DNA Methylation Does Not Stably Lock Gene Expression but Instead Serves as a Molecular Mark for Gene Silencing Memory

Abstract

DNA methylation is commonly thought of as a "molecular lock" that leads to permanent gene silencing. To investigate this notion, we tested 24 different HDAC inhibitors (HDACi) on colon cancer cells that harbor a GFP locus stably integrated and silenced by a hypermethylated CMV promoter. We found that HDACi efficiently reactivated expression of GFP and many other endogenous genes silenced by DNA hypermethylation. After treatment, all promoters were marked with active chromatin, yet DNA hypermethylation did not change. Thus, DNA methylation could not prevent gene reactivation by drug-induced resetting of the chromatin state. In evaluating the relative contribution of DNA methylation and histone modifications to stable gene silencing, we followed expression levels of GFP and other genes silenced by DNA hypermethylation over time after treatment with HDACi or DNA demethylating drugs. Reactivation of methylated loci by HDACi was detectable for only 2 weeks, whereas DNA demethylating drugs induced permanent epigenetic reprogramming. Therefore, DNA methylation cannot be considered as a lock for gene expression, but rather as a memory signal for long-term maintenance of gene silencing. These findings define chromatin as an important druggable target for cancer epigenetic therapy and suggest that removal of DNA methylation signals is required to achieve long-term gene reactivation.

Epigenetics in the Biology and Treatment of Multiple Myeloma

“…17 DNA methylation at transcription start sites and gene promoters has been associated with the formation of heterochromatin and long-term gene silencing. 30,31 Methylation is also an inactivating mechanism of the tumour suppressor p16 gene in MM, which is associated with high plasma cell proliferation and short survival. 32 Furthermore, p16 methylation may be a marker for overall epigenetic changes associated with disease progression.…”

Epigenetic aspects of MDS and its molecular targeted therapy

“…There are HDIs currently in clinical trials; however, less data is available for their use in MDS, though anecdotal responses have been reported [75]. Recently, it has also been reported that DNA methylation per se is not a permanent lock for silencing gene expression, but rather a combination of DNA methylation inhibitors with other drugs (such as HDIs) can be used to reactivate gene expression [76,77]. Furthermore, there is increasing evidence that such combinations have encouraging results [78,79].…”

The Sequential Combination Paradigm in Epigenetic Therapy

“…The use of a DNA Methyl Transferase (DNMT) inhibitor followed by an HDAC inhibitor is required for the re-expression of genes silenced by promoter hypermethylation. Consequently, the sequential or overlapping combination of DNA hypomethylating agents and HDAC inhibitors has been utilized in several clinical trials based on this hypothesis [2,3].Although few previous reports did not support this hypothesis, they were considered exceptions for specific genes or for a specific class of HDAC inhibitors (SIRT1 inhibitors) [4][5][6]. Recently, a more comprehensive study challenged this hypothesis and demonstrated that different classes of HDAC inhibitors induce transient expression of promoter hypermethylated genes without the loss of DNA promoter hypermethylation [5].…”

“…Recently, a more comprehensive study challenged this hypothesis and demonstrated that different classes of HDAC inhibitors induce transient expression of promoter hypermethylated genes without the loss of DNA promoter hypermethylation [5]. On the other hand, DNA hypomethylating agents induced permanent and stable epigenetic reprogramming.…”

“…Although few previous reports did not support this hypothesis, they were considered exceptions for specific genes or for a specific class of HDAC inhibitors (SIRT1 inhibitors) [4][5][6]. Recently, a more comprehensive study challenged this hypothesis and demonstrated that different classes of HDAC inhibitors induce transient expression of promoter hypermethylated genes without the loss of DNA promoter hypermethylation [5].…”