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Departments of
*
Immunology,
Biophysics, and
Pathology, Roswell Park Cancer Institute, Buffalo, NY 14263; and
§
Departments of Medicine and Microbiology, State University of New York, School of Medicine and Biomedical Sciences, Buffalo, NY 14214
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Abstract |
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 Top Abstract IntroductionMaterials and MethodsResultsDiscussionReferences |
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. The magnitude
of induction of these genes cannot be explained by the effect of DAIs
on the cell cycle or enhanced apoptosis. Induction of class II genes by
DAIs was accompanied by activation of a repressed class II
transactivator gene in a plasma cell tumor but, in several other tumor
cell lines, class II was induced in the apparent absence of class II
transactivator transcripts. These findings also suggest that the
abnormalities observed in some tumors in the expression of genes
critical to tumor immunity may result from epigenetic alterations in
chromatin and gene regulation in addition to well-established
mutational mechanisms. |
Introduction |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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. However, a few cell types, including mature
oligodendrocytes, sensory neurons, plasma cells, and trophoblasts,
cannot be induced to express class II by IFN-. MHC class II
expression is tightly regulated primarily at the level of transcription
by multiple transcriptional activators binding to the W/S, X, and Y box
promoter sequences. In addition, coactivators, such as class II
transactivator
(CIITA),3 Bob1, and
CBP/p300, play important roles in the transcriptional regulation of
class II (1, 2, 3). Recently, CIITA has been reported to
facilitate IFN- induction of MHC class I in addition to class II Ags
(4, 5), and this, together with the similarities in the
structure of the class I and II promoters (6), suggests a
commonality in the regulation of MHC genes.
Many tumors are MHC class II negative and, although some can be induced
to express class II by IFN-, a subset of tumors is noninducible by
cytokines. In a few noninducible tumors, defects in IFN- receptors
or Janus kinase-STAT signaling pathway components have been described
and have been associated with tumor progression (7). In
plasmacytoma cell lines, and in selected melanomas and hepatocellular
carcinomas, the lack of expression of all class II isotypes appears to
be due to a defect in the transcription of the CIITA gene. In these
cells, transfection of CIITA results in class II expression (8, 9) and, in certain tumors, this restores their ability to
present intact Ag (9). That MHC class II may play a role
in tumor immunity is suggested by studies demonstrating that
transfection of MHC class II-negative tumor cells with class II genes
inhibits tumorigenicity and elicits immunity to subsequent challenge
with wild-type tumor cells (10). However, induced
expression of MHC class II does not always enhance immunity and, in the
absence of costimulatory factors, may even promote tumor progression by
inducing anergy (11). Thus, differences in the functional
role of class II may be related to the context in which the MHC genes
are expressed in different tumors, i.e., the presence of associated
"escape" defects (reviewed in Refs. 12, 13).
Considerable evidence has accumulated recently that gene expression is influenced by epigenetic mechanisms that alter chromatin structure (14). Acetylation of lysine residues on the N-terminal tails of the basic core histones in nucleosomes is generally associated with enhanced transcription, whereas deacetylation compacts chromatin and represses transcription. Thus, while targeting molecules with histone acetyltransferase activity to promoters results in gene activation, repression of certain genes is accomplished by transcriptional repressors and corepressors that bind and target histone deacetylases (HDACs) (15, 16). Several noncompetitive inhibitors of HDACs have been described, including sodium butyrate (NaBu) and the highly specific inhibitor trichostatin A (TSA), that arrest cells in both the G1 and G2-M phases of the cell cycle (17), influence growth, and induce apoptosis (18). These inhibitors have been shown to activate a number of important genes by altering histone acetylation and chromatin structure, although not all genes are activated by HDAC inhibitors (DAIs), many are unaffected and some are inhibited (19).
We show here that MHC class II mRNA and cell surface protein can be
induced by DAIs in mouse and human tumor cell lines that cannot be
activated to express class II by IFN-. Evidence is presented that,
in some tumors, DAIs activate a repressed CIITA gene, whereas, in
others, induction may occur via a route different from the classical
CIITA-mediated pathway. Importantly, DAIs are also shown to enhance the
constitutive expression of MHC class I, as well as the costimulatory
molecule CD40, on some tumor cells that were not induced by IFN-.
