The E(-ret mouse carries a transgene that produces a chimeric protein, Rfp-Ret with constitutive tyrosine kinase activity in B lineage cells. Adult E(-ret mice develop B precursor leukemias that appear to arise from the progeny of fetal-derived pro B cells. Furthermore, the leukemias are resistant to stimuli that induce apoptosis in the pro B cells from fetal and young E(-ret mice. The applicant hypothesizes that the Rfp-Ret product selectively enhances the proliferation and survival of fetal-derived pro B cells, and that transformation is dependent on the accumulation of other mutations which further enhance these growth properties. The goals of this proposal are to more thoroughly characterize the multistep process of leukemogenesis in the E(-ret mouse.
In aim 1, he will establish whether fetal and adult pro B cells differ in their susceptibility to transformation by the Rfp-Ret transgene product. He will transfer fetal and adult E(-ret pro B cells into Rag-1- mice to determine whether these populations differ in their ability to induce leukemia.
In aim 2, he will establish whether Rfp-Ret activity mimics the effect of bone marrow stromal contact. He will determine whether pro B cells from Rag-1- mice acquire the ability to grow in IL-7 without stromal cell contact when they express the Rfp-Ret product (E(-ret/Rag-1-), and will determine whether Rfp-Ret enhances the proliferative effect of IL-7.
In aim 3, he will establish whether the development of interferon (IFN)-( resistance in pro B cells allows for malignant outgrowth. He will isolate IFN-( resistant cells from within the late pro B cell populations of healthy adult E(-ret mice, and sequence the rearranged immunoglobulin heavy chain loci of the IFN-( resistant cells to determine whether these cells represent a polyclonal (nontransformed) or clonal (transformed) outgrowth.
In aim 4, he will establish whether the level of Rfp-Ret activity increases during the transformation process. He will immunoprecipitate Rfp-Ret from the pro B cell populations of fetal and healthy adult E(-ret mice as well as from leukemias and compare the levels of tyrosine phophorylation of Rfp-Ret and Rfp-Ret-bound proteins.
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