Expression of MHC II genes

Innate and adaptive immunity are connected via antigen processing and presentation (APP), which results in the presentation of antigenic peptides to T cells in the complex with the major histocompatibility (MHC) determinants. MHC class II (MHC II) determinants present antigens to CD4+ T cells, which are the main regulators of the immune response. Their genes are transcribed from compact promoters that form first the MHC II enhanceosome, which contains DNA-bound activators and then the MHC II transcriptosome with the addition of the class II transactivator (CIITA). CIITA is the master regulator of MHC II transcription. It is expressed constitutively in dendritic cells (DC) and mature B cells and is inducible in most other cell types. Three isoforms of CIITA exist, depending on cell type and inducing signals. CIITA is regulated at the levels of transcription and post-translational modifications, which are still not very clear. Inappropriate immune responses are found in several diseases, including cancer and autoimmunity. Since CIITA regulates the expression of MHC II genes, it is involved directly in the regulation of the immune response. The knowledge of CIITA will facilitate the manipulation of the immune response and might contribute to the treatment of these diseases

Recombinant gamma interferon provokes resistance of human breast cancer cells to spontaneous and IL-2 activated non-MHC restricted cytotoxicity.

Natural and lymphokine activated killer cells (NK and LAK) are believed to play an important role in the control of tumour progression and metastasis. Their specific receptors on tumours cells are still unknown. Several studies suggest that these cells recognise and eliminate abnormal cells with deleted or reduced expression of MHC class I molecules. Previous reports suggest that interferons (IFN), by increasing MHC class I expression on target cells, induce resistance to killing by NK cells. We investigated the role of MHC molecule expression by two human breast cancer cell lines T47D and ZR75-1 in their susceptibility to NK and LAK cells. These two cell lines spontaneously express low levels of HLA class I antigens but no HLA class II molecules. After IFN-gamma treatment they both overexpressed MHC class I and de novo expressed class II molecules as detected by flow cytometry, quantified by a radioimmunoassay and analysed by two-dimensional gel electrophoresis. Opposed to untreated cells these IFN-gamma treated cells were resistant to NK and LAK lysis. Furthermore, preincubation of IFN-gamma treated breast cancer cells with F(ab')2 fragments of monoclonal antibodies to HLA class I and HLA class II molecules was unable to restore lysis. In contrast, several complete monoclonal antibodies including anti-HLA class I and HLA class II induced the lysis of target cells whether or not they had been treated by IFN-gamma. The therapeutic use of monoclonal antibodies directed against antigens expressed on tumour cells (ADCC) in conjunction with interferon therapy should be discussed in lymphokine-based strategies for treatment of cancer patients

Binding and cooperative interactions between two B cell-specific transcriptional coactivators.

International audienceThe class II transactivator (CIITA) and B cell octamer-binding protein 1/octamer-binding factor 1/Oct coactivator from B cells (Bob1/OBF-1/OCA-B) represent two B cell-specific transcriptional coactivators. CIITA and Bob1 interact with proteins that bind to conserved upstream sequences in promoters of class II major histocompatibility genes and octamer-binding transcription factors Oct-1 and Oct-2, respectively. Both CIITA and Bob1 increase the expression from the DRA promoter, which is a prototypic class II promoter. Moreover, in the presence of CIITA, interactions between class II promoters and Bob1 are independent of the octamer-binding site. Using in vivo and in vitro binding assays, we confirm that Bob1 binds to CIITA. Thus, CIITA not only activates the expression of class II genes but recruits another B cell-specific coactivator to increase transcriptional activity of class II promoters in B cells

Experimental ovine toxoplasmosis: influence of the gestational stage on the clinical course, lesion development and parasite distribution

P. 1-14The relation between gestational age and foetal death risk in ovine toxoplasmosis is already known, but the mechanisms involved are not yet clear. In order to study how the stage of gestation influences these mechanisms, pregnant sheep of the same age and genetic background were orally dosed with 50 oocysts of Toxoplasma gondii (M4 isolate) at days 40 (G1), 90 (G2) and 120 (G3) of gestation. In each group, four animals were culled on the second, third and fourth week post infection (pi) in order to evaluate parasite load and distribution, and lesions in target organs. Ewes from G1 showed a longer period of hyperthermia than the other groups. Abortions occurred in all groups. While in G2 they were more frequent during the acute phase of the disease, in G3 they mainly occurred after day 20 pi. After challenge, parasite and lesions in the placentas and foetuses were detected from day 19 pi in G3 while in G2 or G1 they were only detected at day 26 pi. However, after initial detection at day 19 pi, parasite burden, measured through RT-PCR, in placenta or foetus of G3 did not increase significantly and, at in the third week pi it was lower than that measured in foetal liver or placenta from G1 to G3 respectively. These results show that the period of gestation clearly influences the parasite multiplication and development of lesions in the placenta and foetus and, as a consequence, the clinical course in ovine toxoplasmosis.S

Ets-1 activates the DRA promoter in B cells.

