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The Journal of Immunology, 2006, 177: 3865-3873.
Copyright © 2006 by The American Association of Immunologists, Inc.

Epigenetic Regulation during B Cell Differentiation Controls CIITA Promoter Accessibility1

Myesha R. Green, Hyesuk Yoon and Jeremy M. Boss2

Department of Microbiology and Immunology, Emory University School of Medicine, Atlanta, GA 30322


   Abstract
Top
 Abstract
Introduction
 Materials and Methods
 Results
 Discussion
 Disclosures
 References
 
B cell to plasma cell maturation is marked by the loss of MHC class II expression. This loss is due to the silencing of the MHC class II transcriptional coactivator CIITA. In this study, experiments to identify the molecular mechanism responsible for CIITA silencing were conducted. CIITA is expressed from four promoters in humans, of which promoter III (pIII) controls the majority of B cell-mediated expression. Chromatin immunoprecipitation assays were used to establish the histone code for pIII and determine the differences between B cells and plasma cells. Specific histone modifications associated with accessible promoters and transcriptionally active genes were observed at pIII in B cells but not in plasma cells. A reciprocal exchange of histone H3 lysine 9 acetylation to methylation was also observed between B cells and plasma cells. The lack of histone acetylation correlated with an absence of transcription factor binding to pIII, particularly that of Sp1, PU.1, CREB, and E47. Intriguingly, changes in chromatin architecture of the 13-kb region encompassing all CIITA promoters showed a remarkable deficit in histone H3 and H4 acetylation in plasma cells, suggesting that the mechanism of silencing is global. When primary B cells were differentiated ex vivo, most of the histone marks associated with pIII activation and expression were lost within 24 h. The results demonstrate that CIITA silencing occurs by controlling chromatin accessibility through a multistep mechanism that includes the loss of histone acetylation and transcription factor binding, and the acquisition of repressive histone methylation marks.


   Introduction
Top
 Abstract
 Introduction
Materials and Methods
 Results
 Discussion
 Disclosures
 References
 
The MHC class II (MHC-II)3 molecule presents exogenously derived Ag to CD4+ T cells to initiate and regulate adaptive immune responses (1, 2, 3). MHC-II molecules are cell surface glycoproteins constitutively expressed on the surface of professional APCs, including macrophages, dendritic cells (4, 5), thymic epithelia, and B cells (6). By exposure to the immune cytokine IFN-{gamma}, MHC-II expression can also be induced on most non-APCs, such as fibroblasts (7, 8). Expression of MHC-II genes is transcriptionally regulated by a group of transcription factors that bind a conserved regulatory region located 150 bp upstream of the transcription start site. These factors, including regulatory factor X (RFX) (9, 10, 11, 12), CREB (13, 14), and NF-Y (15) are required for MHC-II gene expression. Although the binding of these factors to the MHC-II promoters is necessary, it is not sufficient for expression. CIITA, a non-DNA-binding transcriptional coactivator, is required for class II expression (16, 17). CIITA interacts directly with NF-Y, CREB, and RFX, while recruiting histone acetyltransferases such as CBP, as well as other basal transcriptional machinery to the promoters of MHC-II genes (reviewed in Refs. 18, 19, 20).

CIITA is transcriptionally regulated by four separate promoters (pI, pII, pIII, and pIV), each transcribing a unique first exon that will ultimately encode distinct CIITA isoforms (21). Differential expression of the various CIITA isoforms contributes to CIITA’s ability to regulate MHC-II expression in a cell type and developmentally specific manner. Promoter pI drives CIITA expression in splenic dendritic cells and macrophages (22, 23). Promoter pII is expressed at very low levels in humans, and little is known about its biological significance. Mice, however, lack pII. Promoter pIV controls IFN-{gamma}-inducible CIITA expression in most cell types (21, 24, 25). Finally, pIII primarily confers developmental and constitutive CIITA expression in B cells (26), as well as human CD4+ T cells (27, 28), monocytes, and plasmacytoid dendritic cells (29). Low levels of CIITA transcripts from pIV can also be detected in B cells (21, 24, 30).

MHC-II expression is developmentally regulated in B cells (31, 32) and can be observed as early as the pre-B cell stage (33, 34, 35). MHC-II expression increases upon B cell maturation and is constitutively expressed in mature B cells. During the B cell to plasma cell transition both MHC-II and CIITA gene expression are lost (6, 36, 37, 38). Transfection of CIITA into plasma cells can restore MHC-II expression (37), suggesting that silencing of MHC-II is solely due to the silencing of CIITA. Because B cell-specific expression is controlled by pIII, it is the regulation of expression at this promoter that is responsible for MHC-II silencing during this transition.

CIITA pIII is regulated by a 5' promoter proximal regulatory region extending –322 bp upstream to + 124 bp downstream of the transcription start site (21, 26). In vivo genomic footprinting analyses identified several cis-regulatory elements termed site A, site B, site C, and activation response elements (ARE)1 and ARE2 (39). These elements were occupied in B cells but not in plasma cells, suggesting either the loss of the factors that interact with these elements or a loss in accessibility of this region to factor binding. Reporter assays and mutational studies indicated that ARE1 and ARE2 are critical for pIII transcriptional activation in both B cells and human T cells (39). An additional 5'UTR region, as well an E box upstream of site C, are required for maximal transcriptional activity in B cells. The CREB protein has been shown to interact with ARE2 and the 5'UTR of pIII. In addition, PU.1, IRF4, and E47 were shown to interact with site C, as well as two E box elements to potentially confer B cell-specific expression of pIII (40). Although it is suggested that ARE1 binds a TEF-2-like factor in vitro, in vivo evidence for such factors has not been presented.

Originally identified as a B lymphocyte-induced maturation protein, Blimp1 (41) has been shown to function as a transcriptional repressor that controls in part the developmental fate of plasma cells (41, 42). The human ortholog PRD1-BF1 was originally identified due to its ability to bind and repress the IFN-beta promoter through the PRD1 site (43). When ectopically expressed, Blimp1 can silence CIITA expression and was found associated with pIII DNA (44, 45). Blimp1 was also found to interact with members of the GROUCHO family of corepressors (46), as well as with G9a (47, 48). G9a catalyzes the dimethylation of histone H3 lysine 9 (49, 50), a mark associated with transcriptional silencing (reviewed in Ref. 51). Thus, one potential mechanism by which CIITA expression could be silenced during the transition between B cell to plasma cell is through the loss of epigenetic histone modifications associated with gene activation and the gain of silencing marks. Such changes would be predicted to have an impact on the accessibility of the pIII region to its cognate transcription factors. Chromatin remodeling has been previously shown to play an important role in CIITA expression during dendritic cell maturation (22) and may therefore play a role during B cell differentiation.

To investigate whether the above predictions with respect to chromatin accessibility and histone modification changes occur during the B cell to plasma cell transition, the histone code associated with CIITA promoters was examined in model human and murine cell lines. The results showed that in contrast to B cells, plasma cells displayed a general absence of gene activation marks across the entire CIITA upstream region encompassing all four human promoters. At the lymphocyte-specific promoter pIII, H3 lysine 9 acetylation, which was present in B cells, was replaced with a dimethyl mark in plasma cells, indicating a multistep mechanism of gene silencing. Chromatin inaccessibility prevented the binding of specific transcription factors to pIII in plasma cells, despite the fact that the factors were present at the same levels in both cell types. A time course of ex vivo B cell differentiation showed that the transcriptionally active histone marks are lost within 24 h of the process, indicating a rapid mechanism by which CIITA expression is silenced. These data provide evidence that the silencing of CIITA during this transition encompasses a mechanism that orchestrates the transition from an active chromatin state to a repressed and inaccessible state.


