Sp7 Mediated Control of Runx2 Function for Osteoblast Differentiation

Sp7介导的Runx2功能对成骨细胞分化的控制

基本信息

批准号：
10359695
负责人：
Amjad Javed
金额：
$ 35.06万
依托单位：
UNIVERSITY OF ALABAMA AT BIRMINGHAM
依托单位国家：
美国
项目类别：
财政年份：
2012
资助国家：
美国
起止时间：
2012-03-01 至 2025-12-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10359695
关键词：
Address Affinity Anatomy Animals BMP2 gene Biochemical Biological Biological Availability Biological Process Biology Bone Diseases Bone Matrix Bone Morphogenetic Proteins Calvaria Cartilage Cartilage Diseases Cell Differentiation process Cell Maturation Cell model Cells Chondrocytes Chromatin Cleidocranial Dysplasia Competence Complex Cues DNA Binding DNA Binding Domain Data Development Differentiation and Growth Embryo Erinaceidae Event Failure Family Functional disorder Gene Deletion Gene Expression Gene Expression Profile Genes Genetic Genetic Transcription Goals Growth Human In Situ Hybridization Inherited Innovative Therapy Intervention Knockout Mice Knowledge Link MAP Kinase Gene Maintenance Mammals Mediating Mesenchymal Messenger RNA Metabolic Bone Diseases Modeling Modification Molecular Molecular Biology Monitor Mus Mutagenesis Mutation Null Lymphocytes Osteoblasts Osteogenesis Osteogenesis Imperfecta Pathway interactions Pattern Phenotype Physiologic Ossification Post-Translational Protein Processing Process Protein Biochemistry Proteins RBX1 gene RNA Regulation Research Role Runx2 protein Signal Transduction Skeleton Specific qualifier value Specificity Subgroup Sumoylation Pathway Testing Therapeutic Intervention Tissues To specify Tooth eruption Vertebrates Zinc Fingers base bone bone cell bone repair embryo tissue experimental study improved in vivo insight interdisciplinary approach intramembranous bone formation mineralization morphogens mutant novel osteoblast differentiation osteogenic osteoprogenitor cell prevent protein function reconstitution recruit repaired skeletal skeletal disorder skeletal tissue skeletogenesis stable cell line therapy development transcription factor transcription regulatory network ubiquitin ligase ubiquitin mediated proteasome degradation

项目摘要

ABSTRACT: Development of skeleton in mammals is an exceedingly complex process. Completion of both endochondral and intramembranous ossification entails a highly intricate but well-coordinated process of patterning, cell fate commitment, differentiation, growth, and remodeling. These events are specified by a coordinated temporal and spatial pattern of gene expression. At first, secreted morphogens such as hedgehog, bone morphogenetic proteins, wingless proteins, and others, signal to key transcription factors to specify gene expression. Runx2 is an essential transcription factor for both chondrocyte and osteoblast differentiation. Runx2 gene deletion results in embryonic lethality due to a complete failure of bone formation. In humans, mutation of the Runx2 gene causes cleidocranial dysplasia, a dominantly inherited skeletal disorder. Another master regulator of skeletogenesis is the Specificity protein-7 (Sp7). Sp7 belongs to the Sp subgroup of the Krüppel-like family of transcription factors characterized by three zinc-finger DNA- binding domains. Deletion of Sp7 gene results in failure of osteoblasts, and bone formation. In humans, mutation of the Sp7 gene is linked with the recessive form of osteogenesis imperfecta, skeletal fragility and delayed tooth eruption. However, very little is known about the underlying molecular mechanism for the surprisingly similar phenotype from the two seemingly unrelated proteins. Runx2 is required for the expression of Sp7, as mice with targeted disruption of the Runx2 gene completely lack expression of Sp7. In sharp contrast, the Runx2 expression is normal in the skeletal cells of Sp7 null animals. The functional incompetency of Runx2 in the Sp7 null mice suggests that Sp7 presence is obligatory for completion of the Runx2 osteogenic activity. It is important to note that the observation of Runx2 expression in Sp7 null mice is limited to only RNA, determined by in situ hybridization of embryonic tissues. Our data show that despite normal levels of Runx2 mRNA, Runx2 protein is highly unstable in skeletal tissues of Sp7 null mice. We further demonstrate that Runx2 and Sp7 proteins form a molecular complex and their transcriptional activity is regulated by unique posttranslational modifications. Our findings strongly suggest that in skeletal cells, Sp7 acts as a molecular rheostat and is necessary for functional stability and turnover of Runx2 protein. Our experiment will assess endogenous levels of Runx2 protein in Sp7 null background and by a regulated and selective gene reconstitution in osteoprogenitor cells. The goal of this application is to identify and define a) spatial and temporal organization and assembly of Runx2 and Sp7 regulatory complexes for the formation and/or maintenance of osteoblasts and b) mechanisms supporting the stable complex formation and retention of competency for skeletal gene expression. Knowledge obtained from this study will provide molecular insights into components of a bone regulatory complex that can be targeted for innovative therapy to improve cartilage and bone formation and repair.

摘要：哺乳动物骨骼的发育是一个极其复杂的过程。软骨内和膜内骨化需要一个高度复杂但协调良好的过程这些事件由细胞命运决定、分化、生长和重塑来指定。协调基因表达的时间和空间模式首先，分泌形态发生素，例如刺猬蛋白、骨形态发生蛋白、无翅蛋白等向关键转录因子发出信号 Runx2 是软骨细胞和成骨细胞必需的转录因子。 Runx2 基因缺失导致胚胎因骨骼完全衰竭而死亡。在人类中，Runx2 基因的突变会导致锁骨颅骨发育不良，这是一种显性遗传的疾病。骨骼形成的另一个主要调节因子是特异性蛋白 7 (Sp7)。属于以三个锌指 DNA 为特征的 Krüppel 样转录因子家族的 Sp 亚组 Sp7 基因的缺失会导致人类成骨细胞和骨形成的失败。 Sp7 基因突变与隐性成骨不全、骨骼脆弱和然而，人们对牙齿萌出延迟的潜在分子机制知之甚少。令人惊讶的是，这两个看似无关的蛋白质的表型是必需的。 Sp7 的表达，因为靶向破坏 Runx2 基因的小鼠完全缺乏 Sp7 的表达。与此形成鲜明对比的是，Runx2 表达在 Sp7 缺失动物的骨骼细胞中是正常的。 Sp7 缺失小鼠中 Runx2 的无能表明 Sp7 的存在对于完成 Runx2 成骨活性值得注意的是，Sp7 缺失小鼠中 Runx2 表达的观察。仅限于RNA，通过胚胎组织的原位杂交确定，尽管如此。尽管 Runx2 mRNA 水平正常，但 Sp7 缺失小鼠的骨骼组织中 Runx2 蛋白高度不稳定。进一步证明Runx2和Sp7蛋白形成分子复合物及其转录活性我们的研究结果强烈表明，在骨骼细胞中， Sp7 充当分子变阻器，对于 Runx2 蛋白的功能稳定性和周转是必需的。我们的实验将评估 Sp7 无效背景中 Runx2 蛋白的内源水平，并通过调节和骨祖细胞中的选择性基因重建该应用的目标是识别和重建。定义 a) Runx2 和 Sp7 调控复合体的空间和时间组织和组装成骨细胞的形成和/或维持以及 b) 支持稳定复合物形成的机制从这项研究中获得的知识将提供骨骼基因表达能力的保留。对骨调节复合体成分的分子洞察，可用于创新改善软骨和骨形成和修复的疗法。