These data also suggest that failure of tumors to express certain
molecules critical to immunity (tumor escape) may result from chromatin
repression, which is reversed by HDAC inhibitors.
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Materials and Methods |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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The human neuroblastoma tumor cell line SK-N-MC (MC) and mouse plasmacytoma J558 cell line were obtained from American Type Culture Collection (Manassas, VA). Mouse adenocarcinoma Colon 26, generously provided by Elizabeth A. Repasky (Roswell Park Cancer Institute, Buffalo, NY), was maintained in DMEM growth media supplemented with 10% FCS (Life Technologies, Grand Island, NY) as a cell line derived from the original transplantable tumor. RJ2.2.5 cells were a generous gift from Cheong-Hee Chang (University of Michigan, Ann Arbor, MI). Stock and working dilutions of TSA (Wako Biochemicals, Osaka, Japan) were prepared using ethanol as a diluent. Abs to murine surface markers were obtained from the following suppliers: 1) PharMingen (San Diego, CA): PE-conjugated anti-H-2Dd (clone 34-2-12), I-Ad (AMS-32.1), CD40 (3/23), CD44 (IM7), anti-CD45R (RA3-6B2), B7 integrin (M293), strepavidin; biotinylated anti-CD25 (7D4), CD29 (HA2/5), CD119 (GR20); purified anti-CD104 (346-11A), and FITC-anti-H-2Dd (34-2-12); 2) Caltag (South San Francisco, CA): PE-conjugated anti-H-2Kk (CTKk), I-Ak (14V.18), CD62L (MEL-14), CD80 (RMMP-1), CD86 (RMMP-2); FITC- and RPE-annexin-V; and 3) Southern Biotechnology Associates (Birmingham, AL): PE-conjugated anti-CD54 (KAT-1), CD154 (MRI). Flow cytometry reagents for human cells, PE-anti HLA-A, B, C (clone G46-2.6) were obtained from PharMingen, PE-anti HLA-DR (clone 243) from Becton Dickinson (Mountain View, CA), and PE-anti CD40 (clone mAb 89) from Immunotech/Coulter (Miami, FL).
Inhibition of HDAC activity
Toxicity at various concentrations and incubation times with TSA
and NaBu were first determined using three techniques: 1) trypan-blue
dye exclusion; 2) apoptosis as measured by DNA laddering on agarose
gels; and 3) flow cytometric analysis with propidium iodide (PI) or
annexin-V. Preliminary titration was performed on each cell line at 5,
10, 25, 50, 100, and 250 nM TSA for 6, 12, 24, 48, and 72 h and
with the MC cell line at 1 mM NaBu for 12, 24, and 48 h, and
concentrations and incubation times were selected to exclude toxicity
and maximize expression. IFN- was titrated at 10, 50, 100, and 500
U/ml for each cell line. In most experiments, 100 U/ml was
employed.
RT and quantitative real-time PCR
Standard RT-PCR was performed as previously described
using primers for human HLA-DR
and CIITA (20) and mouse
IA (21). Other primers used included human and mouse
GAPDH (Clontech Laboratories, Palo Alto, CA) and
5'-ACACCTGGACCTGGACTCAC-3' (forward) and 5'-GCTCTTGGCTCCTTTGTCAC-3'
(reverse) for mouse CIITA, 5'-GAAACTGGTGAGTGACTGC-3' (forward) and
5'-CACATTGGAGAAGAAGCC-3' (reverse) for human CD40 (22),
and 5'-GTTTAAAGTCCCGGATGCGA-3' (forward) and 5'-CTCAAGGCTATGCTGTCTGT-3'
(reverse) for mouse CD40 (23).