   Materials and Methods
Top
 Abstract
 Introduction
 Materials and Methods
Results
 Discussion
 Disclosures
 References
 
Cell culture

Raji B cells (CCL-86; American Type Culture Collection (ATCC)), a Burkett’s lymphoma-derived cell line, were grown in RPMI 1640 medium (Mediatech) supplemented with 5% FBS (HyClone Laboratories), 5% bovine calf serum (HyClone Laboratories), 100 U/ml penicillin, 100 U/ml streptomycin, and 0.29 mg/ml L-glutamine (Invitrogen Life Technologies). NCI-H929 plasma cells (CRL-9068; ATCC), a plasmacytoma-derived cell line, were grown in RPMI 1640 medium with 2 mM L-glutamine, 1.5 g/L sodium bicarbonate, 4.5 g/L glucose, 10 mM HEPES, 1.0 mM sodium pyruvate (Mediatech), and 0.05 mM 2-ME (Sigma-Aldrich). BCL1 clone 3B3 murine B cells (CRL-1669; ATCC) cells were grown in RPMI 1640 medium with 2 mM L-glutamine and 0.05 mM 2-ME and 10% heat-inactivated FBS. P3X63Ag8.653 (P3XAG, clone CRL-1580; ATCC) murine plasmacytoma cells and primary B cells were grown in RPMI 1640 medium with 2 mM L-glutamine, 1.5 g/L sodium bicarbonate, 4.5 g/L glucose, 10 mM HEPES, 1.0 mM sodium pyruvate, 0.05 mM 2-ME, and 10% heat-inactivated FBS. Primary splenocytes were obtained from 5- to 6-wk-old C57BL/6 mice. The use of animals for these experiments was approved by Emory University Institutional Animal Care and Use Committee. B lymphocytes were isolated by CD43 depletion using MACS beads (Miltenyi Biotec). Cells were treated for 5 days with 20 ng/ML IL-2 (Sigma-Aldrich or PeproTech), 10 ng/ML IL-5 (Sigma-Aldrich), and 20 µg LPS (Sigma-Aldrich) in RPMI 1640 medium with 2 mM L-glutamine, 1.5 g/L sodium bicarbonate, 4.5 g/L glucose, 10 mM HEPES, 1.0 mM sodium pyruvate, and 0.05 mM 2-ME.

Abs to transcription factors

Abs to modified histone residues, as well as the Ab to Sp1, were purchased from Upstate Biotechnology. PU.1, IRF4, and E47 Abs were purchased from Santa Cruz Biotechnology. The rabbit Ab against CREB was previously described (14).

Chromatin immunoprecipitation (ChIP) assays

ChIP assays were preformed as previously described (52). One-tenth of the chromatin isolated from 4 x 107 cells was used for each immunoprecipitation. The chromatin was precleared with protein A beads to avoid nonspecific binding. Immunoprecipitation of specific chromatin was performed overnight with 5 µg of each indicated Ab. The amount of immunoprecipitated ChIP DNA was determined by quantitative real-time PCR through comparison to a five point genomic DNA standard curve established during each experiment for each primer pair. To average samples between experiments and apply statistics, the ChIP immunoprecipitated DNA was normalized to 1/30 of the amount of input chromatin used in each assay. The data were then plotted as fold over the irrelevant Ab. In the transcription factor binding experiments, pIV primers were used as a negative control. In the histone acetylation studies, HLA-DRA promoter primers (52) served as a positive control for acetylation, whereas HLA-DRA promoter primers for the MCP1 gene (53), which is not expressed, were used as a negative control. Each ChIP analysis was repeated a minimum of three times from independent chromatin preparations. The Student t test was used to determine whether the obtained values were statistically significant. The primer sequences include: RealCIITApIII, 5'-TCACCAAATTCAGTCCACAGTAAGG and 3'-GCCCCAAGCGGTCAGATTTC; RealtimepONE, 5'-GGTTCCCTCGTCCTGGTTTTCAC and 3'-AGTTGTTCATGGCAGCCCTTGG; RealtimepTWO, 5'-CCGCCTCAGTTTCCCCATCTATAAAGTG and 3'-TGCCTCAGACGCCAGCCTCAAG; and RealtimepFOUR, 5'-CACTGTGAGGAACCGACTGGAG and 3'-TGGAGCAACCAAGCACCTACTG.

Real-time RT-PCR analysis

RNA was prepared by using TRIzol Reagent (Invitrogen Life Technologies), and reverse transcription was conducted using the GeneAmp RNA PCR kit (Applied Biosystems) according to the manufacturer’s instructions. There was 2 µg of RNA used per sample. Each reaction contained a parallel control with no reverse transcriptase. One-tenth of a reverse transcriptase reaction mixture was used as a template in a real-time PCR. Real-time RT-PCR was performed as previously described (54). For each sample, a parallel PCR with primers for GAPDH transcripts was also performed as an input control. To compare samples, the results for all PCR assays were normalized to results obtained for the corresponding GAPDH RT-PCR assays, providing a relative quantitation value. The sequences of the primers used for real-time RT-PCR were as follows: HGAPDH, 5'-CCATGGGGAAGGTGAAGGTCGGAGTC and 3'-GGTGGTGCAGGCATTGCTGATG; HCIITA, 5'-CTGAAGGATGTGGAAGACCTGGGAAAGC and 3'-GTCCCCGATCTTGTTCTCACTC; HLA-DR, 5'-GAGTTTGATGCTCCAAGCCCTCTCCCA and 3'-CAGAGGCCCCCTGCGTTCTGCTGCAAT; HSyndecan1, 5'-GAGCAGGACTTCACCTTTGA and 3'-TTCGTCCTTCTTCTTCATGC; HPRDM1BF1, 5'-ACTTGCAACAAGGGCTTTAC and 3'-ACCTTGCATGGTATGGTTT; MGAPDH, 5'-CAGTATGACTCCACTCACGGCA and 3'-CCTTCTCCATGGTGGTGAAGAC; MCIITA, 5'-GATGTGGAAGACCTGGATCGT and 3'-AAGAGAAGGCTGGAAGGATCTTT; I-A, 5'-GACGCAGCGCATACGGCTC and 3'-CCGCCGCAGGGAGGTGCT; and MBLIMP1 5'-ATGGAGGACGCTGATATGC and 3'-CCTTACTTACCACGCCAATAAC.

FACS analysis

For analysis of human markers, 1 x 106 cells were stained with anti-HLA-DR or anti-Syndecan-1 PE-conjugated Abs (BD Biosciences) and analyzed on a FACSCalibur apparatus. For analysis of mouse markers, 1 x 106 cells were stained with anti-IA/E or anti-syndecan-1 PE-conjugated Abs (BD Biosciences) and analyzed on a FACSCalibur.

EMSA analysis

DNA protein interactions were analyzed by EMSA according to protocols previously described (13, 55). A DNA probe containing the ARE1 element of CIITA pIII was synthesized: ARE1, 5'-GGGGAGGGCTTAAGGGAGTGTGGTAAAATTAGA-3'. Mutated probes were also used as competitors, including Mut1, 5'-GGGGAAAATTTAAGGGAGTGTGGTAAAATTAGA; Mut2, 5'-GGGGAGGGCTTATAAATGTGTGGTAAAATTAGA; and Mut3 5'-GGGGAGGGCTTAAGGGAGTAAACCAAAATTAGA. Ab supershift assays using anti-Sp1, Sp3, CIITA, AML1, p65 and p50 NF-{kappa}B, I{kappa}B, USF1, Stat1, IRF1, IRF4, CBP, cFos, cJun, AP2, ATF2, CBP, CREB, CREM, NF1, cMyc, cMyb, and RAR{alpha} (Santa Cruz Biotechnology) were incubated with the DNA-binding reaction mixture for 20 min before electrophoresis.


