Transcriptional Regulation of Osteoclasts by Mitf

Mitf 对破骨细胞的转录调控

• 批准号: 6323978
• 负责人: Yi-Ling Lin
• 金额: $ 13.04万
• 依托单位: DANA-FARBER CANCER INST
• 依托单位国家: 美国
• 财政年份: 2001
• 资助国家: 美国
• 起止时间: 2001-09-01 至 2006-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6323978
• 关键词: Adenoviridae; Retroviridae; biological signal transduction; cell growth regulation; dimer; embryo /fetus tissue /cell culture; enzyme activity; gene expression; genetic regulation; laboratory mouse; laboratory rat; microphthalmos; mitogen activated protein kinase; monoclonal antibody; mutant; osteoclasts; osteopetrosis; phosphorylation; protein kinase A; protein structure function; transcription factor; transfection /expression vector

Recent advances in bone biology have revolutionized dentistry. With better understanding of how osteoblasts respond to bone-forming signals, dentists now can use a variety of grafting materials to perform bone augmentation procedures and grow bone "de novo". In contrast, it is still largely unknown how osteoclasts regulate their activity in response to local factors. This knowledge is of great importance to the dental profession since so many dental diseases are directly or indirectly the result of unwanted osteoclast activation. One critical regulator of osteoclast activity is Microphthalmia transcription factor (Mitt). Mice carrying semidominant Mitf alleles exhibit various degrees of osteopetrosis, indicating Mitf is a promising tool for studying gene regulation in osteoclasts. The objective of the proposed study is to gain knowledge of the transcriptional regulation of osteoclasts by Mitf and three other family members, TFEB, TFEC and TFE3 (the four proteins form the "MiT" family). Since Mitf semidominant mutant mice exhibit an age-dependent phenotype, the chronological expression profiles of MiT proteins during osteoclast development will be examined since this would be consistent with functional rescue of Mitf by family members. MAPK and Rsk/PKA have previously been demonstrated to phosphorylate Mitf in melanocytes, which resulting in Mitf activation. To examine if similar processes occur in osteoclasts, recombinant MiT mutant viruses carrying disrupted MAPK and Rsk/PKA phosphorylation sites will be used, and the resulting phenotypic changes will be analyzed. Mitf has been found to regulate cytoskeletal changes in response to maturation signals during fetal osteoclast fusion. Overexpression of the individual MiT protein in fetal mi/mi osteoclasts (which do not fuse or form multinucleated giant cells) will be tested for the ability to rescue the fusion defect. Since Mitf may affect multinucleate giant cell formation by regulating the expression of molecules essential for osteoclast fusion, candidates known to be essential for this process (such as vitronectin receptor and osteopontin) will be examined for their relationship with Mitf.

骨生物学的最新进展使牙科发生了革命性的变化。随着对成骨细胞对骨形成信号的反应有了更好的了解，牙医现在可以使用各种移植材料来进行骨增强手术，并“从头”长出骨。相比之下，破骨细胞是如何根据局部因素调节其活性的，这在很大程度上仍然是未知的。这方面的知识对牙科专业来说是非常重要的，因为许多牙科疾病都是由不想要的破骨细胞激活直接或间接造成的。一个关键的破骨细胞活性调节因子是小眼转录因子（Mitt）。携带半显性Mitf等位基因的小鼠表现出不同程度的骨质疏松，这表明Mitf是研究破骨细胞基因调控的一个很有前途的工具。本研究的目的是了解Mitf和其他三个家族成员TFEB、TFEC和TFE3（“MiT”家族的四种蛋白）对破骨细胞的转录调控。由于Mitf半显性突变小鼠表现出年龄依赖性表型，因此将检查破骨细胞发育过程中MiT蛋白的时间顺序表达谱，因为这将与家庭成员对Mitf的功能拯救相一致。MAPK和Rsk/PKA先前已被证明可以磷酸化黑色素细胞中的Mitf，从而导致Mitf激活。为了检查是否在破骨细胞中发生类似的过程，将使用携带被破坏的MAPK和Rsk/PKA磷酸化位点的重组MiT突变病毒，并分析由此产生的表型变化。已发现Mitf在胎儿破骨细胞融合过程中调节细胞骨架变化以响应成熟信号。在胎儿mi/mi破骨细胞（不能融合或形成多核巨细胞）中过度表达单个MiT蛋白将被测试以挽救融合缺陷的能力。由于Mitf可能通过调节破骨细胞融合所必需的分子的表达来影响多核巨细胞的形成，因此我们将研究对这一过程至关重要的候选分子（如玻璃体连接蛋白受体和骨桥蛋白）与Mitf的关系。