The real-time fluorescent PCR technique was used to quantitate fold
changes in GAPDH, CIITA, MHC class II, and CD40. The detection system
and quantitative PCR have been described in detail elsewhere
(24). The threshold cycle number
(CT) corresponds to the cycle number at
which the fluorescent emission reaches 10 SDs above the mean
fluorescent emission measured during the early "baseline" cycles of
the PCR. The CT values for triplicates were
averaged and normalized to GAPDH levels (i.e., average
CT for HLA-DR on sample cDNA minus average
CT for GAPDH on same sample =
normalized CT or
CT for that sample). Assuming reverse
transcription efficiencies were approximately the same for each sample,
this allowed us to determine relative amounts of mRNA using the
comparative CT
(CT) method (Perkin-Elmer Taqman User
Bulletin 2). Fold changes in mRNA levels were calculated as
2x, where x = the difference between the
GAPDH normalized CT values of the
control and experimental samples.
Validation experiments showed a linear relationship between input cDNA
and CT values over a range of serial
dilutions of cDNA extending from 1 to 10-6. The
slopes of these lines generated for each primer/probe set indicated
that PCR efficiencies for each of the primer sets were close to 100%
and validated the use of the CT
(comparative CT) method for relative
quantitation. These experiments also demonstrated the linear range of
amplification and allowed determination of the maximum
CT value at which linear amplification was
reliable. For most of our primer/probe sets, this value was 40; if the
samples did not amplify by 40 cycles, the gene for that sample was
considered not detectable (ND), because CT
values beyond this point would not meet the criteria for calculation of
fold change by the comparative CT method.
In such cases, to calculate a minimum fold change for other samples in
the same group, the CT value for the not
detectable sample was arbitrarily set at 40. Primer sets for real-time
PCR are available at http://realtime.nwu.edu.
Flow cytometry
Flow cytometric analyses were conducted by published methods (25) with cells fixed with 1.0% paraformaldehyde and analyzed on a FACScan (Becton Dickinson). Cell cycle analyses were conducted by flow cytometry with PI staining of genomic DNA. Cells were fixed and permeabilized in methanol, treated with RNaseA and PI and analyzed for DNA content by determination of FL3-area on the FACScan. The FL3-area data were analyzed with the ModFit software for determination of the percent cells in each phase of the cell cycle. In some cases, FITC-conjugated Ab staining was conducted before PI treatment and surface staining was analyzed as a function of the cell cycle phase. Apoptosis was analyzed by staining with annexin V directly conjugated with either FITC or R-PE and by sub-G0 analysis of DNA staining. Data are representative of at least three independent experiments.
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Results |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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Our experiments were initially designed to explore whether the
failure of certain tumor cells to express MHC class II might be a
result of a repressive chromatin structure. Therefore, we tested the
effect of treatment with the DAIs, NaBu and TSA, on the activation of
class II genes in selected tumor cell lines. As shown in Fig. 1
, treatment of the human neuroblastoma
cell line SK-N-MC (MC) with 100 nM TSA or 1 mM NaBu resulted in a
substantial increase in the levels of mRNA for HLA-DR (DR) as measured
by RT-PCR, whereas CIITA transcription remained not detectable. Levels
of mRNA for the housekeeping gene, GAPDH, were not significantly
changed after treatment. Low levels of mRNA for the DP and DQ isotypes
of MHC class II were also induced by TSA (data not shown). Although TSA
and NaBu activate transcription of MHC class II in MC tumor cells,
these cells cannot be induced to express mRNA for DR by IFN- (Fig. 1A). This is not due to the failure of MC cells to express
IFN- receptors or to a defect in the IFN- signaling pathway,
because mRNA for two prototype IFN--inducible genes, IFN regulatory
factor-1 and guanylate binding protein, is enhanced by IFN-
treatment (data not shown). Essentially identical results to those of
MC were obtained with JAR, a human trophoblast cell line, that was
refractory to IFN- but inducible for class II by TSA and
NaBu.