   Results
Top
 Abstract
 Introduction
 Materials and Methods
 Results
Discussion
 Disclosures
 References
 
Sp1 binds the ARE1 region of CIITA pIII in B cells

CIITA and MHC-II expression is regulated by pIII in B lymphocytes (21, 26). In vivo genomic DNA footprinting and reporter assays have previously revealed several DNA-binding elements (Fig. 1A) that were occupied in B cells (our unpublished data and Ref. 28). Two regions, ARE1 and ARE2 were required for transcriptional activation of CIITA pIII (28, 39). Although CREB was identified as an ARE2-binding element (28), the factors interacting with ARE1 were not identified. To better understand the transcriptional regulation of CIITA pIII, it was necessary to first identify factors interacting with this critical region. EMSA to identify putative binding factors for the ARE1 region in vitro was performed using nuclear extracts prepared from the B cell line Raji. A specific protein-DNA complex was resolved with the ARE1 probe (Fig. 1B). EMSA analysis using Abs to various transcription factors identified the zinc finger transcription factor Sp1 as a potential candidate, as the Sp1 Ab supershifted the complex. Although a failure to supershift an EMSA product does not eliminate the possibility that the factor does bind the ARE1, a supershift was not seen by any of the other 16 Abs tested, including Sp3, a protein that shares sequence homology with Sp1. Additional EMSA analysis was conducted using labeled mutant DNA probes to demonstrate DNA binding specificity (Fig. 1, A and B). Of the mutant probes, only Mut2, a DNA probe containing mutations at the 5' end of the ARE1 region, failed to bind the factor. Additional Ab supershifts demonstrated that Sp1 was the factor binding to the probes. These data indicate that Sp1 binds the ARE1 element in vitro.


Figure 1
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  		FIGURE 1. Sp1 binds the ARE1-binding element in pIII. A, A schematic of DNA-binding elements in CIITA pIII, and the sequences of wild-type and mutant ARE1 probes are shown. B, Nuclear extracts from Raji B cells were incubated with wild-type (left) or mutant (right) ARE1 probes in the presence or absence of Abs to the indicated transcription factors. The arrows indicate the position of the main reactive protein/DNA ARE1 complex and the complex supershifted by Ab. Comparison of the mutant probes to wild-type shows that Mut2 is the only probe with reduced activity to the binding of the factors. C, ChIP assays were conducted using control and Sp1 Abs. Real-time PCR was used to quantify the immunoprecipitated DNA. All values obtained using ChIP assays were normalized to input chromatin and then normalized to the irrelevant Ab control. The mean raw value for the irrelevant Ab control was 2.8 ng ± 1.7. The mean of three independent experiments + SEM is shown.

 
To verify these in vitro findings, ChIP assays were conducted using Abs to an irrelevant Ab or Sp1. Real-time PCR analysis of the immunoprecipitated DNA with primers for CIITA pIII DNA indicated a significant level of Sp1 binding to pIII DNA (Fig. 1C). Sp1 binding was not observed at an irrelevant locus control (data not shown). This result indicates that Sp1 not only binds CIITA pIII in vitro, but also in vivo.

CIITA and MHC-II expression in plasma cells

To establish model cell lines and end points to monitor CIITA and MHC-II expression during the differentiation of B cells into plasma cells, the expression of CIITA and MHC-II genes were evaluated in human and murine B and plasma cell lines. Raji and BCL-1 were used as model mature human and murine B cell lines, respectively, and NCI-H929 and P3XAG were examined as human and murine plasma cell lines. Real-time RT-PCR conducted on RNA isolated from these cells showed that both MHC-II gene expression as well as CIITA expression was highly expressed in the B cell lines but silenced in the plasma cell lines (Fig. 2, A and B). CIITA expression in this case was assayed from pIII. Analysis by flow cytometry demonstrated a lack of MHC-II cell surface expression in the H929 plasma cell line as well (Fig. 2C), demonstrating that the cell lines provide a model system to investigate CIITA silencing.


Figure 2
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  		FIGURE 2. CIITA and MHC-II expression in B cells and plasma cells. RNA was isolated from Raji and BCL-1 B cell lines or H929 and P3XAG plasma cell lines. mRNA levels of human or mouse CIITA (A) and MHC-II (B) were quantitated by real-time RT-PCR. The expression of each mRNA level is expressed relative to GAPDH control. C, Raji and H929 cells were incubated with PE-conjugated HLA-DR-specific Abs. Cell surface expression of MHC-II was then analyzed by flow cytometry.

 
B cell factor occupancy is absent at CIITA pIII in plasma cells

In addition to our identification of Sp1, CREB (56), Pu.1, IRF4, and E47 have been shown to interact with CIITA pIII in B cells (40). To determine whether the occupancy of these factors differs between B cells and plasma cells, ChIP assays for the presence of these factors were conducted on CIITA pIII in Raji and H929 cells. Sp1, CREB, PU.1, and E47 but not IRF4 were associated with CIITA pIII DNA by (Fig. 3A). Intriguingly, in H929 plasma cells the binding of Sp1, CREB, and E47 was the same as a nonspecific control Ab, and the binding of PU.1 was reduced by 3- to 4-fold. IRF4 also showed no binding. The failure to assemble at pIII may be due to the loss of expression of one or all of these factors. To determine whether this loss was the case, Western blot analysis for each for these proteins was conducted (Fig. 3B). As shown, CREB, Sp1, E47, and PU.1 were expressed similarly in both Raji and H929 cells. This finding suggests that a more active or alternative epigenetic mechanism may be at play to regulate the occupancy of pIII in plasma cells. These findings are in agreement with previous in vivo genomic footprinting comparisons between B cells and plasma cells (our unpublished observations) and (39).


Figure 3
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  		FIGURE 3. Transcription factor binding is lost in plasma cells, although all requisite factors are present. A, ChIP assays using Abs for Sp1, CREB, E47, PU.1, and IRF4 were used to compare factor binding at pIII in Raji and H929 cells. The DNA recovered from the ChIP assays were normalized to input chromatin and then normalized to an irrelevant Ab control. The mean raw value for the irrelevant Ab control was 2.8 ng ± 1.7. B, Western blot analysis was conducted on equal amounts of Raji or H929 whole cell extract and probed with the indicated antisera.

 
Changes in histone architecture at CIITA pIII

One mechanism that may limit the accessibility of factors to CIITA pIII in plasma cells may be due to changes in the posttranslational modifications of the local histones that package the pIII DNA, as such modifications regulate the accessibility and transcriptional competence of a locus (reviewed in Refs. 57 and 58). To determine the histone code associated with pIII, as well as characterize changes in local chromatin architecture during B cell to plasma cell differentiation, ChIP analysis using Abs to acetylated histones H3 and H4 was performed on the model cell lines. In Raji and BCL-1 B cells, high levels of histone H3 and modest levels of histone H4 acetylation were observed (Fig. 4A). In contrast, these marks were near absent in the human and mouse plasma cell lines. This absence of histone acetylation infers a closed chromatin conformation at pIII, and correlates perfectly with a silencing of CIITA in plasma cells.


Figure 4
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  		FIGURE 4. CIITA pIII histone acetylation is lost in human and mouse plasma cells. A, Comparisons between CIITA promoter II acetylation in Raji B cells and H929 plasma or BCL-1 B cells and P3XAG plasma cells using ChIP for diacetylated H3 and tetra-acetylated H4 modifications. B, ChIP assays were conducted using Abs to acetylated histone H3 lysine 9, lysine 18, and lysine 27 and to dimethylated and trimethylated H3 lysine 4 or to histone H3 lysine 9 dimethylation (C). Real-time PCR was used to quantitate the immunoprecipitated DNA. All values obtained using ChIP assays were normalized to input chromatin and then normalized to the irrelevant Ab control. The mean of four independent experiments is shown with SEM. Differences between dimethyl lysine 9 in B cells and plasma cells were evaluated using the Student t test and were found to be statistically significant (p = 0.0363 and p = 0.0386, respectively). The mean raw values for the irrelevant Ab controls in the human and mouse cells were 2.1 ng ± 0.29 and 4.2 ng ± 1.2, respectively.

