Hematopoiesis proceeds from a bone marrow resident population of stem cells responsible for generation of all lineages within the blood. Distinct molecular programs within hematopoietic stem cells regulate maintenance of this population under homeostatic conditions, however the coordination of these programs remains largely undefined. DNA methylation is an epigenetic means of gene regulation in a mammals carried out by a family of DNA methyltransferases. Of these, the de novo methyltransferase is highly expressed in hematopoietic stem cells as compared to other members of this family and somatic mutations in DNMT3b lead to a syndrone characterized by immunodeficiency. Therefore we hypothesized that DNMT3b regulates hematopoietic stem cells via its ability to methylate DNA. By knock-down of DNMT3b with a retrovirally delivered shRNA or Cre mediated recombination of floxed DNMT3b alleles work presented in this thesis demonstrates a critical role for DNMT3b in hematopoiesis in mice. Loss of DNMT3b leads to limited reconstitution of hematopoiesis in irradiated recipients associated with a proliferative defect in vitro and a failure of hematopoietic stem cell self-renewal in vivo. Targeted deletion of DNMT3b in hematopoietic stem cells leads to decreased engraftment following transplantation and decreased proliferation in vitro. DNMT3bs function in hematopoietic cells requires the methyltransferase activity of the enzyme and the defects in hematopoiesis are associated with loss of DNA methylation and decreased expression of MLL. Therefore DNMT3b is necessary for maintenance of the proliferative ability and engraftment capacity of hematopoietic cells and hematopoiesis is dependent upon appropriate DNA methylation.
| Identifer | oai:union.ndltd.org:PITT/oai:PITTETD:etd-12032010-144803 |
| Date | 14 December 2010 |
| Creators | Boyer, Matthew Jacob |
| Contributors | Kyle Orwig, Richard Chaillet, Laura Niedernhofer, Eric Lagasse, Tao Cheng |
| Publisher | University of Pittsburgh |
| Source Sets | University of Pittsburgh |
| Language | English |
| Detected Language | English |
| Type | text |
| Format | application/pdf |
| Source | http://etd.library.pitt.edu/ETD/available/etd-12032010-144803/ |
| Rights | restricted, I hereby certify that, if appropriate, I have obtained and attached hereto a written permission statement from the owner(s) of each third party copyrighted matter to be included in my thesis, dissertation, or project report, allowing distribution as specified below. I certify that the version I submitted is the same as that approved by my advisory committee. I hereby grant to University of Pittsburgh or its agents the non-exclusive license to archive and make accessible, under the conditions specified below, my thesis, dissertation, or project report in whole or in part in all forms of media, now or hereafter known. I retain all other ownership rights to the copyright of the thesis, dissertation or project report. I also retain the right to use in future works (such as articles or books) all or part of this thesis, dissertation, or project report. |
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