Transcriptional Regulation of Osteoclasts by Mitf

Mitf 对破骨细胞的转录调控

• 批准号: 6775714
• 负责人: Yi-Ling Lin
• 金额: $ 13.5万
• 依托单位: UNIVERSITY OF KENTUCKY
• 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/6775714
• 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彻底改变了牙科。随着对成骨细胞如何响应骨形成信号的更好理解，牙医现在可以使用各种移植材料来执行骨增量程序并“重新”生长骨。相比之下，破骨细胞如何调节其活动以响应局部因素在很大程度上仍然未知。这方面的知识是非常重要的牙科专业，因为这么多的牙科疾病是直接或间接的结果不必要的破骨细胞激活。破骨细胞活性的一个关键调节因子是小眼症转录因子（Mitt）。携带半显性Mitf等位基因的小鼠表现出不同程度的骨硬化，表明Mitf是一个有前途的工具，用于研究破骨细胞的基因调控。本研究的目的是了解Mitf和其他三个家族成员TFEB、TFEC和TFE 3（这四种蛋白形成“MiT”家族）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期间MiT蛋白的时序表达谱，因为这与家族成员对Mitf的功能性拯救一致。MAPK和Rsk/PKA已被证实可使黑素细胞中的Mitf磷酸化，从而导致Mitf活化。为了检查在破骨细胞中是否发生类似的过程，将使用携带破坏的MAPK和Rsk/PKA磷酸化位点的重组MiT突变体病毒，并将分析所得的表型变化。已发现Mitf在胎儿破骨细胞融合期间响应成熟信号调节细胞骨架变化。将检测胎儿mi/mi破骨细胞（不融合或形成多核巨细胞）中单个MiT蛋白的过表达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形成，因此将检查已知对该过程至关重要的候选物（如玻连蛋白受体和骨桥蛋白）与Mitf的关系。