The goal of this competing renewal is to further our elucidation of the molecular events leading to myeloid leukemic cell differentiation in response to the hormone 1,25 dihydroxyvitamin D3 [1 ,25(OH)2D3}. We will focus on two vitamin D receptor (VDR? target genes central to the induction of growth arrest and differentiation in the myeloid cell system: p21waf1,cip, and HoxAlO. In addition, myeloid differentiation will be used as cellular readout for the 1 ,25(OH)2D3 response in the context of nuclear receptor coactivator function, as well as when VDR function is attenuated by chromosomal translocations involving the RARa gene.
The specific aims during the proposed grant period are to: 1. Characterize the complex regulation of the p21 promoter by VDR and HoxAlO, A cluster of multiple VDr and HoxAlO binding sites in the p21 promoter individually and collectively contribute to 1,25(OH)2D3 regulation of this promoter. Nucleosomal structure and in viva occupancy of these regions will be examined in response to 1,25(OH)2D3 and HoxAlO. 2.Define the role of HoxAlO in growth arrest and cell differentiation. HoxAlO regulation of the p21 promoter, and its possible role as a transcriptional modulator of other cell cycle regulator genes, will be addressed. A screen for novel HoxAlO target genes using array technology will also be generated. 3. Describe the molecular basis for PLZF and PLZF-RARcx attenuation of 1,25 (OH)2D3-inducible myeloid differentiation. The mechanisms by which 1 ,25(OH)2D3-dependent differentiation is attenuated by PLZF and PLZFRARa will be characterized at the molecular level. 4. Utilize cell differentiation as a biological read-out for coactivator function. The functional requirement of the DRIP coactivator complex in mediating VDR and RAR transactivation, in the context of nuclear receptor-dependent myeloid differentiation, will be determined.
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