REGULATING MITF'S ACTIVATION OF OSTEOCLAST TARGET GENES

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DESCRIPTION (provided by applicant): Microphthalmia-associated transcription factor (Mitf) and Tfe3 are basic helix-loop-helix leucine zipper transcription factors that are expressed in osteoclasts. We have demonstrated that Mitf and Tfe3 colloborate to activate genes important for osteoclast differentiation. Evidence that Mitf and Tfe3 are necessary for osteoclast differentiation is demonstrated by the fact that mice null for expression of Mitf and Tfe3 are osteopetrotic. Using a yeast two hybrid screen, we identified Cdc25C-associated kinase 1 (C-TAK1) and POH1, a component of the proteasome lid, as interacting with the amino terminus of Mitf. This proposal attempts to 1) determine how Mitf is translocated to the nucleus in osteoclasts and 2) to identify proteins that affect Mitfs stability in osteoclasts. To define the interaction between C-TAK1 and Mitf (aim 1), we will determine which amino acid residue of Mitf is phosphorylated by C-TAK1, determine which Mitf amino acid residues are critical for binding to 14-3-3 proteins and determine what effect C-TAK1 and 14-3-3 interaction has on Mitfs cellular location. 14-3-3 are a family of proteins that recognize specific phosphorylated serines and bind to and sequester a protein in the cytoplasm. To determine the effect of Mitf and POH1's interaction (aim 2), we will determine if Mitf's stability is affected by CSF-1 and RANKL signaling, map the region of Mitf that interacts with POH1 and determine the effect of POH1's interaction with Mitf on Mitfs activation of osteoclast target genes. By understanding how Mitf activates ostoeclast specific genes, drug targets may be suggested that will lead to therapies to control the differentiation and activation of osteoclasts.