Modulation of DNA methylation by catechol-O-methyltransferase

Abstract

2749

The DNA methyltransferase (DNMT)-mediated methylation of DNA at the 5-position of cytosine within the CpG dinucleotides represents a basic mechanism for epigenetic control of gene expression and maintenance of genome integrity. In the present investigation, we studied the modulating effects of catechol-O-methyltransferase (COMT) and S-adenosyl-L-homocysteine (AdoHcy) on DNA methylation catalyzed by prokaryotic SssI DNMT and human DNMT1. We found that the presence of COMT (at physiologically-relevant concentrations) enhanced the rate of DNA methylation in vitro catalyzed by SssI DNMT and human DNMT1, with an enhancement of up to 1-fold over the control rate. Additional experiments using cultured human MCF-7 cells showed that transfection of the COMT siRNAs (at 100 nM) into these cells caused ∼50% suppression of COMT expression, and this reduction of the cellular COMT levels resulted in a decrease of the methylation status of the RARβ gene (a representative gene), although significant changes of the global DNA methylation status were not detected. This data demonstrated that decreased expression of the COMT by ∼50% in short period of 10 days resulted in a detectable change in the methylation status of certain genes. Kinetic studies showed that AdoHcy strongly and noncompetitively inhibited the methylation of DNA by competing S-adenosyl-L-methionine off the DNMT, thus shifting more enzyme molecules to a form that is bound with AdoHcy. Consequently, the V_max values were reduced in the presence of AdoHcy, but the K_m values were not changed. In the presence of COMT, the V_max values for enzymatic DNA methylation were increased, but the K_m values were essentially not altered. Our data showed that the enhancement of DNA methylation by COMT likely is due to the sequestration of AdoHcy by COMT, which reduces the availability of the free AdoHcy for DNMT inhibition.