Discovery of osteoblast and osteoclast bone mass effector genes using advanced genomics

利用先进基因组学发现成骨细胞和破骨细胞骨量效应基因

• 批准号: 10675631
• 负责人: Struan F A Grant
• 金额: $ 66.98万
• 依托单位: UNIVERSITY OF MICHIGAN AT ANN ARBOR
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-08-03 至 2027-04-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10675631
• 关键词: ATAC-seq; Age Months; American; Bioinformatics; Biology; Bone Density; Bone Diseases; Bone Resorption; CRISPR/Cas technology; Cell Differentiation process; Cell Lineage; Cells; Child; Chromatin; Chromosome Mapping; Clustered Regularly Interspaced Short Palindromic Repeats; Collaborations; Collection; Communities; Complex; Data; Development; Disease; Enhancers; Etiology; Female; Fracture; Gene Expression; Genes; Genetic; Genetic Diseases; Genome; Genomics; Goals; Grant; Human; In Vitro; International; Knockout Mice; Laboratories; Link; Mechanics; Methods; Minerals; Mission; Modeling; Mus; Osteoblasts; Osteoclasts; Osteogenesis; Osteoporosis; Pathogenesis; Pathway Analysis; Phenotype; Physiology; Play; Process; Public Domains; Publishing; Regulation; Reporting; Research; Resolution; Role; Sample Size; Sampling; Signal Transduction; Small Interfering RNA; Testing; Time; Validation; Variant; Work; bone; bone geometry; bone loss; bone mass; causal variant; chromosome conformation capture; clinically relevant; follow-up; fracture risk; gene discovery; genetic variant; genome wide association study; genome-wide; genomic data; in vivo; interest; male; microCT; mouse model; novel; novel therapeutics; osteoblast differentiation; prevent; promoter; trait; transcriptome sequencing

Osteoporosis is a devastating disease of bone that impacts over 10 million Americans. While the cellular basis for osteoporosis includes an imbalance in bone formation by osteoblasts and bone resorption by osteoclasts, there are relatively few validated, clinically relevant genes directly linked to osteoporosis. There is a significant need to discover new genes that influence osteoporosis pathogenesis. Discovery of new osteoporosis genes will eventually permit the field of bone and mineral biology to achieve the long-term goal of developing new therapies to both prevent and treat this debilitating disease. The existing, long-term collaboration between the Hankenson and Grant laboratories has been focused on understanding the functional significance of genome wide association study (GWAS) signals associated with bone mass, osteoporosis and fracture risk. We have developed methods to use those signals to identify novel genes putatively involved in disease pathogenesis. While GWAS efforts by numerous research groups have been successful in discovering genomic variants robustly associated with bone mineral density (BMD) and fracture, GWAS only reports signals associated with a given trait and not necessarily culprit genes. In this proposal we utilize a computationally advanced, multi-step process that integrates genome level data to identify novel osteoblast and osteoclast genes. This bioinformatically-driven `genome-wide variant to gene mapping' effort combines RNA-seq, ATAC-seq and high-resolution chromatin conformation capture methods to implicate culprit effector genes. We have already used this approach in osteoblast lineage cells and 30% of osteoporosis-associated GWAS signals were shown to have direct physical contact with genes in these cells, totaling 86 putative target genes. Several of these targets (ex. EPDR1, ING3) have already had extensive functional follow-up. However, many more need functional follow-up and there are still 70% of osteoporosis associated GWAS loci that remain unresolved. Importantly, our initial work focused only on discovering osteoblast-associated genes, and thus genes that play a role in osteoclasts were not revealed. Furthermore, our published work to date has only focused on one time-point during the osteoblast differentiation process, thus genes that play roles at later points in cell differentiation have not been discovered. This comprehensive application will functionalize GWAS findings, and in doing so, reveal novel genes involved in regulating bone formation and resorption. Our established pipeline from gene discovery to gene validation has been robustly tested and thus far, although our sampling has been small, we have had a 100% hit rate for validating putative effector genes. Thus, it is our hypothesis that we can uncover many more BMD effector genes by conducting high resolution `genome-wide variant to gene mapping' in osteoclasts and osteoblasts. The relevance of genes will be validated using both in vitro and in vivo approaches in mouse models. Upon completion, we will provide the bone community with new targets to pursue for understanding mechanism.

骨质疏松症是一种毁灭性的骨骼疾病，影响着超过 1000 万美国人。虽然细胞基础 骨质疏松症包括成骨细胞的骨形成和破骨细胞的骨吸收失衡， 与骨质疏松症直接相关的经过验证的临床相关基因相对较少。有一个显着的 需要发现影响骨质疏松症发病机制的新基因。发现新的骨质疏松症基因 最终将使骨和矿物生物学领域实现开发新产品的长期目标 预防和治疗这种使人衰弱的疾病的疗法。 汉肯森和格兰特实验室之间现有的长期合作重点是 了解与相关的全基因组关联研究 (GWAS) 信号的功能意义 骨量、骨质疏松症和骨折风险。我们已经开发出使用这些信号来识别新颖的方法 推测与疾病发病机制有关的基因。尽管众多研究小组的 GWAS 努力已经 成功发现与骨矿物质密度（BMD）密切相关的基因组变异 对于骨折，GWAS 仅报告与给定性状相关的信号，而不一定是罪魁祸首基因。 在此提案中，我们利用先进的计算多步骤过程，将基因组水平数据集成到 鉴定新的成骨细胞和破骨细胞基因。这种生物信息驱动的“全基因组变异” Mapping 的工作结合了 RNA-seq、ATAC-seq 和高分辨率染色质构象捕获方法 涉及罪魁祸首效应基因。我们已经在成骨细胞谱系细胞中使用了这种方法，并且 30% 骨质疏松症相关的 GWAS 信号被证明与这些细胞中的基因有直接的物理接触， 总共86个假定的靶基因。其中一些目标（例如 EPDR1、ING3）已经得到了广泛的应用 功能性跟进。然而，更多的人需要进行功能随访，仍有 70% 的人患有骨质疏松症 相关的 GWAS 位点仍未解决。重要的是，我们最初的工作只专注于发现 成骨细胞相关基因，以及在破骨细胞中发挥作用的基因尚未被揭示。此外， 迄今为止，我们发表的工作仅关注成骨细胞分化过程中的一个时间点， 因此，尚未发现在细胞分化后期发挥作用的基因。 这一综合应用将发挥 GWAS 研究结果的作用，并在此过程中揭示涉及的新基因 调节骨形成和吸收。我们已建立从基因发现到基因验证的管道 经过严格测试，到目前为止，虽然我们的样本很小，但我们的命中率是 100% 验证假定的效应基因。因此，我们假设我们可以发现更多的 BMD 效应子 通过在破骨细胞和成骨细胞中进行高分辨率“全基因组变异到基因图谱”来分析基因。 将使用小鼠模型中的体外和体内方法来验证基因的相关性。之上 完成后，我们将为骨骼界提供新的目标来追求理解机制。