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ORCID

http://orcid.org/0000-0002-0968-2947

Date of Award

Winter 12-15-2019

Author's School

Graduate School of Arts and Sciences

Author's Department

Biology & Biomedical Sciences (Human & Statistical Genetics)

Degree Name

Doctor of Philosophy (PhD)

Degree Type

Dissertation

Abstract

Clonal hematopoiesis of indeterminate potential (CHIP) is near ubiquitous in healthy, aging adults. Genetic variants associated with CHIP are commonly located within the epigenetic regulators DNA (cytosine-5-)-methyltransferase 3 alpha (DNMT3A) and tet methylcytosine dioxygenase 2 (TET2). DNMT3A and TET2 are also recurrently mutated in a range of hematological diseases, and mutations in these enzymes occurring at the hematopoietic stem cell (HSC) level are initiating events preceding progression to disorders such as myelodysplastic syndromes (MDS). Additionally, murine HSCs genetically-deficient for Dnmt3a (Dnmt3aKO) or Tet2 (Tet2KO) demonstrate increased self-renewal and impaired differentiation when stressed through serial transplantation. Together, these observations suggest that outgrowth of a dominant clone are not due to stochastic events but result from dysfunction of Dnmt3a or Tet2 conferring an enhanced fitness to HSCs. Using conditional ablation, we show loss of function of these opposing epigenetic enzymes results in different functional effects in hematopoietic stem cells (HSCs) despite similar disease phenotypes. Dnmt3a deficiency bestows enhanced self-renewalon HSCs in serial, competitive repopulation assays while Tet2 loss functionally depletes HSCs after a tertiary transplant despite an initial competitive advantage. Molecular characterization of transcriptomes and chromatin accessibility reflects the functional differences between the genotypes as Dnmt3a-null cells reside in a more stem cell-like state while Tet2 loss leads to functional attrition of down-stream progenitors. Collectively the data suggests loss of Dnmt3a imparts enhanced self-renewal while loss of Tet2 conditions HSCs to persist following replicative stress. One common source of such replicative stress is the inflammatory response associated with infection. The cytokines responsible for altering bone marrow (BM) homeostasis to mount an immune response also effect HSC function. One such cytokine, interferon gamma (IFNγ), promotes both stem cell division and differentiation, while chronic IFNγ signaling precipitates HSC exhaustion and contributes to BM failure syndromes. Using competitive transplants we demonstrate that deprivation of IFNγ signaling at the basal level significantly impedes the outgrowth of Dnmt3a-null HSCs. Chronic exposure to inflammation through interferon signaling also impedes clonal expansion through functional attrition but not as significantly as wild-type HSCs are affected. Transcriptome analysis revealed a downregulation of targets of the transcription factor family E2F. Suppression of E2F activity promotes quiescence which functions to preserve stem cell state over an individual’s lifetime thereby providing an avenue for clonal expansion. Together these data demonstrate the both cell-intrinsic and –extrinsic dynamics shape clonal expansion. Environmental forces such as pro-inflammatory signaling can provide a selective pressure for the outgrowth of mutant stem cell clones while altered function resulting from DNMT3A and TET2 mutations lead HSCs down different molecular and functional pathways despite similar disease destinations.

Language

English (en)

Chair and Committee

Grant Challen

Committee Members

Daniel Link, Jeffrey Magee, Shelia Stewart, Laura Schuettpelz,

Comments

Permanent URL: https://doi.org/10.7936/ytxh-gy03

Available for download on Friday, December 15, 2119

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