The c-Myb transcription factor is a key regulator of hematopoietic cell differentiation, proliferation and apoptosis and expression of the c-myb gene is essential for normal hematopoiesis. However, a few minor mutations unleash the latent oncogenic activity of c-Myb, giving it the ability to transform hematopoietic cells in tissue culture and induce leukemias and lymphomas in chickens and mice. Furthermore, alternative RNA splicing produces at least a dozen different variants of human c-Myb protein with dramatically different activities. The alternative splicing is much more frequent in leukemias and appears to be an important mechanism for generating oncogenic variants of c-Myb in human tumors.
Specific Aim 1 will compare the transcriptional and biological activities of the wild type and variant forms of c-Myb produced by normal and leukemic cells. Wild type and variant Myb proteins will be expressed in normal CD34+ hematopoietic progenitor cells, then microarrays will detect changes in gene expression and colony formation and other cell-based assays will measure changes in proliferation, differentiation or apoptosis. By correlating these results, the relationship between specific changes in c-Myb protein structure and its transcriptional and biological functions will be determined.
Specific Aim 2 will use proven and novel models of leukemic cell transformation to directly test whether wild type and variant forms of c-Myb are able to transform immature hematopoietic cells to a leukemia-like phenotype in tissue culture. The results will be used to identify the changes in c-Myb that unleash its oncogenic activity.
Specific Aim 3 will determine how minor changes in c-Myb structure cause such dramatic changes in activity. Structure-function experiments will determine whether wild type c-Myb or variants produced by alternative RNA splicing have altered interactions with co-factors or other proteins or altered stability or sub- cellular localization. The goal will be to understand how minor changes in structure affect c-Myb activity at the biochemical level. The goal of this renewal application is to understand how relatively minor changes in the structure of the c-Myb protein can lead to such dramatic changes in its activity, from normal regulator to oncoprotein, and to characterize mechanisms that could activate the oncogenic activity of c-Myb in human tumors. ? ? ?
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|Zhou, Ye; Ness, Scott A (2011) Myb proteins: angels and demons in normal and transformed cells. Front Biosci (Landmark Ed) 16:1109-31|
|Quintana, Anita M; Liu, Fan; O'Rourke, John P et al. (2011) Identification and regulation of c-Myb target genes in MCF-7 cells. BMC Cancer 11:30|
|O'Rourke, John P; Ness, Scott A (2008) Alternative RNA splicing produces multiple forms of c-Myb with unique transcriptional activities. Mol Cell Biol 28:2091-101|
|Ness, Scott A (2007) Microarray analysis: basic strategies for successful experiments. Mol Biotechnol 36:205-19|
|Liu, F; Lei, W; O'Rourke, J P et al. (2006) Oncogenic mutations cause dramatic, qualitative changes in the transcriptional activity of c-Myb. Oncogene 25:795-805|
|Chand, Hitendra S; Ness, Scott A; Kisiel, Walter (2006) Identification of a novel human tissue factor splice variant that is upregulated in tumor cells. Int J Cancer 118:1713-20|
|Ness, Scott A (2006) Basic microarray analysis: strategies for successful experiments. Methods Mol Biol 316:13-33|
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