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B,
treatment of MC cells with 100 nM TSA for 24 h resulted in a
108-fold increase in expression of DR, but no increase in CIITA was
detectable after 60 cycles of PCR. NaBu treatment (1 mM, 48
h) resulted in an 88-fold enhancement in DR and no activation of CIITA
(Fig. 1B). Thus, within the limits of these quantitative
measurements, the activation of DR gene expression by DAIs appears to
occur with no change in the level of CIITA transcripts. Although
IFN- did not induce HLA-DR transcripts on MC cells at any
concentration or time point studied, this cytokine did elicit low
levels of CIITA mRNA detectable by real-time PCR. Preliminary studies
showed that the sequence of transcripts stimulated by IFN- in MC
cells are very similar to a minor alternatively spliced Raji
transcript, which has a deletion in the N-terminal activation domain,
and is functionally inactive (26).
To determine whether the effect of DAIs on MHC class II was manifested
in other tumors and in different species, we tested the effect of TSA
on two commonly used mouse tumor models: the Colon 26 adenocarcinoma
and the J558 plasmacytoma. Analysis by standard RT-PCR at 35 cycles
showed that IFN- induced expression of CIITA and class II in Colon
26 and, similar to the human MC cell line, CIITA was not detected after
TSA treatment (Fig. 2A).
Real-time PCR analysis of Colon 26 cells was consistent with the RT-PCR
data and demonstrated that TSA induced a 74-fold increase in Ia in the
absence of CIITA, whereas IFN- elicited both CIITA (20-fold) and Ia
(58-fold) expression. The J558 cell line behaved quite differently from
Colon 26 in that IFN- did not activate either CIITA or class II
expression (Fig. 2B). Because the classical
IFN--inducible genes IFN regulatory factor-1 and guanylate binding
protein, as well as MHC class I mRNA, were not induced by IFN- in
J558 (data not shown), this cell line may have a signaling defect in
the IFN- pathway. However, unlike either the MC or Colon 26 cells,
TSA induced both CIITA (102-fold) and Ia (537-fold) in J558 as shown in
Fig. 2. In different experiments, maximum expression at 12 h
occurred at TSA concentrations between 50 and 100 nM and at 24
h between 25 and 50 nM. At higher concentrations or longer incubation
times, decreasing expression was associated with increasing apoptosis.
The contrasting patterns in regard to CIITA induction suggest the
possibility that TSA may function by both CIITA-dependent (in J558) and
-independent (in MC and Colon 26) mechanisms in different cell
types.
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. TSA
treatment of this cell line resulted in a 51-fold increase in mRNA for
HLA-DR (Fig. 3) and in the appearance of
substantial levels of cell surface DR (data not shown). These data
support the existence of a CIITA-independent TSA induction of MHC class
II expression.
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Because of the importance of costimulatory molecules in the
cellular activation mechanism involving MHC complexes, we explored
whether DAIs altered the expression of B7-1, B7-2, or CD40 genes.
Although no consistent changes in B7-1 and B7-2 were detected, CD40
expression was enhanced by TSA in all three cell lines (Fig. 4). MC cells constitutively expressed low
levels of CD40 and IFN- enhanced CD40 mRNA levels 5-fold, whereas
TSA elicited a 12-fold increase by real-time PCR (Fig. 4A).
Colon 26 cells did not constitutively express CD40 and mRNA levels
remained undetectable at 60 cycles after IFN- treatment, although
TSA induced a 147-fold increase in CD40. J558 cells showed low
constitutive levels of CD40 mRNA that did not change after treatment
with IFN-, whereas TSA induced a 161-fold increase in CD40. The lack
of induction of CD40 by IFN- in J558 cells is consistent with the
failure of IFN- to activate other IFN- inducible genes. The
failure of IFN- to induce CD40 mRNA in Colon 26, which has no
apparent defect in the IFN- signaling pathway, is unexplained, but
may be related to the ability of TSA to activate factors in addition to
those elicited by IFN- that are required for CD40 gene
expression.
