All ETDs from UAB

Advisory Committee Chair

Christopher A Klug

Advisory Committee Members

David Chaplin

Peter Burrows

Casey Weaver

John Kearney

Document Type

Dissertation

Date of Award

2009

Degree Name by School

Doctor of Philosophy (PhD) Heersink School of Medicine

Abstract

Hematopoiesis is a highly regulated process directed by the microenvironment or niche in the bone marrow and the transcription factors those signals activate. Gene knockout experiments have identified critical cytokine signals and transcription factors required for promoting the differentiation of a hematopoietic stem cell to a B cell, but the regulatory mechanisms have yet to be fully elucidated. In early B cell development, a transcriptional hierarchy exists wherein E2A proteins activate early B-cell factor (Ebf1), which in turn activates expression of the B cell commitment factor, Pax5. In our studies, we use IL-7 receptor alpha chain (IL-7Rα) knockout animals that are arrested in adult B cell development at the earliest B-specified stage. We found that increasing the expression level of E47 through use of a retroviral vector in IL-7Rα-deficient bone marrow is not sufficient to induce Ebf1 expression. Rather, E47 expression resulted in significant upregulation in the mRNA levels of inhibitors of differentiation proteins, Id2 and Id3. In contrast, enforced expression of Ebf1 in IL-7Rα-/- bone marrow cells restores B cell differentiation in vivo, potently downregulates expression of both Id2 and Id3 mRNA, and it further upregulates the expression of E47 mRNA. This suggests that one major function of Ebf1 in B-lineage specification is to down-regulate Id levels to allow for activation of E2A protein activity. Additionally, we investigated the role of cytokine signaling in regulating the induction of Ebf1 expression. Our studies show a correlation between down-regulation of expression of c-Kit and Flt3 receptors from the cell surface and developmental progression from the common lymphoid progenitor (CLP) to the earliest B cell progenitor in adult bone marrow. Additionally, in vitro stimulation of the receptors with recombinant SCF or Flt3L was sufficient to block IL-7 induction of Ebf1 expression. Therefore, c-Kit and Flt3 are responsible for inhibiting the B cell promoting activity of IL-7 signaling and for maintaining the full lymphoid potential of common lymphoid progenitors. Collectively, these studies suggest a model in which the balance between positive and negative environmental cues determines maintenance of the lymphoid progenitor population versus Ebf1 induction and B cell specification.

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