The long-term objectives of this project are to determine the mechanisms by which chromosome rearrangements activate the expression of the EVI1 gene in leukemic blasts from cases of acute myelogenous leukemia (AML). One of the most common events in murine myeloid leukemia is the activation, by retroviral insertions, of the expression of the ecotropic virus integration-1 gene (Evi-1) which encodes a zinc finger transcriptional factor. The Evi-1 gene is not normally expressed in myeloid cells and, when aberrantly expressed, can block their differentiation. During the past granting period we have shown that leukemic blasts from approximately 5% of cases of AML aberrantly express the EVI1 gene and have rearrangements of chromosome band 3q26 where the EVI1 gene resides. The most commonly involved region is 3q21 in t(3;3)(q21;q26) and inv(3)(q21q26). The rearrangements are detectable over several hundred kilobases, both 5' and 3' of the EVI1 locus and one has been cloned. The proposed studies will address the hypothesis that activation of EVI1 gene expression is due to introduction of sequences that affect chromatin structure over large distances. this hypothesis will be tested through two specific aims. The first specific aim will molecularly characterize the EVI1 gene and flanking regions on 3q26. We will determine the complete structure of the EVI1 gene and will clone flanking regions. Additional breakpoints will be cloned to determine whether sequence similarities exist that would indicate the mechanisms mediating the rearrangements. To determine whether chromatin structure surrounding the EVI1 gene is affected, we will look for changes in DNase I hypersensitivity and DNA methylation. Because the rearrangements map over large regions surrounding the EVI1 gene, the rearrangements may affect genes flanking the EVI1 gene. Identification of such genes may indicate the distances over which rearrangements can affect gene expression. The second specific aim will molecularly characterize the 3q21 region. We will clone regions of 3q21 and map the sites of rearrangements. To test the hypothesis that this region contains elements that confer high levels of expression in myeloid cells, we will look for genes on 3q21. Regions that might contact transcriptional control elements will be identified and will be examined for their biological activity constructs containing the EVI1 promoter either in tissue culture cells or in vivo in transgenic mice. Collectively, these studies will contribute to our understanding of the mechanisms by which chromosomal rearrangements can activate gene expression that result in cellular transformation.
Showing the most recent 10 out of 613 publications
