AML-associated DNMT3A R882H mutants exert dominant-negative effects leading to hypomethylation.
Major finding: AML-associated DNMT3A R882H mutants exert dominant-negative effects leading to hypomethylation.
Mechanism: R882H mutants heterodimerize with WT DNMT3A and prevent the formation of active WT homotetramers.
Impact: R882 and non-R882 DNMT3A mutations may contribute to AML through distinct mechanisms.
Heterozygous DNMT3A mutations affecting R882 within the methyltransferase catalytic domain account for approximately 60% of DNMT3A mutations in acute myeloid leukemia (AML), which are commonly found in patients with normal karyotype AML (NK-AML). These mutations reduce de novo DNA methylation in vitro, but the underlying mechanism is unclear. Russler-Germain and colleagues utilized a cohort of 80 NK-AML patient samples in order to further explore the impact of R882 alleles on wild-type (WT) DNMT3A methyltransferase activity. Allele frequency and mass spectrometry analysis revealed that DNMT3A R882 mutations were typically heterozygous, originated in NK-AML founder clones, and were expressed to the same extent as the WT allele. In addition, heterozygous R882-mutant samples showed a small but statistically significant reduction in genome-wide methylation compared with WT that was not seen in non-R882 DNMT3A-mutant samples. Hypomethylation profiles were nearly identical in NK-AMLs harboring R882H, the most common R882 mutation, as well as R882C mutants. Furthermore, hypomethylation at specific CpGs was correlated with expression changes of associated genes, although genes were both up- and downregulated. Although mixing recombinant WT and R882H DNMT3A in vitro did not affect WT DNMT3A enzymatic activity, coexpression of WT and R882H DNMT3A in human cells resulted in an 80% reduction in methyltransferase activity compared with WT DNMT3A alone, confirming that the R882H mutant possesses dominant-negative activity. The R882H mutant proteins did not inhibit WT DNMT3A activity by affecting subcellular localization or modifying CpG substrate specificity, but rather formed heterodimers with WT DNMT3A that inhibited the formation of active WT homotetramers. The finding that R882 DNMT3A mutants specifically confer a hypomethylation phenotype by inhibiting DNMT3A activity in a dominant-negative manner raises the possibility that R882 and non-R882 DNMT3A-mutant AML may be etiologically distinct.
Russler-Germain DA, Spencer DH, Young MA, Lamprecht TL, Miller CA, Fulton R, et al. The R882H DNMT3A mutation associated with AML predominantly inhibits wild-type DNMT3A by blocking its ability to form active tetramers. Cancer Cell 2014 Mar 20 [Epub ahead of print].
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