Haploinsufficiency for DNA methyltransferase 3A predisposes hematopoietic cells to myeloid malignancies
Christopher B. Cole, … , Christopher A. Miller, Timothy J. Ley
Christopher B. Cole, … , Christopher A. Miller, Timothy J. Ley
Published September 5, 2017
Citation Information: J Clin Invest. 2017;127(10):3657-3674. https://doi.org/10.1172/JCI93041.
View: Text | PDF
Research Article Hematology Oncology Article has an altmetric score of 8

Haploinsufficiency for DNA methyltransferase 3A predisposes hematopoietic cells to myeloid malignancies

Abstract

The gene that encodes de novo DNA methyltransferase 3A (DNMT3A) is frequently mutated in acute myeloid leukemia genomes. Point mutations at position R882 have been shown to cause a dominant negative loss of DNMT3A methylation activity, but 15% of DNMT3A mutations are predicted to produce truncated proteins that could either have dominant negative activities or cause loss of function and haploinsufficiency. Here, we demonstrate that 3 of these mutants produce truncated, inactive proteins that do not dimerize with WT DNMT3A, strongly supporting the haploinsufficiency hypothesis. We therefore evaluated hematopoiesis in mice heterozygous for a constitutive null Dnmt3a mutation. With no other manipulations, Dnmt3a+/– mice developed myeloid skewing over time, and their hematopoietic stem/progenitor cells exhibited a long-term competitive transplantation advantage. Dnmt3a+/– mice also spontaneously developed transplantable myeloid malignancies after a long latent period, and 3 of 12 tumors tested had cooperating mutations in the Ras/MAPK pathway. The residual Dnmt3a allele was neither mutated nor downregulated in these tumors. The bone marrow cells of Dnmt3a+/– mice had a subtle but statistically significant DNA hypomethylation phenotype that was not associated with gene dysregulation. These data demonstrate that haploinsufficiency for Dnmt3a alters hematopoiesis and predisposes mice (and probably humans) to myeloid malignancies by a mechanism that is not yet clear.

Authors

Christopher B. Cole, David A. Russler-Germain, Shamika Ketkar, Angela M. Verdoni, Amanda M. Smith, Celia V. Bangert, Nichole M. Helton, Mindy Guo, Jeffery M. Klco, Shelly O’Laughlin, Catrina Fronick, Robert Fulton, Gue Su Chang, Allegra A. Petti, Christopher A. Miller, Timothy J. Ley

×

Figure 6

Persistent expression of the residual WT Dnmt3a gene in AML samples arising in Dnmt3a+/– mice.

Options: View larger image (or click on image) Download as PowerPoint
Persistent expression of the residual WT Dnmt3a gene in AML samples aris...
(A) Plot demonstrating disease latency for sublethally irradiated WT animals engrafted with Dnmt3a+/– tumors, designated A–F (see Table 1 and Supplemental Table 2). For each of the 6 primary tumors that were transplanted, 3 to 5 secondary recipient mice were assessed. (B) Copy number variation in sequenced tumors. Note that none of the tumors has deletions involving chromosome 12 at the location of the Dnmt3a gene. The locations of 3 cancer-related genes that were amplified (Nras, Foxq1) or deleted (Runx1) in tumor E are shown. Tumor E was derived from a male mouse; the single copy of the X chromosome in these tumors “calibrates” the color value for a single copy deletion. (C) Representative flow cytometry plots for Dnmt3a protein abundance in the CD11b+ compartment of Dnmt3a+/+, Dnmt3a+/–, and Dnmt3a–/– bone marrow samples (top panels) and a WT spleen or Dnmt3a+/– tumors A and B (derived from the unmanipulated spleen samples from the primary mice), showing preserved expression of WT Dnmt3a protein in the CD11b+ cells in each tumor spleen sample. The level of Dnmt3a protein in the tumor cells was similar to that of the haploinsufficient bone marrow cells. (D) VAFs for selected mutations detected in primary tumors (either bulk or sorted to enrich for Gr-1+CD34+ myeloid tumor cells) and in corresponding tumors from transplanted secondary recipients. Kras mutation VAFs are from AmpliSeq data (Supplemental Table 6), while other mutation VAFs are from exome sequencing (Supplemental Tables 3–5).