 
Although the above ChIP analysis of H3 and H4 provides an overall indication of chromatin accessibility, it does not address the dynamics of specific modifications that may ultimately indicate the factors that are regulating accessibility. To further characterize the pIII chromatin architecture, ChIP was conducted using Abs to specific histone acetylation of H3 lysines 9, 18, and 27, modifications associated with transcriptional activation. B cells displayed a high level of histone H3 acetylation at lysines 9, 18, and 27 (Fig. 4B). In the plasma cell lines, acetylation of lysine 9 and lysine 27 was absent or near absent, and acetylation of lysine 18 was reduced 8- to 10-fold when compared with the B cell lines. The levels of histone H3 dimethyl lysine 4, which indicate a region competent for transcription (59), and trimethyl lysine 4, which indicate a promoter region actively being transcribed (60), were also examined. Both human and murine B cells displayed high levels of H3 lysine 4 dimethylation and trimethylation (Fig. 4B). In the plasma cell lines, the level of H3 lysine 4 dimethylation was between two and four times lower than the comparable B cell line, suggesting that the region was less competent for transcription and less accessible. In addition, trimethylation of lysine 4, a mark for active transcription, was robust in both B cell lines and completely absent in the plasma cell lines. Importantly, both human and murine systems provided near identical results, indicating that the modifications observed and the loss of these modifications likely occur through similar mechanisms.

Histone H3 lysine 9 modification has been suggested to represent a molecular switch between active and silenced chromatin. Although acetylation of this site is an indication of transcriptional activation (58), methylation is indicative of transcriptional silencing (61). As discussed, histone H3 lysine 9 acetylation was robust in the B cell lines and absent in the plasma cell lines. To determine whether the switch of modifications had occurred, ChIP assays for H3 dimethyl lysine 9 were conducted. Histone H3 lysine 9 di-methylation was not observed in Raji or BCL-1 B cells but was clearly observed in both plasma cell lines (Fig. 4C), indicating that there is an active mechanism to repress/silence CIITA expression.

Global changes in CIITA promoter histone architecture

The silencing of CIITA pIII as indicated by the loss of activating and gain of silencing epigenetic chromatin marks may not be limited to just pIII and may be more widespread and include the other CIITA promoter regions. To test this hypothesis, the nucleosome modifications at the other CIITA promoters, which span 13 kb of CIITA 5' DNA, were examined by ChIP (Fig. 5). Each promoter region displayed a significant level of acetylation of H3 lysine 9, lysine 18, and lysine 27 in Raji cells, albeit less than that seen at the constitutively active pIII. In H929, this acetylation was completely absent at pI and nearly absent at pIV. A residual level of lysine 18 acetylation was observed at pII. The levels of H3 lysine 4 dimethylation were also high at all three promoters in Raji cells and greatly reduced in the H929 plasma cells. In Raji cells, only pIV displayed trimethylation of H3 lysine 4, a marker for transcriptional activation. This result correlates with the fact that low levels of CIITA transcripts have been observed from pIV in B cells (30, 62). H3 lysine 4 trimethylation was absent in H929 cells. These data suggest that not only is the lymphocyte-specific promoter silenced, but the chromatin in the entire region is deacetylated and likely to be silenced in plasma cells.


Figure 5
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  		FIGURE 5. All CIITA promoters display low acetylation transcriptional competence in plasma cells. The histone modification profiles of CIITA pI, pII, and pIV were evaluated in Raji B cells and H929 plasma cells using ChIP-coupled real-time PCR with the indicated Abs. The mean of three independent experiments is shown with SEM. The mean raw value for the irrelevant Ab control was 2.2 ng ± 0.8.

 
CIITA pIII regulation in primary differentiated cells

Although model cell lines are invaluable tools for understanding molecular mechanisms, their immortalization may prevent an accurate representation of the primary cell situation. CIITA silencing in the context of a primary cell system was therefore examined. CD43– splenic B cells were isolated from C57BL/6 mice. The cells were stimulated with IL-2, IL-5, and LPS to induce plasma cell differentiation. After 5 days of stimulation, mRNA levels of both CIITA and class II molecules were greatly reduced (Fig. 6A). In addition, CIITA protein expression was lost (Fig. 6B) and cell surface MHC-II expression was reduced significantly (data not shown). Increased mRNA levels of the plasma cell markers Blimp1, XBP1, and syndecan 1, as well as decreased levels of B cell-specific factors, Bcl-6 and Pax5 were also observed, verifying the plasma cell phenotype (data not shown).


Figure 6
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  		FIGURE 6. Primary B cell differentiation induces histone deacetylation and loss of methylation accessibility marks at CIITA pIII. Primary B cells isolated from ~5-wk-old mouse spleens were treated with IL-2, IL5, and LPS for 0, 12, or 24 h or 5 days as indicated. A, RNA was isolated from 0-day and 5-day treated cells and levels of CIITA and I-A{alpha} mRNA were quantitated by real-time RT-PCR. The expression of each mRNA level is expressed relative to GAPDH mRNA level control. B, Western blot analysis for CIITA was conducted on whole cell lysates isolated from 0-day and 5-day treated cells. C, ChIP assays using Abs for Sp1, CREB, E47, Pu.1, and IRF4 were used to compare factor binding at pIII in 0-day and 5-day treated cells. The mean raw value for the irrelevant Ab control was 3.6 ± 1.7 ng. D and E, Cells treated with IL-2, IL5, and LPS for the designated time were subjected to ChIP real-time PCR analysis using the indicated Abs and were processed as described. The mean raw values for the irrelevant Ab controls used were 1.2 ng ± 0.6, 2.8 ng ± 1.9, 1.5 ng ± 0.6, and 2.8 ng ± 0.6 for the 0, 12, and 24 h and 5-day time course experiment.

 
ChIP was used to analyze factor binding and histone acetylation in these primary differentiated cells. After 5 days, a significant loss in the binding of Sp1, CREB, E47, and PU.1 to pIII DNA was observed (Fig. 6C). Acetylation and methylation were also examined in a time course following stimulation with IL-2, IL-5, and LPS. Whereas unstimulated cells displayed a high level of histone acetylation at pIII, differentiated cells showed marked decreases in histone acetylation over the time course (Fig. 6D). Examination of individual histone modifications showed a rapid decrease in acetylation at 12 h, with histone H3 acetylation of lysine 9 and lysine 27 approaching background levels (Fig. 6E). Histone H3 lysine 18 acetylation, although markedly decreased at 5 days, was delayed. Similarly, H3 lysine 4 dimethylation did not reach a plateau until after 24 h (Fig. 6E). In contrast, H3 lysine 4 trimethylation decreased rapidly. Taken together, these results suggest that histone deacetylation and loss of histone methylation plays a significant role in the silencing of CIITA pIII, not only in human but in mouse as well.


   Discussion
Top
 Abstract
 Introduction
 Materials and Methods
 Results
 Discussion
Disclosures
 References
 
The developmental control of MHC-II expression in the B lymphocyte lineage reflects the biology and function of B cells during the generation of humoral immune responses. MHC-II gene expression and subsequent Ag presentation in B lymphocytes is principally regulated by CIITA driven from pIII. As B cells differentiate into plasma cells where their terminal function is to secrete Ab, Ag presentation and MHC-II expression are no longer necessary. The loss of CIITA pIII expression is marked and controlled by changes in chromatin accessibility and transcriptional competence and can be divided into several stages based on our data.

