Inherited immunodeficiency with a defect in a major histocompatibility complex class II promoter-binding protein differs in the chromatin structure of the HLA-DRA gene.

A defect in a trans-regulatory factor which controls major histocompatibility complex class II gene expression is responsible for an inherited form of immunodeficiency with a lack of expression of human leukocyte antigen (HLA) class II antigens. We have recently described and cloned an HLA class II promoter DNA-binding protein, RF-X, present in normal B cells and absent in these class II-deficient regulatory mutants. Here we report that these in vitro results correlate with a specific change in the chromatin structure of the class II promoter: two prominent DNase I-hypersensitive sites were identified in the promoter of the HLA-DRA gene in normal B lymphocytes and found to be absent in the class II-deficient mutant cells. The same two prominent DNase I-hypersensitive sites were observed in normal fibroblastic cells induced by gamma interferon to express class II genes. Interestingly, they were also observed in the uninduced class II-negative fibroblastic cells, which have also been shown to have a normal RF-X binding pattern. We conclude that the two DNase I-hypersensitive sites in the HLA-DRA promoter reflect features in chromatin structure which correlate with the binding of the trans-acting factor RF-X and which are necessary but not sufficient for the expression of class II genes.     

Two DNA-binding proteins discriminate between the promoters of different members of the major histocompatibility complex class II multigene family.

The regulation of major histocompatibility complex (MHC) class II gene expression is a key feature of the control of normal and abnormal immune responses. In humans, class II alpha - and beta-chain genes are organized in a multigene family with three distinct subregions, HLA-DR, -DQ, and -DP. The regulation of these genes is generally coordinated, and their promoters contain highly conserved motifs, in particular the X and Y boxes. We have identified five distinct proteins that bind to specific DNA sequences within the first 145 base pairs of the HLA-DR promoter, a segment known to be functionally essential for class II gene regulation. Among these, RF-X is of special interest, since mutants affected in the regulation of MHC class II gene expression have a specific defect in RF-X binding. Unexpectedly, RF-X displays a characteristic gradient of binding affinities for the X boxes of three alpha-chain genes (DRA greater than DPA much greater than DQA). The same observation was made with recombinant RF-X. We also describe a novel factor, NF-S, which bound to the spacer region between the X and Y boxes of class II promoters. NF-S exhibited a reverse gradient of affinity compared with RF-X (DQA greater than DPA much greater than DRA). As expected, RF-X bound well to the mouse IE alpha promoter, while NF-S bound well to IA alpha. The drastic differences in the binding of RF-X and NF-S to different MHC class II promoters contrasts with the coordinate regulation of HLA-DR, -DQ, and -DP genes.     

NF-X, a transcription factor implicated in MHC class II gene regulation

The X box has been shown in several assay systems to be a critical element of MHC class II gene promoters. Several X box-binding activities have been discovered in nuclear extracts from a variety of cell lines. The critical question is: which of these are responsible for mediating X box function? This report provides a further characterization of NF-X, a highly specific X box-binding activity we described previously. The cell-type distribution, structural features, and binding site characteristics of NF-X are analyzed in detail, to facilitate comparison with other reported activities. Most importantly, the functional relevance of NF-X is assessed by scanning mutagenesis, and the results indicate that this complex is indeed involved in regulating MHC class II gene expression. With these data in mind, the relationship between NF-X and RF-X, an X box-binding activity reported to be absent in patients with severe combined immunodeficiency, is discussed.     

The genes for MHC class II regulatory factors RFX1 and RFX2 are located on the short arm of chromosome 19.

RFX1 is a transacting DNA-binding regulatory factor involved in the control of MHC class II gene expression. RFX2 is a structurally very similar protein with identical DNA binding features. A member of the family of RFX factors is affected in an autosomal recessive disease, MHC class II deficient combined immunodeficiency (CID), caused by a defect in a trans-acting regulatory factor controlling MHC class II gene expression. In situ hybridization with 3H-labeled RFX1 cDNA has allowed us to identify two distinct targets on the short arm of chromosome 19 (19p13.1 and 19p13.2-p13.3). With the use of biotinylated genomic cosmid clones specific for RFX1 and RFX2, respectively, it was then possible to localize RFX1 at 19p13.1 and RFX2 at 19p13.2-p13.3. These two regulatory genes are thus assigned to a region of high gene density and RFX1 is close to another DNA-binding factor, LYL1.     

Class I major histocompatibility proteins are an essential component of the simian virus 40 receptor.

The class I molecules encoded by the major histocompatibility complex (MHC) present endogenously synthesized antigenic peptide fragments to cytotoxic T lymphocytes. We show here that these proteins are an essential component of the cell surface receptor for simian virus 40 (SV40). First, SV40 binding to cells can be blocked by two monoclonal antibodies against class I human lymphocyte antigen (HLA) proteins but not by monoclonal antibodies specific for other cell surface proteins. Second, SV40 does not bind to cells of two different human lymphoblastoid cell lines which do not express surface class I MHC proteins because of genetic defects in the beta 2-microglobulin gene in one line and in the HLA complex in the other. Transfection of these cell lines with cloned genes for beta 2-microglobulin and HLA-B8, respectively, restored expression of their surface class I MHC proteins and resulted in concomitant SV40 binding. Finally, SV40 binds to purified HLA proteins in vitro and selectively binds to class I MHC proteins in a cell surface extract.     

Regulation of interlocking gene regulatory network subcircuits by a small molecule inhibitor of retinoblastoma protein (RB) phosphorylation: Cancer cell expression of HLA-DR

The induction of the major histocompatibility (MHC), antigen-presenting class II molecules by interferon-gamma, in solid tumor cells, requires the retinoblastoma tumor suppressor protein (Rb). In the absence of Rb, a repressosome blocks the access of positive-acting, promoter binding proteins to the MHC class II promoter. However, a complete molecular linkage between Rb expression and the disassembly of the MHC class II repressosome has been lacking. By treating A549 lung carcinoma cells with a novel small molecule that prevents phosphorylation-mediated, Rb inactivation, we demonstrate that Rb represses the synthesis of an MHC class II repressosome component, YY1. The reduction in YY1 synthesis correlates with the advent of MHC class II inducibility; with loss of YY1 binding to the promoter of the HLA–DRA gene, the canonical human MHC class II gene; and with increased Rb binding to the YY1 promoter. These results support the concept that the Rb gene regulatory network (GRN) subcircuit that regulates cell proliferation is linked to a GRN subcircuit regulating a tumor cell immune function.     

B-cell factor 1 is required for optimal expression of the DRA promoter in B cells.

The X box in the DRA promoter of the human histocompatibility complex is required for expression of the DRA gene in B cells. We show that a B-cell factor binds to a sequence that is clearly distinguishable from binding sites for the previously described X box binding nuclear proteins RF-X, NF-X, NF-Xc, NF-S, hXBP, and AP-1. Mutations in the DRA X box that disrupt the binding of this factor result in a lower level of gene expression, as does the presence of Id (a trans-dominant regulatory protein that negatively regulates helix-loop-helix proteins). Furthermore, this factor is recognized by antibodies directed against the helix-loop-helix protein A1, a mouse homolog of the immunoglobulin enhancer binding proteins E12/E47, and it binds to sequences in other genes that were previously shown to bind these proteins. By these criteria, this factor is BCF-1.     

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. Received: 12 January 1998