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To determine the effect of TSA on cell surface protein expression,
the J558 cell line was subjected to FACScan analyses. Of 13 markers
analyzed, only class I, II, and CD40 were enhanced at low
concentrations of TSA. Levels of several nonexpressed Ags were not
elicited (including CD25, ß7 integrin, CD40L,
H-2Kk, and I-Ak). Other
markers were expressed, but were not altered (including CD44, CD54,
CD80, CD86, and CD119) by TSA treatment. In each cell line tested, TSA
significantly induced the expression of MHC class II protein on the
cell surface with induction ranging from 3- to 21-fold in different
cells, as indicated by the percent positive cells shown in Fig. 5. The highest expression (21-fold
induction) was achieved on J558 cells after 24 h of treatment with
25 nM TSA. IFN- did not induce class II on any of the cell lines
described here. MHC class I expression was also found to be enhanced by
TSA in each cell type, but, in contrast to MHC class II, TSA was less
effective than IFN- in MC and Colon 26, whereas J558 was
unresponsive to IFN-. However, similar to the class II data, TSA
induced expression of CD40 on all three cell types and did so more
effectively than IFN-.
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TSA has been described as a cell cycle inhibitor that induces both
G1 and, especially, G2-M
blocks (17). MHC class II, but not class I, has been
reported (29, 30) to be regulated differentially during
the cell cycle. We therefore analyzed the cell cycle of our tumor lines
(by DNA staining and flow cytometry) with each of the treatment
conditions under study. From these studies, we learned which TSA
concentrations significantly altered the cell cycle, and therefore
designed our experiments to include conditions that did not affect cell
cycle progression in the cell lines examined. In Fig. 5, the conditions
employed did not significantly alter the cell cycle (see cell cycle
analysis Fig. 6A) and yet 29%
of cells were positive for class II expression in MC, 53% in Colon 26,
and 75% in J558. Two additional methods were used to analyze cell
cycle effects on MHC expression. Because TSA arrests the cell cycle
mainly in G2/M, and class II has been reported to
accumulate in G2-M (30), we studied
the effects of the cell cycle inhibitor nocodazole, which arrests cells
in G2-M (Fig. 6B). Nocodazole arrested
nearly 100% of J558 cells in G2-M and resulted
in only 23% MHC class II-positive cells (Fig. 6C). However,
10 nM TSA resulted in only 19% of cells in G2-M
and yet this treatment produced 75% MHC class II-positive cells (Fig. 6D). This indicated that a significant proportion of the MHC
class II expression in the TSA-treated population was independent of
G2-M. Also, using double staining with PI and Ab
to class I on J558 cells, we found TSA induced expression throughout
the cell cycle and enhancement was not restricted to a specific phase.
From the above experiments, we conclude that TSA has specific effects
on MHC gene expression that are not solely a result of its action on
the cell cycle. Although our data demonstrated CD40 expression at TSA
concentrations that do not alter the cell cycle, more detailed studies
are required to clarify the cell cycle regulation of CD40.
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, selected for a normal cell cycle, apoptosis was not significantly
different from untreated controls. To demonstrate that the MHC class II
expression induced by TSA was, in large part, independent of apoptosis,
we stained J558 cells for I-Ad and annexin V.
Subsequent analyses were gated on the annexin V-negative population and
assayed for class II expression (Fig. 7).
We conclude that TSA has specific effects on gene expression over and
above any effects that may result from apoptosis. However, at higher
concentrations (differing for each cell type) apoptosis may result in
changes in gene expression and complicate the interpretation of
experiments using DAIs.
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Discussion |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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failed to
induce MHC class II Ags on the cell surface, but, following TSA
treatment, each cell line elicited substantial expression detected by
flow cytometry. Enhanced expression of class I and CD40 were elicited
by low concentrations of TSA, whereas the expression of 10 other
surface Ags remained essentially unchanged. Higher concentrations of
TSA induced Fas and FasL expression and apoptosis. Thus, as expected,
various genes appear to differ in their sensitivity to DAIs that likely
reflects inherent differences in chromatin structure and nucleosomal
positioning in the promoters regulating these genes.