Mature B cells express high levels of MHC-II and CIITA. PIII is highly acetylated at multiple histone residues, each serving as activation markers and substrates for additional chromatin remodeling enzymes and components of the transcriptional machinery. The chromatin is fully accessible to at least four transcription factors, Sp1, E47, PU.1, and CREB. Moreover histone methylation marks, associated with active transcription, are present. Additionally, low levels of transcripts have been observed from the downstream pIV region (21, 24, 30, 62), demonstrating that this region is also competent for transcription.

In the initial stage of B cell differentiation to plasma cell, histone H3 acetylation marks that are associated directly with transcriptional activation, such as lysine 9 and lysine 27, are lost, as is histone H3 lysine 4 trimethylation, a mark associated with actively transcribed genes. The loss of these marks occurs within the first 12 h of the ex vivo differentiation process. The loss of H3 lysine 9 acetylation can have profound effects as this mark is associated with the recruitment of components of the general transcription machinery to promoters (63). Acetylation marks can be rapidly removed by histone deacetylases (HDACs), suggesting that the first step may be the recruitment of a HDAC. Alternatively, removal of the acetylation marks can be mediated by the replacement of histones (reviewed in Ref. 64). Recently, H3 replacement with variant H3.3 has been shown to be associated with transcriptional activation of previously silenced genes (65, 66). It is possible that an exchange of modified H3 with histone variants (67) is associated with initial steps toward gene silencing. Also occurring at the same time is the loss of histone H3 lysine 4 trimethylation. The mechanism by which histones lose trimethylated lysine modifications is not known, although it is possible again that these histones are replaced/exchanged with unmodified histones (reviewed in Ref. 64).

A second or a parallel event is the loss of acetylation of histone H3 lysine 18 and the decrease of dimethylated lysine 4. Lysine 18 deacetylation displayed slower kinetics than the loss of lysine 9 or lysine 27 acetylation, suggesting that it may be mediated by a different HDAC/complex or that the HDAC responsible has a lower affinity for lysine 18 acetyl residues. The delay in the loss of H3 dimethyl lysine 4 may indicate a stepwise process of lysine 4 dimethylation (i.e., tri- to di- to mono-). H3 dimethyl lysine 4 can be removed by the coREST complex containing the histone lysine-specific demethylase LSD1 (68, 69), suggesting a possible role for this factor in this process. This mark is associated with overall accessibility (59); thus, it is somewhat intriguing that some pIII regions still carry this mark. This finding was also true in the plasma cell lines, which are fully differentiated, possibly indicating some plasticity even at silenced genes. The loss of DNA binding factor occupancy likely occurs at a time point that is concordant with the loss in acetylation and accessibility. This timing naturally represents a significant step as these factors are likely responsible for the recruitment of the histone acetyltransferases to pIII.

The ARE1 binding element within pIII was identified previously as a critical region required for CIITA transcription in both B cells and human T cells (28, 39); however, no factors associated with this element were identified. Sp1 was found to bind to this site. Sp1 is ubiquitously expressed in both B cells and T cells (reviewed in Ref. 70) and belongs to a large family of factors known to bind GC-rich sequences. Sp1 as well as other Sp1 like molecules have been shown to interact with CBP/p300 coactivators (71, 72) as well as TFIID (73). In addition to Sp1, the factors CREB, PU.1, and E47 are also known to recruit CBP/p300 (74, 75, 76), basal transcriptional machinery (77), and RNA polymerase II (78) to promoters. CBP and p300 are responsible for modification of histone H3 lysine 18 in vivo (79). Thus, it is likely that the combination of these DNA binding factors mediate the recruitment of coactivators that are responsible for the histone code associated with the CIITA locus.

The last stage involved in CIITA silencing is the dimethylation of histone H3 lysine 9. Histone H3 lysine 9 dimethylation, a mark of transcriptional silencing, was observed in both human and murine plasma cell lines. Thus, the switch from a transcriptionally active to silent state is revealed by this lysine modification exchange. Despite several attempts, this mark was not observed in the primary differentiated mouse B cells. Several reasons could account for failure to detect this mark. The first is that the 5-day ex vivo culture may not be long enough to allow this complete change in chromatin structure to have occurred. A second possibility is that the ex vivo differentiation process lacks the necessary signaling events required to completely silence CIITA and therefore only shows the initial stages of the process. Because the endpoint stages of in vivo differentiation are revealed by the plasma cell lines, this possibility is favored. The dimethyl lysine 9 mark can be catalyzed by histone methyltransferases G9a, GLP, SUVar39h1, or SUVar9h2 (reviewed in Ref. 80) and can serve as a substrate for the heterochromatin protein-1 (HP1) (81). Minimal binding of HP1 was observed by ChIP in H929 cells (data not shown).

Previous studies have suggested that CIITA silencing is mediated by expression and binding of Blimp1 to pIII in plasma cells (44). Blimp1 is a transcriptional repressor responsible in part for initiating plasma cell differentiation (41, 82). Exogenously expressed tagged Blimp1 in a pre-B cell line or in IgM- and LPS-stimulated splenocytes down-modulated CIITA 4-fold, and was found associated with pIII, indicating a role for this factor in orchestrating CIITA silencing (44, 45). Experiments in which Blimp1 was exogenously expressed by either retrovirus or adenovirus infection in mature human or murine B cells displayed a modest increase in plasma cell markers but failed to ablate CIITA to the levels seen in our system (data not shown). Based on these results, we suggest that Blimp1 may be necessary but not sufficient to completely silence CIITA. It is intriguing that Blimp1 can interact with G9a in vitro (47). This interaction may serve as the mechanism to epigenetically mark CIITA chromatin in the silent state once differentiation is complete.

Epigenetic mechanisms are critical to B cell development. Treatment of cells with HDAC inhibitors (trichostatin and butyrate) alone can induce splenic B cell to plasma cell transition as indicated by increased expression of plasma cell specific genes, such as Blimp1, J chain, and syndecan-1, and CD43, and down-modulation of IgM, Pax5, and c-myc. In these experiments, however, MHC-II cell surface expression was not affected (83), suggesting that simple inhibition of deacetylases is not sufficient for MHC-II expression in splenic B cells. Conversely, trichostatin can induce endogenous CIITA pI and pIII CIITA expression in macrophage and dendritic cells and in plasmacytomas, respectively (data not shown) and (84). It remains unclear whether this re-expression of CIITA is due to specific effects on the HDACs targeted toward the promoters or results from more global effects. CIITA expression from pI in dendritic cells is regulated by epigenetic mechanisms as well (22). Immature dendritic cells express CIITA until exposed to TNF-{alpha}, LPS, or other maturation stimuli. Stimulated dendritic cells down-regulate CIITA mRNA and protein levels. Unlike the B cell to plasma cell transition, analysis of the pI region did not show changes in the in vivo footprint region during this differentiation, but rather a global deacetylation of histones H3 and H4 across the entire 13 kb region encompassing the CIITA promoters (22). In this study, a similar change in global acetylation during the B cell to plasma cell transition was observed. Together, these data suggest that histone deacetylation and global chromatin remodeling of the CIITA upstream regulatory region is a common mechanism for developmental silencing of CIITA among APCs.


   Acknowledgments
 
We thank members of the laboratory and Dr. Paul Wade for their comments on this work.


   Disclosures
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 Abstract
 Introduction
 Materials and Methods
 Results
 Discussion
 Disclosures
References
 
The authors have no financial conflict of interest.


   Footnotes
 
The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked advertisement in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

1 This work was supported by a National Research Service Award F31 GM 20675 (to M.G.) and Grant AI34000 (to J.M.B.) from the National Institutes of Health. Back

2 Address correspondence and reprint requests to Dr. Jeremy M. Boss, Department of Microbiology and Immunology, Room 3131, Rollins Research Center, Emory University School of Medicine, 1510 Clifton Road, Atlanta, GA 30322. E-mail address: boss{at}microbio.emory.edu Back

3 Abbreviations used in this paper: MHC-II, MHC class II; Blimp, B late-induced maturation protein; ChIP, chromatin immunoprecipitation; HDAC, histone deacetylase; RFX, regulatory factor X; ARE, activation response element. Back

Received for publication April 25, 2006. Accepted for publication June 29, 2006.


