15 research outputs found
DNMT3A Loss Drives Enhancer Hypomethylation in FLT3-ITD-Associated Leukemias.
DNMT3A, the gene encoding the de novo DNA methyltransferase 3A, is among the most frequently mutated genes in hematologic malignancies. However, the mechanisms through which DNMT3A normally suppresses malignancy development are unknown. Here, we show that DNMT3A loss synergizes with the FLT3 internal tandem duplication in a dose-influenced fashion to generate rapid lethal lymphoid or myeloid leukemias similar to their human counterparts. Loss of DNMT3A leads to reduced DNA methylation, predominantly at hematopoietic enhancer regions in both mouse and human samples. Myeloid and lymphoid diseases arise from transformed murine hematopoietic stem cells. Broadly, our findings support a role for DNMT3A as a guardian of the epigenetic state at enhancer regions, critical for inhibition of leukemic transformation.L.Y. is funded by the Robert and Janice McNair Foundation as an MD/PhD McNair Scholar. This project was funded by CPRIT (RP110028, RP110471 and RP150292 ), the NIH (DK092883 and HG007538), and the Samuel Waxman Cancer Research Foundation. We also thank the Cytometry and Cell Sorting and Genomic and RNA Profiling Cores (NCI P30CA125123, P30 AI036211, P30 CA125123, and S10 RR024574 ) at Baylor College of Medicine. Authors declare no conflicts of interest.This is the author accepted manuscript. The final version is available from Cell Press via http://dx.doi.org/10.1016/j.ccell.2016.05.00
Abrupt climate change as an important agent of ecological change in the Northeast U.S. throughout the past 15,000 years
Author Posting. © The Author(s), 2009. This is the author's version of the work. It is posted here by permission of Elsevier B.V. for personal use, not for redistribution. The definitive version was published in Quaternary Science Reviews 28 (2009): 1693-1709, doi:10.1016/j.quascirev.2009.04.005.We use a series of tests to evaluate two competing hypotheses about the association of climate
and vegetation trends in the northeastern United States over the past 15 kyrs. First, that abrupt
climate changes on the scale of centuries had little influence on long-term vegetation trends,
and second, that abrupt climate changes interacted with slower climate trends to determine the
regional sequence of vegetation phases. Our results support the second. Large dissimilarity
between temporally-close fossil pollen samples indicates large vegetation changes within 500
years across >4° of latitude at ca. 13.25-12.75, 12.0-11.5, 10.5, 8.25, and 5.25 ka. The evidence of
vegetation change coincides with independent isotopic and sedimentary indicators of rapid
shifts in temperature and moisture balance. In several cases, abrupt changes reversed long-term
vegetation trends, such as when spruce (Picea) and pine (Pinus) pollen percentages rapidly
declined to the north and increased to the south at ca. 13.25-12.75 and 8.25 ka respectively.
Abrupt events accelerated other longâterm trends, such as a regional increase in beech (Fagus)
pollen percentages at 8.5-8.0 ka. The regional hemlock (Tsuga) decline at ca. 5.25 ka is unique
among the abrupt events, and may have been induced by high climatic variability (i.e., repeated
severe droughts from 5.7-2.0 ka); autoregressive ecological and evolutionary processes could
have maintained low hemlock abundance until ca. 2.0 ka. Delayed increases in chestnut
(Castanea) pollen abundance after 5.8 and 2.5 ka also illustrate the potential for multi-century
climate variability to influence speciesâ recruitment as well as mortality. Future climate changes
will probably also rapidly initiate persistent vegetation change, particularly by acting as broad,
regional-scale disturbances.This work was supported by NSF grants to B. Shuman (EARâ0602408; DEBâ0816731) and J.
Donnelly (EARâ0602380)
Highly Efficient Genome Editing of Murine and Human Hematopoietic Progenitor Cells by CRISPR/Cas9
Summary: Our understanding of the mechanisms that regulate hematopoietic stem/progenitor cells (HSPCs) has been advanced by the ability to genetically manipulate mice; however, germline modification is time consuming and expensive. Here, we describe fast, efficient, and cost-effective methods to directly modify the genomes of mouse and human HSPCs using the CRISPR/Cas9 system. Using plasmid and virus-free delivery of guide RNAs alone into Cas9-expressing HSPCs or Cas9-guide RNA ribonucleoprotein (RNP) complexes into wild-type cells, we have achieved extremely efficient gene disruption in primary HSPCs from mouse (>60%) and human (âŒ75%). These techniques enabled rapid evaluation of the functional effects of gene loss of Eed, Suz12, and DNMT3A. We also achieved homology-directed repair in primary human HSPCs (>20%). These methods will significantly expand applications for CRISPR/Cas9 technologies for studying normal and malignant hematopoiesis. : Gundry et al. develop an efficient and simple method implementing CRISPR/Cas9-mediated gene disruption and HDR in murine and human HSPCs. This method enables quick evaluation of the function of genes by performing in vitro or transplantation assays using the modified HSPCs. Keywords: HSC, hematopoietic stem cells, progenitor, human CD34, genome editing, CRISPR/Cas9, sgRNA, homology-directed repair, gene therapy, transplantatio
DOT1L as a therapeutic target for the treatment of DNMT3A-mutant acute myeloid leukemia
Mutations in DNA methyltransferase 3A (DNMT3A) are common in acute myeloid leukemia and portend a poor prognosis; thus, new therapeutic strategies are needed. The likely mechanism by which DNMT3A loss contributes to leukemogenesis is altered DNA methylation and the attendant gene expression changes; however, our current understanding is incomplete. We observed that murine hematopoietic stem cells (HSCs) in which Dnmt3a had been conditionally deleted markedly overexpress the histone 3 lysine 79 (H3K79) methyltransferase, Dot1l. We demonstrate that Dnmt3a(-/-) HSCs have increased H3K79 methylation relative to wild-type (WT) HSCs, with the greatest increases noted at DNA methylation canyons, which are regions highly enriched for genes dysregulated in leukemia and prone to DNA methylation loss with Dnmt3a deletion. These findings led us to explore DOT1L as a therapeutic target for the treatment of DNMT3A-mutant AML. We show that pharmacologic inhibition of DOT1L resulted in decreased expression of oncogenic canyon-associated genes and led to dose- and time-dependent inhibition of proliferation, induction of apoptosis, cell-cycle arrest, and terminal differentiation in DNMT3A-mutant cell lines in vitro. We show in vivo efficacy of the DOT1L inhibitor EPZ5676 in a nude rat xenograft model of DNMT3A-mutant AML. DOT1L inhibition was also effective against primary patient DNMT3A-mutant AML samples, reducing colony-forming capacity (CFC) and inducing terminal differentiation in vitro. These studies suggest that DOT1L may play a critical role in DNMT3A-mutant leukemia. With pharmacologic inhibitors of DOT1L already in clinical trials, DOT1L could be an immediately actionable therapeutic target for the treatment of this poor prognosis disease