Precipitous Release of Methyl-CpG Binding Protein 2 and Histone Deacetylase 1 from the Methylated Human Multidrug Resistance Gene (MDR1) on Activation

Abstract

Overexpression of the human multidrug resistance gene 1 (MDR1) is a negative prognostic factor in leukemia. Despite intense efforts to characterize the gene at the molecular level, little is known about the genetic events that switch on gene expression in P-glycoprotein-negative cells. Recent studies have shown that the transcriptional competence of MDR1 is often closely associated with DNA methylation. Chromatin remodeling and modification targeted by the recognition of methylated DNA provide a dominant mechanism for transcriptional repression. Consistent with this epigenetic model, interference with DNA methyltransferase and histone deacetylase activity alone or in combination can reactivate silent genes. In the present study, we used chromatin immunoprecipitation to monitor the molecular events involved in the activation and repression of MDR1. Tumors become resistant to chemotherapy by a variety of mechanisms, including altered DNA repair, alterations in scavenging enzymes, and increased drug efflux. Although clinical drug resistance may be multifactorial, understanding these various mechanisms is important and may afford a means of improving treatment regimens. One of the most extensively studied mechanisms of multidrug resistance is associated with the overexpression of the MDR1 product, P glycoprotein (Pgp) (for reviews see references 27 and 55). A transmembrane protein, Pgp acts as an efflux pump, reducing intracellular drug levels and thus cytotoxic activity. We and others have demonstrated that the human MDR1 promoter is activated by changes in CpG methylation in B-cell chronic lymphocytic leukemia (26), acute myeloid leukemia (38), and tumor cell lines (12).DNA hypermethylation is associated with transcriptional silencing and is accompanied by the accumulation of deacetylated histones on heterochromatin (13,21). Methylation is often associated with the pathological silencing of tumor suppressor genes in human cancer and neurodevelopmental syndrome (20, 23). In contrast, transcriptionally active chromatin is characterized by the enrichment of hyperacetylated histone and is generally associated with hypomethylated chromatin (2, 34). The mechanisms underlying the correlation between DNA methylation and histone deacetylation in the control of gene expression have been established by recent biochemical experiments (24,42). Evidence has emerged that a family of methylCpG binding proteins binds heterochromatin to stably repress transcription (10, 18). The transcriptional repressor methylCpG binding protein 2 (MeCP2 ) is the best-characterized family member (18,41). MeCP2 is typically associated on heterochromatin in a methylation-dependent manner by its methyl-binding domain (MBD) and can displace histone H1 for nucleosome binding, indicating that it can dynamically interact with assembled chromatin (30,40). The transcriptional-repression domain of MeCP2 recruits the corepressors mSin3 and histone deacetylases (HDACs) (24, 42), causing transcriptional repression (41). Moreover, repression by MeCP2 is partial...

The histone deacetylase inhibitor trichostatin A downregulates human MDR1 (ABCB1) gene expression by a transcription-dependent mechanism in a drug-resistant small cell lung carcinoma cell line model

“…Both cell lines appeared to display hypomethylated ABCB1 DNA ( Figure 6B). These data confirm the sensitivity of hypomethylated ABCB1 promoter to the direct action of HDACs (El-Osta et al, 2002). Treatment of H69WT and H69VP by TSA (330 nmol l À1 , 24 h), or the demethylating agent 5azadC (2 mmol l À1 , 72 h), or a combination of both (cells treated with 5azadC for 72 h were resuspended in fresh supplemented medium and harvested 33 h later.…”

“…For instance, ABCB1 gene was overexpressed in SW620 colon carcinoma and CEM-Bcl2 cells exposed to TSA (Jin and Scotto, 1998;Baker et al, 2005), or KU812 and NB4 cells exposed to depsipeptide (Tabe et al, 2006;Yamada et al, 2006). In drug-resistant cells, HDAC inhibition also increased ABCB1 expression in CEM-A7R, Kasumi-1, and Kasumi-6 cell lines (El-Osta et al, 2002;Tabe et al, 2006). On the contrary, our results showed a repression of ABCB1 gene expression by TSA and NaB.…”

“…Epigenetic mechanisms (histone acetylation, DNA methylation) have been shown to play a pivotal role in ABCB1 gene expression in several tumour cell systems (El-Osta et al, 2002;Labialle et al, 2002;Scotto, 2003).…”

The temporal relationship between ABCB1 promoter hypomethylation, ABCB1 expression and acquisition of drug resistance

“…This synergy between DNA methyltransferase inhibitors and histone deacetylase inhibitors in stimulating ABCB1 mRNA expression is supported by another study, which showed that TSA or Aza treatment alone had little effect or weakly induced ABCB1 expression, respectively, in CEM-CCRF cells. 27 In contrast, treatment of these cells with both Aza and TSA induced a 10-fold higher level of ABCB1 expression than with Aza treatment alone. The study further showed that Aza-induced demethylation of the ABCB1 promoter resulted in the release of methyl-CpG-binding protein-2 from the promoter, promoter acetylation and partial transcriptional activation at the downstream ABCB1 promoter.…”

“…The study further showed that Aza-induced demethylation of the ABCB1 promoter resulted in the release of methyl-CpG-binding protein-2 from the promoter, promoter acetylation and partial transcriptional activation at the downstream ABCB1 promoter. 27 Interestingly, another study showed that acetylation of the ABCB1 downstream promoter was dependent upon its methylation state. Partial hypomethylation of the ABCB1 promoter in CEM-Bcl2 cells induced ABCB1 transcription and daunorubicin-induced H3 acetylation of the ABCB1 promoter preceded such changes in ABCB1 expression.…”