecule in the rhesus monkey placenta, and that the common structural features of Mamu-AG and HLA-G may be functionally significant.     

NK cells can recognize different forms of class I MHC

NK recognition and lysis of targets are mediated by activation receptor(s) whose effects may be over-ridden by inhibitory receptors recognizing class I MHC on the target. Incubation of normal lymphoblasts with a peptide that can bind to their class I MHC renders them sensitive to lysis by syngeneic NK cells. By binding to class I MHC, the peptide alters or masks the target structure recognized by an inhibitory NK receptor(s). This target structure is most likely an "empty" dimer of class I heavy chain and beta2m as opposed to a "full" class I trimer formed by binding of specific peptide that is recognized by CTL.     

The making of the dominant MHC class I ligand SYFPEITHI

The proteasome contributes to the generation of most of the peptide ligands of MHC class I molecules. To compare the identity of the peptides generated by the proteasome with those finally presented by MHC class I molecules, we generated a monoclonal antibody recognizing the C-terminal part of the dominant H2-Kd ligand SYFPEITHI derived from the JAK1 tyrosine kinase. Immunoprecipitations of lysates from H2-Kd-expressing or non-expressing cells revealed that only in the presence of H2-Kd SYFPEITHI could be isolated. No longer potential precursor peptide containing SYFPEITHI could be detected. Surprisingly, a peptide lacking the first two amino acids, FPEITHI, was isolated independently of the presence of H2-Kd molecules. The detection of only SYFPEITHI and FPEITHI in cell lysates corresponded with the strong generation of these two peptides in in vitro digests of elongated SYFPEITHI-containing peptides with purified 20S proteasomes. Our results indicate that MHC ligands can be generated directly by the proteasome in vivo and that at least for SYFPEITHI the expression of the corresponding MHC molecule is critical for protection of the ligand in vivo.     

Identification of class I genes in cartilaginous fish, the most ancient group of vertebrates displaying an adaptive immune response

Sharks are members of the most primitive class of vertebrates (Chondrichthyes) shown to have an adaptive immune system. Suprisingly, however, class I genes have not been identified unambiguously in this taxon, and absence of class I loci or a failure to express class I genes might explain some of the relatively "weak" adaptive immune responses documented in cartilaginous fish. We report here the isolation of three unique cDNA clones from two different species of sharks that encode bona fide class I proteins. These clones exhibit different sequence and expression profiles indicating that they are likely to represent both classical and nonclassical class I lineages. In addition, our preliminary analysis suggests that there may be transfer of gene segments among shark class I genes over evolutionary time. The cloning of shark class I genes completes the identification of molecules that define the adaptive immune system (including Ig, TCR, and MHC class II proteins) in this taxon. Thus, simple models invoking a total absence of certain molecular hallmarks of the immune system to account for poor immune responsiveness in cartilaginous fish should be abandoned.     

Production and crystallization of MHC class I B allele single peptide complexes

Major histocompatibility complex class I B alleles, HLA B8, B53 and B3501 have been cloned, expressed, refolded and crystallized in specific complexes with a number of different 8-mer and 9-mer peptides. For some of these crystallization was initiated by cross-seeding between different B allele complexes. All crystallize in the space group P212121, with similar unit cell dimensions of approximately 52 A X 81 A X 112 A, contain one complex per asymmetric unit and diffract to approximately 2.0 A resolution.     

Natural killer cell receptors specific for MHC class I molecules

The aim of this study was to describe serum GH, IGF-I, and IGF binding protein (BP) 3 levels at birth and during the first 2 y of life in intrauterine growth-retarded (IUGR) children and to correlate these hormonal values with auxologic parameters noted during this period to investigate their predictive value on the postnatal growth pattern. Three hundred and seventeen children were included at birth and studied for auxologic and biologic parameters at birth, 3 and 30 d, and 3, 6, 12, 18, and 24 mo of age. At birth, when analyzed according to gestational age, serum GH levels were increased (p = 0.0001) and serum IGF-I and IGFBP3 levels were decreased (p = 0.0001) in IUGR as compared with normal neonates. When two cohorts were established at birth as a function of the ponderal index (PI) (< or = or > 3rd percentile), serum IGF-I and IGFBP3 levels were found to be significantly reduced in the case of low PI. All parameters were within normal limits at 1 mo of age and remained normal thereafter. During the first 3 mo of life, a positive correlation was found between IGF-I increment and weight gain (r = 0.28, p = 0.002). None of the biologic parameters at birth were predictive either of later growth or of short stature at 2 y of age. In conclusion, low serum IGF-I and IGFBP3 levels at birth were related to fetal malnutrition and were not predictive parameters for later growth.     

