Inhibition of intestinal tumor formation by deletion of the DNA methyltransferase 3a.
Aberrant de novo methylation of DNA is considered an important mediator of tumorigenesis. To investigate the role of de novo DNA methyltransferase 3a (Dnmt3a) in intestinal tumor development, we analyzed the expression of Dnmt3a in murine colon crypts, murine colon adenomas and human colorectal cancer using RNA fluorescence in situ hybridization (FISH), quantitative PCR and immunostaining. Following conditional deletion of Dnmt3a in the colon of APC((Min/+)) mice, we analyzed tumor numbers, genotype of macroadenomas and laser dissected microadenomas, global and regional DNA methylation and gene expression. Our results showed increased Dnmt3a expression in colon adenomas of APC((Min/+)) mice and human colorectal cancer samples when compared with control tissue. Interestingly, in tumor tissue, RNA FISH analysis showed highest Dnmt3a expression in Lgr5-positive stem/progenitor cells. Deletion of Dnmt3a in APC((Min/+)) mice reduced colon tumor numbers by ~40%. Remaining adenomas and microadenomas almost exclusively contained the non-recombined Dnmt3a allele; no tumors composed of the inactivated Dnmt3a allele were detected. DNA methylation was reduced at the Oct4, Nanog, Tff2 and Cdkn1c promoters and expression of the tumor-suppressor genes Tff2 and Cdkn1c was increased. In conclusion, our results show that Dnmt3a is predominantly expressed in the stem/progenitor cell compartment of tumors and that deletion of Dnmt3a inhibits the earliest stages of intestinal tumor development.
Division of Epigenetics (A130), German Cancer Research Center (DKFZ), Heidelberg, Germany.
B
B
Weis
Division of Epigenetics (A130), German Cancer Research Center (DKFZ), Heidelberg, Germany.
J
J
Schmidt
Department of Bioengineering, University of Pennsylvania, Philadelphia, PA, USA.
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H
Maamar
Department of Bioengineering, University of Pennsylvania, Philadelphia, PA, USA.
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A
Raj
Department of Ophthalmology, Massachusetts Eye and Ear Infirmary, Harvard Medical School, Boston, MA, USA.
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H
Lin
1] Department of Pathology, University of Heidelberg, National Center for Tumor Diseases (NCT) Tissue Bank, Heidelberg, Germany [2] Department of Pathology, Heinrich Heine University, Düsseldorf, Germany.
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C
Tóth
Division of Epigenetics (A130), German Cancer Research Center (DKFZ), Heidelberg, Germany.
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K
Riedmann
Division of Epigenetics (A130), German Cancer Research Center (DKFZ), Heidelberg, Germany.
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G
Raddatz
Department of Medicine, Salem Medical Center, Alcohol Research Center, University of Heidelberg, Heidelberg, Germany.
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HK
Seitz
Department of Hematology/Oncology, University of Heidelberg Medical Center, Heidelberg, Germany.
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AD
Ho
Division of Epigenetics (A130), German Cancer Research Center (DKFZ), Heidelberg, Germany.
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F
Lyko
1] Department of Medicine, Salem Medical Center, Alcohol Research Center, University of Heidelberg, Heidelberg, Germany [2] Department of Hematology/Oncology, University of Heidelberg Medical Center, Heidelberg, Germany [3] Division of Epigenetics (A130), German Cancer Research Center (DKFZ), Heidelberg, Germany.
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Linhart
D000236:Adenoma / Q000235:genetics* / Q000473:pathology; D000818:Animals; D002471:Cell Transformation, Neoplastic / Q000235:genetics; D003106:Colon / Q000378:metabolism / Q000473:pathology; D015179:Colorectal Neoplasms / Q000235:genetics* / Q000473:pathology; D050761:Cyclin-Dependent Kinase Inhibitor p57 / Q000096:biosynthesis / Q000235:genetics; D004248:DNA (Cytosine-5-)-Methyltransferases / Q000096:biosynthesis / Q000235:genetics*; D019175:DNA Methylation / Q000235:genetics*; D000090205:DNA Methyltransferase 3A; D015972:Gene Expression Regulation, Neoplastic; D018398:Homeodomain Proteins / Q000235:genetics; D006801:Humans; D017404:In Situ Hybridization, Fluorescence; D051379:Mice; D008810:Mice, Inbred C57BL; D008822:Mice, Transgenic; D009077:Mucins / Q000096:biosynthesis / Q000235:genetics; D009124:Muscle Proteins / Q000096:biosynthesis / Q000235:genetics; D000071317:Nanog Homeobox Protein; D014411:Neoplastic Stem Cells / Q000473:pathology*; D050814:Octamer Transcription Factor-3 / Q000235:genetics; D010455:Peptides / Q000235:genetics; D011401:Promoter Regions, Genetic; D060888:Real-Time Polymerase Chain Reaction; D000071164:Trefoil Factor-2
C493965:Cdkn1c protein, mouse; D050761:Cyclin-Dependent Kinase Inhibitor p57; C000717967:DNMT3A protein, human; C000715535:Dnmt3a protein, mouse; D018398:Homeodomain Proteins; D009077:Mucins; D009124:Muscle Proteins; D000071317:Nanog Homeobox Protein; C475540:Nanog protein, mouse; D050814:Octamer Transcription Factor-3; D010455:Peptides; C494174:Pou5f1 protein, mouse; C000604007:TFF2 protein, human; C481546:TFF2 protein, mouse; D000071164:Trefoil Factor-2; D004248:DNA (Cytosine-5-)-Methyltransferases; D000090205:DNA Methyltransferase 3A