Genetic mechanisms regulating B-lymphocyte commitment

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DESCRIPTION (provided by applicant): The pathways and molecular mechanisms responsible for B lymphocyte commitment from oligopotent common lymphoid progenitor cells remain largely unknown. Recent studies have suggested that the transcription factors, early B cell factor (EBF) and Pax5, may play a significant role in regulating both the initiation and maintenance of the B cell lineage. In mice that develop in the absence of EBF or Pax5, B cell development is arrested prior to lineage commitment (EBF) or the maintenance of the B committed state cannot be sustained (Pax5). Findings that EBF can directly bind and activate the Pax5 promoter suggest that these two factors may act in a common regulatory cascade that controls B cell fate. This notion is supported by studies described in this proposal, where enforced expression of either EBF or Pax5 in hematopoietic stem cells is sufficient to determine B versus T cell identity in vivo. All animals reconstituted with cells expressing EBF or Pax5 exhibit greater than 90 percent donor-derived B-lineage cells in the peripheral blood and spleen for at least 10 months post-transplant, with essentially no T lymphopoiesis occurring in the thymus. The strong skew in development to the B cell lineage was not due to malignancy and was specific for expression of either EBF or Pax5. The overlapping phenotypes further suggest that EBF and Pax5 may function in complementary pathways to control the earliest events in B-lineage determination. To test this hypothesis, we propose to (1) map domains within EBF and Pax5 that are responsible for both promotion of B cell development and inhibition of T-lineage commitment, (2) identify downstream target genes of EBF that activate B cell development in the context of EBF-/- animals, (3) define whether EBF and/or Pax5 are sufficient to rescue B cell development in genetic backgrounds deficient in PU. 1, Ikaros, or E2A and (4) determine whether signaling through the Notch1 pathway influences the expression or activity of EBF and/or Pax5. These studies will provide insights into the regulatory cascades that control the earliest stages of B cell development at the molecular level.