   References
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 Abstract
 Introduction
 Materials and Methods
 Results
 Discussion
 Disclosures
 References
 

  1. Benacerraf, B.. 1981. Role of MHC gene products in immune regulation. Science 212: 1229-1238. [Free Full Text]
  2. Long, E. O.. 1992. Antigen processing for presentation to CD4+ T cells. New Biol. 4: 274-282. [Medline]
  3. Cresswell, P.. 1994. Assembly, transport, and function of MHC class II molecules. Annu. Rev. Immunol. 12: 259-293. [Medline]
  4. Cella, M., A. Engering, V. Pinet, J. Pieters, A. Lanzavecchia. 1997. Inflammatory stimuli induce accumulation of MHC class II complexes on dendritic cells. Nature 388: 782-787. [Medline]
  5. Pierre, P., S. J. Turley, E. Gatti, M. Hull, J. Meltzer, A. Mirza, K. Inaba, R. M. Steinman, I. Mellman. 1997. Developmental regulation of MHC class II transport in mouse dendritic cells. Nature 388: 787-792. [Medline]
  6. Kara, C. J., L. H. Glimcher. 1993. Developmental and cytokine-mediated regulation of MHC class II gene promoter occupancy in vivo. J. Immunol. 150: 4934-4942. [Abstract]
  7. Steimle, V., C. A. Siegrist, A. Mottet, B. Lisowska-Grospierre, B. Mach. 1994. Regulation of MHC class II expression by interferon-{gamma} mediated by the transactivator gene CIITA. Science 265: 106-109. [Abstract/Free Full Text]
  8. Collins, T., A. J. Korman, C. T. Wake, J. M. Boss, D. J. Kappes, W. Fiers, K. A. Ault, M. A. Gimbrone, Jr, J. L. Strominger, J. S. Pober. 1984. Immune interferon activates multiple class II major histocompatibility complex genes and the associated invariant chain gene in human endothelial cells and dermal fibroblasts. Proc. Natl. Acad. Sci. USA 81: 4917-4921. [Abstract/Free Full Text]
  9. Durand, B., P. Sperisen, P. Emery, E. Barras, M. Zufferey, B. Mach, W. Reith. 1997. RFXAP, a novel subunit of the RFX DNA binding complex is mutated in MHC class II deficiency. EMBO J. 16: 1045-1055. [Medline]
  10. Masternak, K., E. Barras, M. Zufferey, B. Conrad, G. Corthals, R. Aebersold, J. C. Sanchez, D. F. Hochstrasser, B. Mach, W. Reith. 1998. A gene encoding a novel RFX-associated transactivator is mutated in the majority of MHC class II deficiency patients. Nat. Genet. 20: 273-277. [Medline]
  11. Nagarajan, U. M., P. Louis-Plence, A. DeSandro, R. Nilsen, A. Bushey, J. M. Boss. 1999. RFX-B is the gene responsible for the most common cause of the bare lymphocyte syndrome, an MHC class II immunodeficiency. Immunity 10: 153-162. [Medline]
  12. Reith, W., S. Satola, C. H. Sanchez, I. Amaldi, B. Lisowska-Grospierre, C. Griscelli, M. R. Hadam, B. Mach. 1988. Congenital immunodeficiency with a regulatory defect in MHC class II gene expression lacks a specific HLA-DR promoter binding protein, RF-X. Cell 53: 897-906. [Medline]
  13. Hasegawa, S. L., J. M. Boss. 1991. Two B cell factors bind the HLA-DRA X box region and recognize different subsets of HLA class II promoters. Nucleic Acids Res. 19: 6269-6276. [Abstract/Free Full Text]
  14. Moreno, C. S., G. W. Beresford, P. Louis-Plence, A. C. Morris, J. M. Boss. 1999. CREB regulates MHC class II expression in a CIITA-dependent manner. Immunity 10: 143-151. [Medline]
  15. Wright, K. L., B. J. Vilen, Y. Itoh-Lindstrom, T. L. Moore, G. Li, M. Criscitiello, P. Cogswell, J. B. Clarke, J. P. Ting. 1994. CCAAT box binding protein NF-Y facilitates in vivo recruitment of upstream DNA binding transcription factors. EMBO J. 13: 4042-4053. [Medline]
  16. Riley, J. L., S. D. Westerheide, J. A. Price, J. A. Brown, J. M. Boss. 1995. Activation of class II MHC genes requires both the X box region and the class II transactivator (CIITA). Immunity 2: 533-543. [Medline]
  17. Steimle, V., L. A. Otten, M. Zufferey, B. Mach. 1993. Complementation cloning of an MHC class II transactivator mutated in hereditary MHC class II deficiency (or bare lymphocyte syndrome). Cell 75: 135-146. [Medline]
  18. Ting, J. P., J. Trowsdale. 2002. Genetic control of MHC class II expression. Cell 109: (Suppl.):S21-S33. [Medline]
  19. Boss, J. M., P. E. Jensen. 2003. Transcriptional regulation of the MHC class II antigen presentation pathway. Curr. Opin. Immunol. 15: 105-111. [Medline]
  20. LeibundGut-Landmann, S., J. M. Waldburger, M. Krawczyk, L. A. Otten, T. Suter, A. Fontana, H. Acha-Orbea, W. Reith. 2004. Mini-review: specificity and expression of CIITA, the master regulator of MHC class II genes. Eur. J. Immunol. 34: 1513-1525. [Medline]
  21. Muhlethaler-Mottet, A., L. A. Otten, V. Steimle, B. Mach. 1997. Expression of MHC class II molecules in different cellular and functional compartments is controlled by differential usage of multiple promoters of the transactivator CIITA. EMBO J. 16: 2851-2860. [Medline]
  22. Landmann, S., A. Muhlethaler-Mottet, L. Bernasconi, T. Suter, J. M. Waldburger, K. Masternak, J. F. Arrighi, C. Hauser, A. Fontana, W. Reith. 2001. Maturation of dendritic cells is accompanied by rapid transcriptional silencing of class II transactivator (CIITA) expression. J. Exp. Med. 194: 379-391. [Abstract/Free Full Text]
  23. Waldburger, J. M., T. Suter, A. Fontana, H. Acha-Orbea, W. Reith. 2001. Selective abrogation of major histocompatibility complex class II expression on extrahematopoietic cells in mice lacking promoter IV of the class II transactivator gene. J. Exp. Med. 194: 393-406. [Abstract/Free Full Text]
  24. Piskurich, J. F., C. A. Gilbert, B. D. Ashley, M. Zhao, H. Chen, J. Wu, S. C. Bolick, K. L. Wright. 2006. Expression of the MHC class II transactivator (CIITA) type IV promoter in B lymphocytes and regulation by IFN-{gamma}. Mol. Immunol. 43: 519-528. [Medline]
  25. Muhlethaler-Mottet, A., W. Di Berardino, L. A. Otten, B. Mach. 1998. Activation of the MHC class II transactivator CIITA by interferon-{gamma} requires cooperative interaction between Stat1 and USF-1. Immunity 8: 157-166. [Medline]
  26. Lennon, A. M., C. Ottone, G. Rigaud, L. L. Deaven, J. Longmire, M. Fellous, R. Bono, C. Alcaide-Loridan. 1997. Isolation of a B-cell-specific promoter for the human class II transactivator. Immunogenetics 45: 266-273. [Medline]
  27. Wong, A. W., N. Ghosh, K. P. McKinnon, W. Reed, J. F. Piskurich, K. L. Wright, J. P. Ting. 2002. Regulation and specificity of MHC2TA promoter usage in human primary T lymphocytes and cell line. J. Immunol. 169: 3112-3119. [Abstract/Free Full Text]