IFN-gamma treatment increases insulin binding and MHC class I expression in erythroleukemia cells

We have investigated if interferon-gamma (IFN-gamma) treatment of human K562 tumor cells, which upregulates the expression of MHC class I antigens (MHC-I), simultaneously would influence insulin binding. Treatment of K562 cells with recombinant human IFN-gamma for 48 h caused a significant increase of insulin binding at 37 degrees C. Recombinant human tumor necrosis factor-alpha (TNF-alpha) alone had no effect but acted synergistically with IFN-gamma, leading to a two-fold increase of insulin binding. No change in affinity, number of binding sites or cell surface expression of insulin receptors (IR) after IFN-gamma treatment could be detected. The increased insulin binding observed at 37 degrees C was not seen at 4 degrees C, suggesting alteration of insulin internalization. The dose-response curve, as well as the time curve, for the increase in insulin binding after IFN-gamma treatment correlated with enhanced cell surface expression of MHC-I antigens. However, the correlation was not absolute. Our results show that IFN-gamma treatment alone or together with TNF-alpha, can alter the insulin binding to K562 cells without changing the expression or affinity of the IR. This correlates with the effect of IFN-gamma on MHC-I expression. These results support the findings that MHC-I molecules associate and interact with the IR at the cell surface.     

Regulation of MHC class I membrane expression by beta 2-microglobulin

MHC-I binding peptides and beta 2 microglobulin (beta 2-m) can upregulate the MHC-I heavy chain expression on certain peptide transporter mutant cells. We have further studied this with normal cells and non-mutant cell lines. No MHC-I upregulation was seen with normal, resting or activated T cells. On mouse cell lines P815 and B16, both peptides and human beta 2-m gave an additive upregulation response. With the human small cell lung carcinoma H82, an optimal HLA.A2 binding peptide (GILGFVFTL) gave an upregulation response, whereas beta 2-m alone or in combination with this peptide had no effect. However, beta 2-m potentiated the response of H82 cells to a slightly longer peptide. Using mutant RMA-S cells, it was found that both Brefeldin A (BFA) and chloroquine, but not leupeptin, inhibited MHC-I upregulation response to both peptide and beta 2-m. In contrast to chloroquine, BFA also gave a reduction of background membrane MHC-I expression, presumably due to a block in Golgi transport. Human beta 2-m, which binds to RMA-S cells, and which is known to internalize into endosomes, did not reappear on the cell surface. When Db on RMA-S cells was upregulated by human beta 2-m, the sensitivity of these cells to Db restricted CTL cells increased. Even if beta 2-m did not upregulate the overall MHC-I expression on normal cells, it may still quantitatively increase the expression of optimally presented peptides and endosomal recycling many be important in this process.     

Regulation of MHC class I and II gene transcription: differences and similarities

Major histocompatibility complex (MHC) molecules serve as peptide receptors. These peptides are derived from processed cellular or extra-cellular antigens. The MHC gene complex encodes two major classes of molecules, MHC class I and class II, whose function is to present peptides to CD8+ (cytotoxic) and CD4+ (helper) T cells, respectively. The genes encoding both classes of MHC molecules seem to originate from a common ancestral gene. One of the hallmarks of the MHC is its extensive polymorphism which displays locus and allele-specific characteristics among the various MHC class I and class II genes. Because of its central role in immunosurveillance and in various disease states, the MHC is one of the best studied genetic systems. This review addresses several aspects of MHC class I and class II gene regulation in human and in particular, the contribution to the constitutive and cytokine-induced expression of MHC class I and II genes of MHC class-specific regulatory elements and regulatory elements which apparently are shared by the promoters of MHC class I and class II genes.     

Inhibition of VLA-4 and up-regulation of TIMP-1 expression in B16BL6 melanoma cells transfected with MHC class I genes

The effect of MHC class I gene transfection on the metastatic properties of B16BL6 melanoma cells was investigated. BL6-8 melanoma cells transfected with H-2Kb or H-2Kd, but not H-2Dd or H-2Ld, genes showed a dramatic reduction in their ability to generate experimental metastases in immunosuppressed CB6F1 mice. This observation suggested that some changes in the metastatic phenotype may have been induced in the H-2K- transfected melanoma cells. Analyses of adhesive and invasive properties of BL6-8 melanoma cells transfected with H-2 class I genes have been performed. We found that the loss of metastatic properties in the H-2Kb or H-2Kd gene-transfected melanoma cells was associated with reduced adherence to endothelial cells, laminin and collagen IV, decreased ability to form homotypic cell aggregates and with a complete loss of VLA-4 integrin expression. In addition, BL6-8 melanoma cells transfected with H-2K genes demonstrated reduced ability to invade Matrigel that paralleled up-regulation of TIMP-1 expression. Incubation of untransfected BL6-8 clone or B16F1 cells with 5-azacytidine similarly resulted in up-regulation of TIMP-1, suggesting that the changes in methylation of TIMP-1 gene could be responsible for TIMP-1 expression in the H-2K-transfected BL6-8 melanoma cells. Transfection of BL6-8 cells with the H-2Dd/Ld genes did not affect their adhesive and invasive properties. Previously we reported that reduction in the metastatic properties of the H-2Kb transfected cells was associated with alterations in cell surface carbohydrates with appearance of alpha-galactosyl epitopes and reduction in cell surface sialylation. The present data indicate that, in addition to changes in cell surface carbohydrates, reduction in adhesive properties and up-regulation of TIMP-1 may be responsible for the observed loss of metastatic potential of BL6-8 cells transfected with the H-2K genes.