The current paradigm for MHC gene expression suggests that CIITA is
required for constitutive and induced class II transcription, IFN-
enhanced MHC class I expression, and perhaps plays a nonessential role
in maintaining constitutive levels of transcription of class I. This
study, together with other recently reported evidence (21, 32, 33, 34), suggests that, under specific experimental
circumstances and in certain cell types, class II may be expressed
independently of CIITA. One explanation of our data is that CIITA
expression is defective or repressed in MC and Colon 26, and an
alternative pathway exists that is activated by DAIs that completely
bypasses CIITA. This is especially evident in the CIITA-mutant cell
line RJ2.2.5. In normal IFN- responsive cells, the CIITA protein
interacting with other factors may function to facilitate an open
chromatin structure at the class II promoter. Consistent with this
thesis are the reports that, in certain cells lacking endogenous CIITA,
the DR, Ii, and DM promoters are not occupied (35), and
that transfection of CIITA results in the appearance of DNase
hypersensitivity sites in the class II promoter and expression of the
DR gene (36). These observations, together with the
finding that CIITA does not alter the stability of the higher order
multiprotein complexes that bind to the class II promoter
(37), supports the thesis that the CIITA complex enhances
access of transcription factors to DNA binding sites by altering
chromatin. Several recent studies have reported (2, 38, 39) that the histone acetyltransferase cofactor CBP/p300 binds
to CIITA and significantly enhances its activity. TSA, at least in
part, could replace the function of a CIITA-CBP/p300 coactivator
complex by acetylating histones and relieving nucleosome repression
allowing promoter occupancy and some level of transcription in the
absence of CIITA.
An additional model that is consistent with current dogma and with the
data presented here is that, in certain cell types, DAIs release
chromatin repression of the CIITA gene. The activation of CIITA and MHC
class II after treatment of the J558 plasmacytoma with TSA is
consistent with this hypothesis. Active suppression of CIITA and class
II has been postulated during the differentiation from B cells to
plasma cells (8) and in trophoblast cells (20, 40). Inhibition of CIITA transcription could be mediated by a
repressor that recruits HDACs to the CIITA promoter, analogous to the
repression mediated by HDAC recruitment by nuclear receptors, the
Mad/Max and Rb-E2F complexes, and by the methyl-CpG-binding
proteins (15, 16). A likely candidate is the Blimp-1
transcriptional repressor, a regulator of terminal B cell
differentiation that has been shown to associate with HDACs
(41). The surface expression of class II on TSA-treated
J558 cells is high, approaching that on B cells and IFN- inducible
cells; compared with the relatively low levels found in the
CIITA-negative MC and Colon 26 cells. This suggests that, although
class II expression may occur without CIITA expression, CIITA may be
required for high levels of expression. The difference in inducibility
of CIITA by TSA observed in the cell lines in this study could be
related to the distinct promoters (PI
PIV) and their differential
usage in regulating CIITA in various cell types (42). J558
is of B cell lineage and should preferentially utilize PIII, the B cell
promoter. One possibility is that, in the transition from a B cell to a
plasma cell, the PIII promoter is repressed by chromatin and that this
is reversed by TSA, leading to the activation of CIITA and class II in
plasma cells.
In summary, DAIs could function in activating MHC class II in
different cell types by one of several mechanisms that are not mutually
exclusive: 1) Acetylating histones at the MHC promoter, thus providing
a function normally performed by the CIITA-CBP/p300 coactivation
complex. 2) Activating expression of the CIITA gene that is repressed
by chromatin. This would include CIITA promoters that are methylated on
CpG islands and bind methyl CpG binding proteins that recruit HDACs
(16). In the trophoblast cell line, Jar, the PIV CIITA
promoter is methylated and, although 5-azacytidine alone does not
activate CIITA and class II, such treatment does support low level
expression after IFN- treatment (43). In view of the
potential synergy of demethylation and DAIs in activating certain
epigenetically silenced genes (44), future studies of the
effect of combinations of DAIs and demethylating agents will be of
interest. 3) Altering the levels and/or activation state of essential
cofactors such as CBP/p300. 4) Direct acetylation of the CIITA protein,
or other factors required in class II transactivation, by DAIs could
also affect their activity or nuclear transport.
