Abstract
In mammals, DNA methylation is necessary for the maintenance of genomic stability, gene expression regulation, and other processes. During malignant diseases progression, changes in both DNA methylation patterns and DNA methyltransferase (MTase) genes are observed. Human de novo MTase DNMT3A is most frequently mutated in acute myeloid leukemia (AML) with a striking prevalence of R882H mutation, which has been extensively studied. Here, we investigate the functional role of the missense mutations (S714C, R635W, R736H, R771L, P777R, and F752V) found in the catalytic domain of DNMT3A in AML patients. These were accordingly mutated in the murine Dnmt3a catalytic domain (S124C, R45W, R146H, R181L, P187R, and F162V) and in addition, one-site CpG-containing DNA substrates were used as a model system. The 3–15-fold decrease (S124C and P187R) or complete loss (F162V, R45W, and R146H) of Dnmt3a-CD methylation activity was observed. Remarkably, Pro 187 and Arg 146 are not located at or near the Dnmt3a functional motives. Regulatory protein Dnmt3L did not enhance the methylation activity of R45W, R146H, P187R, and F162V mutants. The key steps of the Dnmt3a-mediated methylation mechanism, including DNA binding and transient covalent intermediate formation, were examined. There was a complete loss of DNA-binding affinity for R45W located in the AdoMet binding region and for R146H. Dnmt3a mutants studied in vitro suggest functional impairment of DNMT3A during pathogenesis.
Highlights
DNA methylation is an important epigenetic modification essential for the regulation of cell processes in both eukaryotes and prokaryotes
DNA methylation is necessary for the maintenance of genomic stability, gene expression regulation, and other processes [1,2]
This study aimed to examine the functional consequences of the widespread acute myeloid leukemia (AML) missense mutations found in the human DNMT3A catalytic domain
Summary
DNA methylation is an important epigenetic modification essential for the regulation of cell processes in both eukaryotes and prokaryotes. DNA methylation is necessary for the maintenance of genomic stability, gene expression regulation, and other processes [1,2]. DNA methylation is maintained by DNA methyltransferases (MTases), which transfer a methyl group from co-factor S-adenosyl-l-methionine (AdoMet) to the five positions of the cytosine residue mainly within. Changes in the methylation process occur in various human pathologies [4]. There are alterations in the pattern of methylation and multiple disorders in the MTases, including missense mutations [4,5,6,7]. The function of mammalian Dnmt3a is to methylate DNA de novo [3]
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