MI: MODULATING OSTEOCLAST GENE EXPRESSION AND FUNCTION

描述（由申请人提供）：小眼转录因子（MITF）是发育无关细胞类型（包括破骨细胞、肥大细胞、色素性视网膜上皮细胞和黑素细胞）的终末分化所必需的，并且调节每种细胞类型中的不同靶基因。在破骨细胞的终末分化过程中，MITF 调节骨吸收所需的一组基因，包括组织蛋白酶 K (Ctsk)、氯离子通道 Ccln7、Ostm1/grey lethal 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，强烈认为 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组靶基因是必要的。 MITF 也是破骨细胞分化过程中 CSF-1/RANKL 信号传导的直接靶点，由丝裂原激活蛋白激酶 (MAPK)、Erk 和 p38 直接激活。在当前的资助期内，我们的工作揭示了相互作用的蛋白质，这些蛋白质暗示了 ITF/PU.1 在破骨细胞中作用的潜在机制。出乎意料的是，在单独用CSF-1处理的定型破骨细胞祖细胞中，MITF/PU.1复合物与阻遏物Eos和共阻遏物复合物相互作用，抑制破骨细胞靶基因的表达。在 CSF-1 和 RANKL 均存在的情况下，Eos/辅阻遏物复合物被包含 p38 MAPK、辅激活剂 CBP/p300 和 BRG-1 染色质重塑复合物的复合物取代。在这些事件之后，转录因子 NFATc1 被招募到目标启动子。这些数据引出了我们的总体假设：MITF/PU.1 复合物在定型骨髓祖细胞中作为骨微环境中遇到的信号整合器起作用，以影响破骨细胞功能所必需的基因表达的变化。公共健康相关性：这项工作将定义在来自骨微环境的信号触发分化为特定细胞类型之前，在定型骨髓祖细胞中调节基因表达模式的机制。这不仅是破骨细胞分化的关键问题，而且是具有普遍生物学意义的问题。此外，这些研究可能直接应用于重大人类疾病。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质破坏是本研究可能产生潜在影响的临床病症的例子。识别癌细胞中的分子靶标并开发选择性干扰这些靶标作用的药物一直是癌症研究的长期目标，并终于开始带来红利。骨疾病中的类似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表明 MITF 途径可能为某些骨疾病提供分子靶点。

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.