Because the class I and II promoters show significant sequence homologies (6), and both utilize CIITA as a coactivator (4, 5), DAIs may be functioning by similar mechanisms in both class I and II. Tissue specific expression of class I is determined, in significant part, by transcriptional mechanisms that depend on upstream negative regulatory elements in the class I promoter (45) and factors binding to these elements may recruit HDACs and repress transcription. The mechanism of TSA activation of CD40 is unclear and we are not aware of data on the promoter structure of CD40 that would help clarify this issue. The observation that TSA can induce surface expression of CD40 has potential biological implications. CD40 ligand binds to the CD40 receptor and promotes tumor killing by CTLs, directly induces apoptosis of certain tumor cells (46), as well as enhances the ability of normal DC to present Ag and elicit CTLs (47). Thus, up-regulating CD40 expression could potentially be useful in future attempts to treat cancers by the administration of soluble CD40L or cells genetically modified to express CD40L genes.
Epigenetic phenomena including methylation and histone acetylation are critical in regulating gene expression through repression (14), and complex patterns of chromatin activation and repression are undoubtedly involved in many fundamental immune processes (48, 49, 50). These alterations in chromatin structure would likely involve changes in the pattern of histone acetylation. Thus, agents that hyperacetylate histones may be able to substitute for natural signals that release repression during cellular differentiation and, as shown in this study, these same agents may elicit expression of repressed genes critical in the immune response against tumors. The work reported here also suggests that some of the variation in immunogenicity and tumorigenicity of different tumors may be mediated by regulatory events involving reversible acetylation patterns, and these could potentially be altered by treatment or targeting with agents that alter acetylation.
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Acknowledgments |
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Footnotes |
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2 Address correspondence and reprint requests to Dr. Thomas B. Tomasi, Department of Immunology, Elm and Carlton Streets, Roswell Park Cancer Institute, Buffalo NY 14263.
3 Abbreviations used in this paper: CIITA, class II transactivator; HDAC, histone deacetylase; DAIs, HDAC inhibitors; TSA, trichostatin A; PI, propidium iodide.
Received for publication June 2, 2000. Accepted for publication September 21, 2000.
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 R. Holtz, J. C. Choi, M. G. Petroff, J. F. Piskurich, and S. P. Murphy Class II Transactivator (CIITA) Promoter Methylation Does Not Correlate with Silencing of CIITA Transcription in Trophoblasts Biol Reprod, September 1, 2003; 69(3): 915 - 924. [Abstract] [Full Text] [PDF]
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 E. Zika, S. F. Greer, X.-S. Zhu, and J. P.-Y. Ting Histone Deacetylase 1/mSin3A Disrupts Gamma Interferon-Induced CIITA Function and Major Histocompatibility Complex Class II Enhanceosome Formation Mol. Cell. Biol., May 1, 2003; 23(9): 3091 - 3102. [Abstract] [Full Text] [PDF]
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S. Das, J.-H. Lin, J. Papamatheakis, Y. Sykulev, and P. N. Tsichlis Differential Splicing Generates Tvl-1/RFXANK Isoforms with Different Functions J. Biol. Chem., November 15, 2002; 277(47): 45172 - 45180. [Abstract] [Full Text] [PDF]
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S. P. Murphy, R. Holtz, N. Lewandowski, T. B. Tomasi, and H. Fuji DNA Alkylating Agents Alleviate Silencing of Class II Transactivator Gene Expression in L1210 Lymphoma Cells J. Immunol., September 15, 2002; 169(6): 3085 - 3093. [Abstract] [Full Text] [PDF]
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R. Mudhasani and J. D. Fontes The Class II Transactivator Requires brahma-Related Gene 1 To Activate Transcription of Major Histocompatibility Complex Class II Genes Mol. Cell. Biol., July 15, 2002; 22(14): 5019 - 5026. [Abstract] [Full Text] [PDF]
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 A. A. Jazaeri, C. J. Yee, C. Sotiriou, K. R. Brantley, J. Boyd, and |