The X box in promoters of class II major histocompatibility complex genes plays a crucial role in the B-cell-specific and gamma interferon-inducible expression of these genes. The sequence TTCC is located in the pyrimidine tract which extends 5' to and partially overlaps the X box of the DRA promoter. This sequence resembles the core binding site for the Ets family of DNA-binding proteins. In this study, we demonstrate that mutations within the pyrimidine tract which change the TTCC motif, but do not affect the binding of regulatory factor X to the X box, decrease the activity of the DRA promoter in B cells. Furthermore, using electrophoretic mobility shift assays and cotransfection experiments, we demonstrate that Ets-1, but not Ets-2 or PU.1, functionally interacts with the pyrimidine tract and activates the DRA promoter

The function of the octamer-binding site in the DRA promoter.

International audienceThe octamer binding site, which is located immediately upstream of the poorly conserved DRA TATA sequence, is important for high levels of expression of this human major histocompatibility class II gene in B cells. In this study, we demonstrate that the substitution of the DRA TATA sequence with the TATA box from the adenovirus E1b promoter removes the requirement for the octamer binding site for high levels of expression from the DRA promoter. Since only the TATA box from the E1b but not the DRA promoters binds the TATA binding protein, we conclude that the octamer binding site helps to recruit TBP to the DRA promoter

MHC class II enhanceosome: how is the class II transactivator recruited to DNA-bound activators?

MHC class II (MHCII) determinants play a crucial role in the immune response by presenting antigenic peptides to T cells. Their expression is controlled from compact promoters at the transcriptional level. Pre-assembled regulatory factor X (RFX) and nuclear factor Y (NFY) complexes form a platform on DNA. The class II transactivator (CIITA) can then be recruited through multiple protein\ub1protein interactions. In this report, we de\uaened domains of CIITA that are responsible for its interactions with these DNA-bound factors. Furthermore, using DNA-af\uaenity precipitation, we demonstrated that although CIITA binds at least \uaeve activators, RFX5, RFXAP, RFXANK/B, NFYB and NFYC, its assembly on the promoter requires the addition of nuclear extracts. We conclude that not only does the platform bind DNA via multiple, spatially constrained nteractions, but that it can recruit only modi\uaeed and/or complexed CIITA to MHCII promoters

Major histocompatibility complex class II transcriptional platform: assembly of nuclear factor Y and regulatory factor X (RFX) on DNA requires RFX5 dimers

Major histocompatibility complex class II (MHC-II) genes are regulated in a B-cell-specific and gamma interferon-inducible manner. Conserved upstream sequences (CUS) in their compact promoters bind nuclear factor Y (NFY) and regulatory factor X (RFX) complexes. These DNA-bound proteins form a platform that attracts the class II transactivator, which initiates and elongates MHC-II transcription. In this report, we analyzed the complex assembly of these DNA-bound proteins. First, we found that NFY can interact with RFX in cells. In particular, NFYA and NFYC bound RFXANK/B in vitro. Next, RFX5 formed dimers in vivo and in vitro. Within a leucine-rich stretch N-terminal to the DNA-binding domain in RFX5, the leucine at position 66 was found to be critical for this self-association. Mutant RFX5 proteins that could not form dimers also did not support the formation of higher-order DNA-protein complexes on CUS in vitro or MHC-II transcription in vivo. We conclude that the MHC-II transcriptional platform begins to assemble off CUS and then binds DNA via multiple, spatially constrained interactions. These findings offer one explanation of why in the Bare Lymphocyte Syndrome, which is a congenital severe combined immunodeficiency, MHC-II promoters are bare when any subunit of RFX is mutated or missing

Complex architecture of major histocompatibility complex class II promoters: reiterated motifs and conserved protein-protein interactions.

International audienceThe S box (also known as at the H, W, or Z box) is the 5'-most element of the conserved upstream sequences in promoters of major histocompatibility complex class II genes. It is important for their B-cell-specific and interferon gamma-inducible expression. In this study, we demonstrate that the S box represents a duplication of the downstream X box. First, RFX, which is composed of the RFX5-p36 heterodimer that binds to the X box, also binds to the S box and its 5'-flanking sequence. Second, NF-Y, which binds to the Y box and increases interactions between RFX and the X box, also increases the binding of RFX to the S box. Third, RFXs bound to S and X boxes interact with each other in a spatially constrained manner. Finally, we confirmed these protein-protein and protein-DNA interactions by expressing a hybrid RFX5-VP16 protein in cells. We conclude that RFX binds to S and X boxes and that complex interactions between RFX and NF-Y direct B-cell-specific and interferon gamma-inducible expression or major histocompatibility complex class II genes