MI: MODULATING OSTEOCLAST GENE EXPRESSION AND FUNCTION

描述(申请人提供)：微眼炎转录因子(MITF)是包括破骨细胞、肥大细胞、有色视网膜上皮细胞和黑素细胞在内的发育无关细胞类型的终末分化所必需的，并调节这些细胞类型中不同的靶基因。在破骨细胞的终末分化过程中，MITF调控一系列骨吸收所需的基因，包括组织蛋白酶K(CTSK)、氯离子通道Ccln7、Ostm1/Grey Laythal和酒石酸抗性酸性磷酸酶/酸性磷酸酶5(Trap/Acp5)。这组基因的突变与人类和小鼠的破骨细胞功能障碍和骨骼疾病有关，有力地证明了MITF在破骨细胞终末分化过程中的基因调控中发挥着核心作用。我们的工作表明，MITF和ETS家族因子PU.1之间的相互作用对于选择性调节破骨细胞中的这一组靶基因是必要的。在破骨细胞分化过程中，MITF也是CSF-1/RANKL信号转导的直接靶点，由丝裂原活化蛋白激酶(MAPK)、ERK和p38直接激活。在目前的授权期，我们的工作揭示了相互作用的蛋白质，表明了ITF/PU.1在破骨细胞中潜在的作用机制。出乎意料的是，在承诺的破骨细胞前体细胞中，仅用CSF-1处理，MITF/PU.1复合体与抑制物Eos和共抑制物复合体相互作用，抑制破骨细胞靶基因的表达。在同时存在CSF-1和RANKL的情况下，Eos/共抑制物复合体被含有p38MAPK、共激活子CBP/p300和BRG-1染色质重塑复合体的复合体所取代。在这些事件之后，转录因子NFATc1被招募到目标启动子。这些数据导致了我们的总体假设：MITF/PU.1复合体作为骨微环境中遇到的信号的整合因子，在承诺的髓系祖细胞中发挥作用，影响破骨细胞功能所必需的基因表达的变化。公共卫生相关性：这项工作将确定在来自骨骼微环境的信号触发分化为特定细胞类型之前，承诺的髓系祖细胞的基因表达模式被调节的机制。这不仅是破骨细胞分化的关键问题，也是一个普遍存在的生物学问题。此外，这些研究可能会直接应用于重大的人类疾病。特别是，绝经后妇女的骨质疏松症以及多发性骨髓瘤、乳腺癌和前列腺癌患者发生的溶骨性骨破坏都是这项研究可能产生潜在影响的临床情况的例子。识别癌细胞中的分子靶点，以及开发选择性干扰这些靶点作用的药理学试剂，一直是癌症研究的长期目标，最终开始见效。在骨骼疾病中，类似的策略应该允许合理设计干扰特定分子靶点的药物。MITF的遗传学和分子生物学研究表明，MITF通路可能为某些骨骼疾病提供分子靶点。

DESCRIPTION (provided by applicant): The microphthalmia transcription factor (MITF) is required for terminal differentiation of developmentally unrelated cell types including osteoclasts, mast cells, pigmented retinal epithelial cells and melanocytes and regulates distinct target genes in each of these cell types. During terminal differentiation of osteoclasts, MITF regulates a set of genes that are required for bone resorption, including Cathepsin K (Ctsk), the chloride channel Ccln7, Ostm1/grey lethal, and tartrate resistant acid phosphatase/acid phosphatase 5 (Trap/Acp5). Mutations in this set of genes are associated with osteoclast dysfunction and bone disorders in humans and mice, strongly arguing that MITF plays a central role in gene regulation during terminal osteoclast differentiation. Our work has demonstrated that interactions between MITF and the ETS-family factor PU.1 are necessary to selectively regulate this set of target genes in osteoclasts. MITF is also a direct target of CSF-1/RANKL signaling during osteoclast differentiation, activated directly by Mitogen Activated Protein Kinases (MAPK), Erk and p38. In the current grant period, our work has revealed interacting proteins that suggest potential mechanisms underlying ITF/PU.1 action in osteoclasts. Unexpectedly, in committed osteoclast progenitors treated with CSF-1 alone, the MITF/PU.1 complex interacts with the repressor Eos and co-repressor complexes to suppress the expression of osteoclast target genes. In the presence of both CSF-1 and RANKL, the Eos/co-repressor complexes are replaced by complexes that contain p38 MAPK, the co-activator CBP/p300, and the BRG-1 chromatin remodeling complex. Subsequent to these events, the transcription factor NFATc1 is recruited to the target promoters. This data leads to our overall hypothesis: MITF/PU.1 complexes act in committed myeloid progenitors as integrators of signals encountered in the bone microenvironment to effect changes in the expression of genes essential for osteoclast function. PUBLIC HEALTH RELEVANCE: This work will define mechanisms by which gene expression patterns are regulated in committed myeloid progenitors before signals from the bone microenvironment trigger differentiation into specific cell types. This is a key problem not only in osteoclast differentiation, but is a problem of general biological interest. In addition, these studies may have direct applications to significant human diseases. In particular, osteoporosis in post-menopausal women and the osteolytic bone destruction that occurs in patients with multiple myeloma, breast cancer and prostate cancer are examples of clinical conditions where this research may have potential impact. The identification of molecular targets in cancer cells and the development of pharmacological agents that selectively interfere with the action of these targets have been long term goals in cancer research that have finally started to pay dividends. A similar strategy in bone diseases should allow for the rational design of agents that interfere with specific molecular targets. The genetics and molecular biology of MITF suggests that the MITF pathway may provide molecular targets for certain bone disorders.