The promyelocytic leukemia zinc finger (PLZF) protein is a transcription factor, expressed in hematopoietic progenitor cells, fused to the retinoic acid receptor-alpha (RAR alpha) in t(11;17)-associated acute promyelocytic leukemia (APL). Over the past 9 years, through two periods of funding, our group characterized t(11;17) APL as a distinct syndrome, unresponsive to retinoic acid. We determined that the PLZF-RARalpha fusion generated in t(11;17) is a dominant negative form of PAR that actively recruits corepressors and histone deacteylase molecules to RAR target genes. The study of the PLZF fusion protein helped solidify the model of aberrant transcriptional repression as a pathogenic basis of leukemia. Though progress has been gratifying, many questions remain. The nature of the critical target genes of the retinoid receptor blocked by the fusion proteins of APL is not certain. The way in which genes are repressed is incompletely understood. Histone deacetylases are critically involved but other modes of chromatin modification, chromatin remodeling and epigenetic silencing of repressed genes are likely. The PLZF protein represses through a number of co-repressors attracted though the BTB/POZ Domain. Further structure of this domain will yield further insights and potential therapeutic modalties in the disease. APL in animal models occurs after a considerable delay, indicating that other mutations are required for the disease to occur. One such cooperating mutation may be the mutation of the fit3 receptor tyrosine kinase molecule. One mode of cooperation may be the ability of the APL fusion proteins to abrogate the p53 pathway and prevent premature cellular senescence in response to activation of ras/map kinasse pathways. PML is a modulator of p53 function and PLZF may be as well. The proposed research will: 1. Determine of how PLZF controls myeloid cell growth and differentiation by elucidation of PLZF target genes which bind the PLZF protein in vitro and in vivo such as IL-6, cyclin A and other to be identified by whole genome PCR. 2. Define how an evolutionarily conserved protein motif, the POZ domain, functions in transcriptional regulation, though mutagensis of conserved residues and identification of partner proteins using the yeast two hybrid system. 3. Define protein-protein interaction networks that play a role in normal myelopoiesis and leukemogenesis (PML-PLZF, N-Cor-PLZF) 4. Extend knowledge of gene regulation in early hematopoiesis through characterization of the cis-acting sequences controlling expression of PLZF.
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