Neoplastic transformation is a multistep process that requires the deregulation of key cellular pathways that control the expansion and the differentiation of progenitor cells. The EVI1 oncogene is inappropriately activated in many tissues resulting in aggressive solid and hematopoietic cancers. The mechanism by which EVI1 operates in the transformation of tissues of different histology is not clear. The overall goal of the proposal that was funded 3 years ago was to determine how EVI1 confers to the cell the ability to escape the control of several cellular pathways including differentiation and proliferation and how it contributes to cell transformation. These studies have shown that EVI1 interacts with master regulatory proteins to disrupt cell cycling, cell proliferation, and normal differentiation. During the prior funding cycle, we have also determined that EVI1 interacts with several transcription coregulators and protein modifiers, which post-translationally modify EVI1 by reversible acetylation or sumoylation. More importantly, by using point mutations we have established that specific interactions with these modifiers are required to alter proliferation and differentiation of primary normal cells by EVI1 and to induce immortalization or transformation. Therefore, we propose that the ability of EVI1 to disrupt multiple pathways is the basis of the very aggressive phenotype characteristic of EVI1-positive cancers. We also propose that EVI1 must be post-translationally modified in order to interact with key cell regulators and induce cell transformation. We determined that EVI1 modifications are required for the assembly of EVI1 in discrete nuclear speckles and we propose that these speckles are centers of EVI1 activity. Therefore, it is likely that disruption of EVI1 modification will impair the ability of EVI1 to alter at least one of these pathways, leading to attenuated or delayed neoplastic transformation. This competing renewal has two overall goals: to identify and analyze multi-protein complexes in which EVI1 participates and to determine which protein-modifications are required to alter proliferation and differentiation.
The specific aims of the proposal are: 1. To characterize EVI1 sumoylation and acetylation and to define their role in transcription regulation. 2. To identify the components and the functions of the nuclear EVI1 multiprotein complexes. 3. To characterize the sumoylation and acetylation of EVI1 in vivo and to define their role in EVI1- induced transformation.
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