A Model System for Studying the DNMT3A Hotspot Mutation (DNMT3A^R882) Demonstrates a Causal Relationship between Its Dominant-Negative Effect and Leukemogenesis.

Abstract

Mutation of DNA methyltransferase 3A at arginine 882 (DNMT3A^R882mut) is prevalent in hematologic cancers and disorders. Recently, DNMT3A^R882mut has been shown to have hypomorphic, dominant-negative, and/or gain-of-function effects on DNA methylation under different biological contexts. However, the causal role for such a multifaceted effect of DNMT3A^R882mut in leukemogenesis remains undetermined. Here, we report TF-1 leukemia cells as a robust system useful for modeling the DNMT3A^R882mut-dependent transformation and for dissecting the cause-effect relationship between multifaceted activities of DNMT3A^R882mut and leukemic transformation. Ectopic expression of DNMT3A^R882mut and not wild-type DNMT3A promoted TF-1 cell transformation characterized by cytokine-independent growth, and induces CpG hypomethylation predominantly at enhancers. This effect was dose dependent, acted synergistically with the isocitrate dehydrogenase 1 (IDH1) mutation, and resembled what was seen in human leukemia patients carrying DNMT3A^R882mut. The transformation- and hypomethylation-inducing capacities of DNMT3A^R882mut relied on a motif involved in heterodimerization, whereas its various chromatin-binding domains were dispensable. Mutation of the heterodimerization motif that interferes with DNMT3A^R882mut binding to endogenous wild-type DNMT proteins partially reversed the CpG hypomethylation phenotype caused by DNMT3A^R882mut, thus supporting a dominant-negative mechanism in cells. In mice, bromodomain inhibition repressed gene-activation events downstream of DNMT3A^R882mut-induced CpG hypomethylation, thereby suppressing leukemogenesis mediated by DNMT3A^R882mut. Collectively, this study reports a model system useful for studying DNMT3A^R882mut, shows a requirement of the dominant-negative effect by DNMT3A^R882mut for leukemogenesis, and describes an attractive strategy for the treatment of leukemias carrying DNMT3A^R882mut. SIGNIFICANCE: These findings highlight a model system to study the functional impact of a hotspot mutation of DNMT3A at R882 in leukemia.