Hematopoiesis is characterized by incessant production of a variety of cell types from hematopoietic stem cells (HSC). During this process, HSC generate progenitor populations with progressively reduced lineage potentials. We have directly shown the existence of such progenitor populations by prospective purification on a fluorescence-activated cell sorter. These include the earliest lymphoid-committed progenitors (common lymphoid progenitor; CLP), the earliest myeloid-committed progenitors (common myeloid progenitors: CMP), the granulocyte/monocyte progenitor (GMP), and the megakaryocyte/ erythrocyte progenitor (MEP). Using these prospectively-purified progenitor populations, we will evaluate whether cytokines play instructive as well as permissive roles in lineage determination. In this proposal, we will focus on three important branch points of hematopoiesis; 1) CLP vs. CMP development from HSC, 2) MEP vs. GMP development from CMP, and 3) eosinophil, mast cell, and neutrophil development from GMP. First, the physiological roles of cytokines in hematopoiesis will be evaluated at the progenitor level. We will analyze coexpression of various cytokine receptors in progenitors by using single cell RT-PCR systems. Then we will evaluate the effect of cytokine depletion on differentiation potentials of each progenitor subset. We will evaluate biological characteristics of progenitors isolated from mice deficient for myeloid and lymphoid-related cytokines. Next, we will evaluate whether ectopic IL-7R signals can reprogram myeloid progenitors to transdifferentiate into lymphoid lineages, whether ectopic GM-CSFR or G-CSFR signals can transdifferentiate lymphoid progenitors into myeloid lineage, and whether transgenic IL-5R signals can instruct GMP to differentiate into eosinophils. This system will also be used to evaluate the effect of oncogenic tyrosine kinase fusions on lineage determination, whose signals considerably resemble those of cytokines. We will retrovirally transduce BCR-ABL and TEL-PTK fusions into progenitors to test whether lineage conversion occurs after the transduction. We hope that this study will help elucidate the mechanisms of lineage commitment in normal and malignant hematopoiesis.
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