  28. Holling, T. M., N. van der Stoep, E. Quinten, P. J. van den Elsen. 2002. Activated human T cells accomplish MHC class II expression through T cell-specific occupation of class II transactivator promoter III. J. Immunol. 168: 763-770. [Abstract/Free Full Text]
  29. LeibundGut-Landmann, S., J. M. Waldburger, C. Reis e Sousa, H. Acha-Orbea, W. Reith. 2004. MHC class II expression is differentially regulated in plasmacytoid and conventional dendritic cells. Nat. Immunol. 5: 899-908. [Medline]
  30. Pai, R. K., D. Askew, W. H. Boom, C. V. Harding. 2002. Regulation of class II MHC expression in APCs: roles of types I, III, and IV class II transactivator. J. Immunol. 169: 1326-1333. [Abstract/Free Full Text]
  31. Noelle, R. J., W. A. Kuziel, C. R. Maliszewski, E. McAdams, E. S. Vitetta, P. W. Tucker. 1986. Regulation of the expression of multiple class II genes in murine B cells by B cell stimulatory factor-1 (BSF-1). J. Immunol. 137: 1718-1723. [Abstract]
  32. Dellabona, P., F. Latron, A. Maffei, L. Scarpellino, R. S. Accolla. 1989. Transcriptional control of MHC class II gene expression during differentiation from B cells to plasma cells. J. Immunol. 142: 2902-2910. [Abstract]
  33. Lala, P. K., G. R. Johnson, F. L. Battye, G. J. Nossal. 1979. Maturation of B lymphocytes. I. Concurrent appearance of increasing Ig, Ia, and mitogen responsiveness. J. Immunol. 122: 334-341. [Abstract/Free Full Text]
  34. Kincade, P. W., G. Lee, T. Watanabe, L. Sun, M. P. Scheid. 1981. Antigens displayed on murine B lymphocyte precursors. J. Immunol. 127: 2262-2268. [Abstract]
  35. Miki, N., M. Hatano, K. Wakita, S. Imoto, S. Nishikawa, T. Tokuhisa. 1992. Role of I-A molecules in early stages of B cell maturation. J. Immunol. 149: 801-807. [Abstract]
  36. Sartoris, S., G. Tosi, A. De Lerma Barbaro, T. Cestari, R. S. Accolla. 1996. Active suppression of the class II transactivator-encoding AIR-1 locus is responsible for the lack of major histocompatibility complex class II gene expression observed during differentiation from B cells to plasma cells. Eur. J. Immunol. 26: 2456-2460. [Medline]
  37. Silacci, P., A. Mottet, V. Steimle, W. Reith, B. Mach. 1994. Developmental extinction of major histocompatibility complex class II gene expression in plasmocytes is mediated by silencing of the transactivator gene CIITA. J. Exp. Med. 180: 1329-1336. [Abstract/Free Full Text]
  38. Lennon, A. M., C. Ottone, M. Rosemblatt, M. Fellous, M. R. Bono, C. Alcaide-Loridan. 1998. CIITA B-cell-specific promoter suppression in MHC class II-silenced cell hybrids. Immunogenetics 48: 283-291. [Medline]
  39. Ghosh, N., J. F. Piskurich, G. Wright, K. Hassani, J. P. Ting, K. L. Wright. 1999. A novel element and a TEF-2-like element activate the major histocompatibility complex class II transactivator in B-lymphocytes. J. Biol. Chem. 274: 32342-32350. [Abstract/Free Full Text]
  40. van der Stoep, N., E. Quinten, M. Marcondes Rezende, P. J. van den Elsen. 2004. E47, IRF-4, and PU. 1 synergize to induce B-cell-specific activation of the class II transactivator promoter III (CIITA-PIII). Blood 104: 2849-2857. [Abstract/Free Full Text]
  41. Turner, C. A., Jr, D. H. Mack, M. M. Davis. 1994. Blimp-1, a novel zinc finger-containing protein that can drive the maturation of B lymphocytes into immunoglobulin-secreting cells. Cell 77: 297-306. [Medline]
  42. Shapiro-Shelef, M., K. I. Lin, L. J. McHeyzer-Williams, J. Liao, M. G. McHeyzer-Williams, K. Calame. 2003. Blimp-1 is required for the formation of immunoglobulin secreting plasma cells and pre-plasma memory B cells. Immunity 19: 607-620. [Medline]
  43. Keller, A. D., T. Maniatis. 1991. Identification and characterization of a novel repressor of beta-interferon gene expression. Genes Dev. 5: 868-879. [Abstract/Free Full Text]
  44. Piskurich, J. F., K. I. Lin, Y. Lin, Y. Wang, J. P. Ting, K. Calame. 2000. BLIMP-I mediates extinction of major histocompatibility class II transactivator expression in plasma cells. Nat. Immunol. 1: 526-532. [Medline]
  45. Ghosh, N., I. Gyory, G. Wright, J. Wood, K. L. Wright. 2001. Positive regulatory domain I binding factor 1 silences class II transactivator expression in multiple myeloma cells. J. Biol. Chem. 276: 15264-15268. [Abstract/Free Full Text]
  46. Ren, B., K. J. Chee, T. H. Kim, T. Maniatis. 1999. PRDI-BF1/Blimp-1 repression is mediated by corepressors of the Groucho family of proteins. Genes Dev. 13: 125-137. [Abstract/Free Full Text]
  47. Gyory, I., J. Wu, G. Fejer, E. Seto, K. L. Wright. 2004. PRDI-BF1 recruits the histone H3 methyltransferase G9a in transcriptional silencing. Nat. Immunol. 5: 299-308. [Medline]
  48. Osipovich, O., R. Milley, A. Meade, M. Tachibana, Y. Shinkai, M. S. Krangel, E. M. Oltz. 2004. Targeted inhibition of V(D)J recombination by a histone methyltransferase. Nat. Immunol. 5: 309-316. [Medline]
  49. Tachibana, M., K. Sugimoto, T. Fukushima, Y. Shinkai. 2001. Set domain-containing protein, G9a, is a novel lysine-preferring mammalian histone methyltransferase with hyperactivity and specific selectivity to lysines 9 and 27 of histone H3. J. Biol. Chem. 276: 25309-25317. [Abstract/Free Full Text]
  50. Tachibana, M., K. Sugimoto, M. Nozaki, J. Ueda, T. Ohta, M. Ohki, M. Fukuda, N. Takeda, H. Niida, H. Kato, Y. Shinkai. 2002. G9a histone methyltransferase plays a dominant role in euchromatic histone H3 lysine 9 methylation and is essential for early embryogenesis. Genes Dev. 16: 1779-1791. [Abstract/Free Full Text]
  51. Martin, C., Y. Zhang. 2005. The diverse functions of histone lysine methylation. Nat. Rev. Mol. Cell. Biol. 6: 838-849. [Medline]
  52. Beresford, G. W., J. M. Boss. 2001. CIITA coordinates multiple histone acetylation modifications at the HLA-DRA promoter. Nat. Immunol. 2: 652-657. [Medline]
  53. Boekhoudt, G. H., Z. Guo, G. W. Beresford, J. M. Boss. 2003. Communication between NF-{kappa}B and Sp1 controls histone acetylation within the proximal promoter of the monocyte chemoattractant protein 1 gene. J. Immunol. 170: 4139-4147. [Abstract/Free Full Text]
  54. Gomez, J. A., P. Majumder, U. M. Nagarajan, J. M. Boss. 2005. X box-like sequences in the MHC class II region maintain regulatory function. J. Immunol. 175: 1030-1040. [Abstract/Free Full Text]
  55. Hasegawa, S. L., J. L. Riley, J. H. Sloan, III, J. M. Boss. 1993. Protease treatment of nuclear extracts distinguishes between class II MHC X1 box DNA-binding proteins in wild-type and class II-deficient B cells. J. Immunol. 150: 1781-1793. [Abstract]
