Function and Regulation of Osteonectin in Bone

描述(申请人提供)：随着年龄的增长，骨丢失是由于骨重建不平衡造成的，成骨细胞数量减少，破骨细胞数量增加，骨髓中脂肪细胞数量增加。间充质干细胞(MSCs)可同时分化为成骨细胞和脂肪细胞，随着年龄的增长，MSC的谱系分配发生改变。MSC的谱系分配受不同的细胞内信号、细胞间相互作用和骨微环境的控制。骨微环境中最丰富的非胶原基质蛋白是基质糖蛋白Osteonectin(富含半胱氨酸的酸性分泌蛋白，SPARC；BM-40)。在骨骼中，骨连接蛋白促进成骨细胞的承诺，抑制脂肪生成，并调节甲状旁腺素治疗后骨形成和骨吸收之间的平衡。它在成骨细胞分化早期高表达，但随着细胞获得成熟成骨细胞的特征，其表达减弱。相反，在成骨细胞分化过程中，骨连接蛋白转录水平变化不大，表明在翻译水平上存在调控。MicroRNAs(MiRNAs)是一种小的非编码RNA，通过与靶mRNAs的3‘非翻译区(UTR)相互作用而介导翻译抑制。我们发现miR-29a和-29c作用于骨连接蛋白3‘非编码区，并在成骨细胞中介导翻译抑制。我们推测miR-29a和-29c调控成骨细胞分化。重要的是，骨结素基因3‘端非编码区的单核苷酸多态(SNPs)与人类骨密度相关，这些单核苷酸多态(SNPs)调节3’端非编码区的功能。由于骨连蛋白对正常的骨重建和骨合成代谢甲状旁腺素治疗的反应至关重要，我们工作的目标是了解其在骨骼中表达的转录后调控机制。我们将1.确定人骨连接蛋白3‘非编码区SNPs如何在体外调节成骨细胞分化过程中的蛋白水平；2.利用携带人UTR基因敲入突变的小鼠，在体内测定人骨连接蛋白3’非编码区单倍型的活性；3.确定miR-29在体外成骨细胞分化中的作用。这些研究将填补在调节骨量的关键机制方面的知识空白。此外，我们获得的信息可以应用于其他被认为在病理中起作用的疾病，如肥胖症和癌症。这项建议包含基本和翻译部分，我们将获得与基础科学和临床研究相关的信息。

DESCRIPTION (provided by applicant): Bone loss with aging results from imbalanced bone remodeling, with decreased osteoblast number, increased osteoclast number, and increased adipocyte number in the marrow. Mesenchymal stem cells (MSCs) give rise to both osteoblasts and adipocytes, and MSC lineage allocation is altered in aging. MSC lineage allocation is controlled by diverse intracellular signals, cell-cell interactions and the bone microenvironment. The most abundant non-collagen matrix protein in the bone microenvironment is the matricellular glycoprotein osteonectin (secreted protein acidic rich in cysteine, SPARC; BM-40). In the skeleton, osteonectin promotes osteoblast committment, suppresses adipogenesis, and regulates the balance between bone formation and resorption in response to PTH treatment. It is highly expressed early in osteoblastic differentiation, but its expression decreases as the cells acquire characteristics of mature osteoblasts. In contrast, osteonectin transcript levels change little during osteoblastic differentiation, indicating regulation at the level of translation. MicroRNAs (miRNAs) are small non-coding RNAs that mediate translational repression by interacting with the 3' untranslated region (UTR) of target mRNAs. We found that miR-29a and -29c act on the osteonectin 3' UTR and mediate translational repression in committed osteoblasts. We hypothesize that miR-29a and -29c regulate osteoblastic differentiation. Importantly, single nucleotide polymorphisms (SNPs) in the 3' UTR of osteonectin gene are associated with bone density in humans, and these SNPs modulate 3' UTR function. Since osteonectin is critical for normal bone remodeling and response to bone anabolic PTH therapy, the goal of our work is to understand post-transcriptional mechanisms regulating its expression in the skeleton. We will 1. determine how human osteonectin 3' UTR SNPs modulate protein levels during osteoblastic differentiation in vitro; 2. determine the activity of human osteonectin 3' UTR haplotypes in vivo, using mice carrying knock-in mutations of the human UTR and 3. determine the role of miR-29 in osteoblast differentiation in vitro. These studies will fill a substantial void in the knowledge of key mechanisms regulating bone mass. In addition, the information we acquire could be applied to other diseases in which osteonectin is thought to play a role in pathology, such as obesity and cancer. This proposal contains basic and translational components, and we will obtain information relevant to both basic science and clinical studies.