ROLE OF PU1 IN HEMATOPOIETIC PROGENITOR CELL DIFFERENTIATION

University Of Pennsylvania, Philadelphia PA

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Abstract

Description (taken directly from the application): Hematopoiesis is a regulated developmental cascade that generates at least eight distinct lineages which differentiate into the mature cell types of the blood and lymph. Hematopoiesis is an accessible model system in mammals to study the development of distinct cell lineages. However, the regulatory pathways that control hematopoiesis are as yet poorly understood. The proto-oncogene PU.1 is a member of the Ets-family of transcription factors and has been suggested to be a regulator of hematopoiesis based on its restricted expression pattern (primarily in B-cells and monocytes) and by identification of many presumptive target genes in the B-lymphoid and monocyte/macrophage lineages. Inappropriate expression of PU.1 mediated by retroviral integration can result in erythroleukemia, or deregulated polyclonal B-cell proliferation resulting in an immunodeficiency syndrome in mice. To understand the function of PU.1 in the mouse, the gene was targeted via homologous recombination in embryonic stem (ES) cells and "knock-out" mice were generated. Analysis of the hematopoietic system revealed that lymphoid and myeloid development was completely abrogated in mutant embryos, suggesting the existence of a PU.1-dependent lymphoid/myeloid multipotent progenitor (Scott et al., 1994). In vitro clonogenic and radioprotection assays with PU.1-/- hematopoietic cells have demonstrated that the defect in myeloid and lymphoid development is cell intrinsic and not due to a defect in the fetal liver microenvironment. The myeloid differentiative capacity of PU.1-/- ES cells can be rescued with a transgene which expresses PU.1 under the control of its own promoter, and PU.1 transactivation can be modulated by fusing PU.1 and the estrogen receptor (Olson et al., 1995 and preliminary results). In order to further our understanding of the regulatory role that PU.1 plays during fetal and adult hematopoiesis, we propose the following Specific Aims: Aim I. To characterize the effect of the PU.1 mutation on hematopoietic progenitor cell populations in vivo. Aim II. Complement the PU.1-/- mutation with a PU.1::Estrogen Receptor fusion. Aim III. Determine the role of PU.1 in the later stages of lymphoid and myeloid development.