  56. van der Stoep, N., E. Quinten, P. J. van den Elsen. 2002. Transcriptional regulation of the MHC class II trans-activator (CIITA) promoter III: identification of a novel regulatory region in the 5'-untranslated region and an important role for cAMP-responsive element binding protein 1 and activating transcription factor-1 in CIITA-promoter III transcriptional activation in B lymphocytes. J. Immunol. 169: 5061-5071. [Abstract/Free Full Text]
  57. Peterson, C. L., M. A. Laniel. 2004. Histones and histone modifications. Curr. Biol. 14: R546-R551. [Medline]
  58. Jenuwein, T., C. D. Allis. 2001. Translating the histone code. Science 293: 1074-1080. [Abstract/Free Full Text]
  59. Santos-Rosa, H., R. Schneider, A. J. Bannister, J. Sherriff, B. E. Bernstein, N. C. Emre, S. L. Schreiber, J. Mellor, T. Kouzarides. 2002. Active genes are tri-methylated at K4 of histone H3. Nature 419: 407-411. [Medline]
  60. Strahl, B. D., R. Ohba, R. G. Cook, C. D. Allis. 1999. Methylation of histone H3 at lysine 4 is highly conserved and correlates with transcriptionally active nuclei in Tetrahymena. Proc. Natl. Acad. Sci. USA 96: 14967-14972. [Abstract/Free Full Text]
  61. Sims, R. J., III, K. Nishioka, D. Reinberg. 2003. Histone lysine methylation: a signature for chromatin function. Trends Genet. 19: 629-639. [Medline]
  62. Fujita, N., D. L. Jaye, C. Geigerman, A. Akyildiz, M. R. Mooney, J. M. Boss, P. A. Wade. 2004. MTA3 and the Mi-2/NuRD complex regulate cell fate during B lymphocyte differentiation. Cell 119: 75-86. [Medline]
  63. Agalioti, T., G. Chen, D. Thanos. 2002. Deciphering the transcriptional histone acetylation code for a human gene. Cell 111: 381-392. [Medline]
  64. Hake, S. B., C. D. Allis. 2006. Histone H3 variants and their potential role in indexing mammalian genomes: the "H3 barcode hypothesis". Proc. Natl. Acad. Sci. USA 103: 6428-6435. [Abstract/Free Full Text]
  65. Ahmad, K., S. Henikoff. 2002. The histone variant H3.3 marks active chromatin by replication-independent nucleosome assembly. Mol. Cell 9: 1191-1200. [Medline]
  66. Janicki, S. M., T. Tsukamoto, S. E. Salghetti, W. P. Tansey, R. Sachidanandam, K. V. Prasanth, T. Ried, Y. Shav-Tal, E. Bertrand, R. H. Singer, D. L. Spector. 2004. From silencing to gene expression: real-time analysis in single cells. Cell 116: 683-698. [Medline]
  67. Hake, S. B., B. A. Garcia, E. M. Duncan, M. Kauer, G. Dellaire, J. Shabanowitz, D. P. Bazett-Jones, C. D. Allis, D. F. Hunt. 2006. Expression patterns and post-translational modifications associated with mammalian histone H3 variants. J. Biol. Chem. 281: 559-568. [Abstract/Free Full Text]
  68. Shi, Y., F. Lan, C. Matson, P. Mulligan, J. R. Whetstine, P. A. Cole, R. A. Casero. 2004. Histone demethylation mediated by the nuclear amine oxidase homolog LSD1. Cell 119: 941-953. [Medline]
  69. Lee, M. G., C. Wynder, N. Cooch, R. Shiekhattar. 2005. An essential role for CoREST in nucleosomal histone 3 lysine 4 demethylation. Nature 437: 432-435. [Medline]
  70. Kaczynski, J., T. Cook, R. Urrutia. 2003. Sp1- and Krüppel-like transcription factors. Genome Biol. 4: 206[Medline]
  71. Chu, B. Y., K. Tran, T. K. Ku, D. L. Crowe. 2005. Regulation of ERK1 gene expression by coactivator proteins. Biochem. J. 392: 589-599. [Medline]
  72. Song, C. Z., K. Keller, K. Murata, H. Asano, G. Stamatoyannopoulos. 2002. Functional interaction between coactivators CBP/p300, PCAF, and transcription factor FKLF2. J. Biol. Chem. 277: 7029-7036. [Abstract/Free Full Text]
  73. Gill, G., E. Pascal, Z. H. Tseng, R. Tjian. 1994. A glutamine-rich hydrophobic patch in transcription factor Sp1 contacts the dTAFII110 component of the Drosophila TFIID complex and mediates transcriptional activation. Proc. Natl. Acad. Sci. USA 91: 192-196. [Abstract/Free Full Text]
  74. Chrivia, J. C., R. P. Kwok, N. Lamb, M. Hagiwara, M. R. Montminy, R. H. Goodman. 1993. Phosphorylated CREB binds specifically to the nuclear protein CBP. Nature 365: 855-859. [Medline]
  75. Yamamoto, H., F. Kihara-Negishi, T. Yamada, Y. Hashimoto, T. Oikawa. 1999. Physical and functional interactions between the transcription factor PU.1 and the coactivator CBP. Oncogene 18: 1495-1501. [Medline]
  76. Qiu, Y., A. Sharma, R. Stein. 1998. p300 mediates transcriptional stimulation by the basic helix-loop-helix activators of the insulin gene. Mol. Cell. Biol. 18: 2957-2964. [Abstract/Free Full Text]
  77. Xing, L., V. K. Gopal, P. G. Quinn. 1995. cAMP response element-binding protein (CREB) interacts with transcription factors IIB and IID. J. Biol. Chem. 270: 17488-17493. [Abstract/Free Full Text]
  78. Felinski, E. A., J. Kim, J. Lu, P. G. Quinn. 2001. Recruitment of an RNA polymerase II complex is mediated by the constitutive activation domain in CREB, independently of CREB phosphorylation. Mol. Cell. Biol. 21: 1001-1010. [Abstract/Free Full Text]
  79. Schiltz, R. L., C. A. Mizzen, A. Vassilev, R. G. Cook, C. D. Allis, Y. Nakatani. 1999. Overlapping but distinct patterns of histone acetylation by the human coactivators p300 and PCAF within nucleosomal substrates. J. Biol. Chem. 274: 1189-1192. [Abstract/Free Full Text]
  80. Zhang, Y., D. Reinberg. 2001. Transcription regulation by histone methylation: interplay between different covalent modifications of the core histone tails. Genes Dev. 15: 2343-2360. [Free Full Text]
  81. Lachner, M., D. O’Carroll, S. Rea, K. Mechtler, T. Jenuwein. 2001. Methylation of histone H3 lysine 9 creates a binding site for HP1 proteins. Nature 410: 116-120. [Medline]
  82. Shaffer, A. L., K. I. Lin, T. C. Kuo, X. Yu, E. M. Hurt, A. Rosenwald, J. M. Giltnane, L. Yang, H. Zhao, K. Calame, L. M. Staudt. 2002. Blimp-1 orchestrates plasma cell differentiation by extinguishing the mature B cell gene expression program. Immunity 17: 51-62. [Medline]
  83. Lee, S. C., A. Bottaro, R. A. Insel. 2003. Activation of terminal B cell differentiation by inhibition of histone deacetylation. Mol. Immunol. 39: 923-932. [Medline]
  84. Chou, S. D., A. N. Khan, W. J. Magner, T. B. Tomasi. 2005. Histone acetylation regulates the cell type specific CIITA promoters, MHC class II expression and antigen presentation in tumor cells. Int. Immunol. 17: 1483-1494. [Abstract/Free Full Text]



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