Epigenetic changes, such as DNA methylation and histone modifications, alter chromatin structure and regulate gene transcription. Cancer cells are characterized by abnormal DNA methylation: Repetitive DNA sequences and some gene promoters are hypomethylated and transcriptionally active, whereas many tumor suppressor gene promoters are hypermethylated and transcriptionally inactive without the presence of mutations. My laboratory discovered that cancer cells exhibit aberrant splicing of the DNMT3B gene, which encodes one of the three DNA methyltransferases. The aberrant splicing produces DNMT3B transcripts containing premature stop codons and encoding truncated proteins lacking the catalytic domain. Tissue culture cells expressing DNMT3B7, the most frequently observed aberrant DNMT3B transcript in cancer cells, show DNA methylation changes that correlate with altered gene expression. Furthermore, transgenic mice that express DNMT3B7 display disrupted embryonic development and changes in DNA methylation that are dependent on DNMT3B7 transgene levels. We hypothesize that truncated DNMT3B proteins influence DNA methylation in cancer cells, and we propose to test this idea using three Specific Aims: (1) To examine the effect of DNMT3B7 on mouse development by: (A) determining the pattern of DNMT3B7 transgene expression within embryos by in situ hybridization;and (B) examining the effects of DNMT3B7 transgene expression on DNA methylation, histone modifications, and gene expression;(2) To study the effect of DNMT3B7 expression on the DNA methylation patterns and phenotypes of cancer cells by: (A) inhibiting DNMT3B7 expression in breast cancer cells via shRNA and examining the effects on DNA methylation;(B) correlating DNMT3B7 expression with particular phenotypes in two distinctive types of neuroblastoma cell lines;and (C) quantifying DNMT3B7 levels in primary leukemia samples and correlating those with DNA methylation levels;and (3) To determine how DNMT3B7 could alter DNA methylation by: (A) testing the predictions of our models, and (B) further characterizing the interactions between DNMT3B/DNMT3B7 and three exciting interacting proteins, ZNF445, CHTF18, and NPM. Our proposed studies address the mechanism by which epigenetic alterations originate within cancer cells. The knowledge gained from the proposed work is likely to provide a basis for novel diagnostic and therapeutic strategies applicable to virtually all forms of cancer. Moreover, the pathways found to mediate the effects of DNMT3B7 are likely to reveal paradigms common to other processes that use DNA methylation to control gene expression.

Public Health Relevance

The DNA within a cell can be modified by methylation to alter its structure and affect gene expression. DNA methylation is involved in many normal cellular processes and is abnormally distributed in cancer cells, leading to some of the phenotypes of cancer cells. The mechanism by which cancer cells acquire and maintain abnormal DNA methylation is not understood. We have discovered that cancer cells express shortened forms of DNMT3B, one of the enzymes that carries out the DNA methylation reaction, and we hypothesize that truncated DNMT3B proteins contribute to abnormal DNA methylation patterns in cancer cells. The knowledge gained from the proposed experiments is likely to provide a basis for novel diagnostic and therapeutic strategies that will be applicable to virtually all forms of cancer. Furthermore, the cellular pathways found to mediate the effects of truncated DNMT3B proteins are likely to reveal paradigms common to other processes that involve DNA methylation, such as mammalian embryonic development, X-chromosome inactivation, genomic imprinting, and aging.

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Research Project (R01)
Project #
3R01CA129831-03S1
Application #
8150184
Study Section
Cancer Genetics Study Section (CG)
Program Officer
Okano, Paul
Project Start
2008-04-01
Project End
2013-02-28
Budget Start
2010-06-01
Budget End
2011-02-28
Support Year
3
Fiscal Year
2010
Total Cost
$93,600
Indirect Cost
Name
University of Chicago
Department
Internal Medicine/Medicine
Type
Schools of Medicine
DUNS #
005421136
City
Chicago
State
IL
Country
United States
Zip Code
60637
Yamazaki, Jumpei; Jelinek, Jaroslav; Lu, Yue et al. (2015) TET2 Mutations Affect Non-CpG Island DNA Methylation at Enhancers and Transcription Factor-Binding Sites in Chronic Myelomonocytic Leukemia. Cancer Res 75:2833-43
Vasanthakumar, Aparna; Zullow, Hayley; Lepore, Janet B et al. (2015) Epigenetic Control of Apolipoprotein E Expression Mediates Gender-Specific Hematopoietic Regulation. Stem Cells 33:3643-54
Vasanthakumar, Aparna; Godley, Lucy A (2015) 5-hydroxymethylcytosine in cancer: significance in diagnosis and therapy. Cancer Genet 208:167-77
Rampal, Raajit; Alkalin, Altuna; Madzo, Jozef et al. (2014) DNA hydroxymethylation profiling reveals that WT1 mutations result in loss of TET2 function in acute myeloid leukemia. Cell Rep 9:1841-1855
Madzo, Jozef; Liu, Hui; Rodriguez, Alexis et al. (2014) Hydroxymethylation at gene regulatory regions directs stem/early progenitor cell commitment during erythropoiesis. Cell Rep 6:231-244
Clozel, Thomas; Yang, ShaoNing; Elstrom, Rebecca L et al. (2013) Mechanism-based epigenetic chemosensitization therapy of diffuse large B-cell lymphoma. Cancer Discov 3:1002-19
Vasanthakumar, Aparna; Lepore, Janet B; Zegarek, Matthew H et al. (2013) Dnmt3b is a haploinsufficient tumor suppressor gene in Myc-induced lymphomagenesis. Blood 121:2059-63
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Ostler, Kelly R; Yang, Qiwei; Looney, Timothy J et al. (2012) Truncated DNMT3B isoform DNMT3B7 suppresses growth, induces differentiation, and alters DNA methylation in human neuroblastoma. Cancer Res 72:4714-23
Yamazaki, Jumpei; Taby, Rodolphe; Vasanthakumar, Aparna et al. (2012) Effects of TET2 mutations on DNA methylation in chronic myelomonocytic leukemia. Epigenetics 7:201-7

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