Investigations of the molecular genetics and pathogenesis of scoliosis.

摘要 这是一项既定计划的续展申请，以调查开发和 脊柱的动态平衡。在第一个资助期，我们的研究表明Adgrg6基因， 与一种称为青春期特发性脊柱侧凸的常见人类脊柱疾病有关，在 维持小鼠脊柱的排列。我们发现G蛋白偶联受体Adgrg6调控基因 脊柱间盘和致密结缔组织的表达和生物力学特性。 此外，我们还证明了Adgrg6刺激cAMP信号调节因子 脊柱纤维软骨组织的动态平衡。我们的发现提出了一个新的假设，即刺激 CAMP信号转导可降低Adgrg6缺失所致脊柱侧弯的发病和严重程度 发信号。此外，人类对脊柱侧弯的遗传学分析发现了一种新的变异，位于 转录因子SOX9的转录激活结构域。值得注意的是，有针对性地破坏这种 Sox9结构域引起的小鼠脊柱侧弯及纤维软骨组织中基因表达异常 脊椎。在这里，我们将检验Adgrg6和Sox9在功能上与调控相关的假设 脊柱的动态平衡和对齐。 为了增加我们的计划目标的广度，我们继续进行正向遗传筛选，以分离出一系列 脊椎紊乱突变斑马鱼。我们最近发现了两个斑马鱼突变体，它们未能补充一种新的 胸椎侧弯的表型，提示了一种控制脊柱形态发生的新途径。人物刻画 这种独特的胸椎侧弯表型的研究将把我们的知识扩展到细胞和分子 脊柱疾病的异质性。在这里，我们将检验斑马鱼胸椎侧弯是 由嘌呤能信号中断导致脊索生物发生缺陷引起的。 我们将通过分成三个具体目标的研究来检验这些假设。具体目标1将深化 我们对脊髓中Adgrg6信号效应器的机制理解，并测试刺激是否 CAMP可恢复脊柱纤维软骨组织的动态平衡，延缓脊椎病的发生和发展 脊柱侧弯。特殊目标2将描述一个新的Sox9突变体中脊柱侧弯的细胞和分子原因 并使用此模型来测试Adgrg6和Sox9变体之间的遗传交互作用是否会增加 易患脊柱侧弯。特定目标3将表征新的胸椎侧弯突变斑马鱼并测试 在斑马鱼的脊索生物发生和脊椎形态发生中，嘌呤能信号是必不可少的模型。 我们的研究结果将为人类进化所必需的分子遗传学和生物学过程提供新的见解。 脊柱的发育和动态平衡。这些研究可能提供对生物学的基本见解。 与人类骨骼发育不良和脊柱侧弯相关的过程和途径。

ABSTRACT This is a renewal application of an established program to investigate the regulation of the development and homeostasis of the spine. During the first funding period, our studies demonstrated that the gene Adgrg6, implicated in a common human spine disorder called adolescent idiopathic scoliosis, has an essential role in maintaining spine alignment in mice. We showed that the G-protein coupled rector Adgrg6 regulates gene expression and biomechanical properties of the intervertebral discs and dense connective tissue of the spine. Furthermore, we demonstrated that Adgrg6 stimulates cAMP signaling regulate factors essential for homeostasis of fibrocartilaginous tissue of the spine. Our findings suggest a new hypothesis that stimulation of cAMP signaling can decrease the onset and severity of scoliosis caused by the loss of Adgrg6 signaling. In addition, human genetics analysis of scoliosis identified a novel variant located in the transcriptional activation domain of the transcription factor SOX9. Significantly, targeted disruption of this domain of Sox9 in mice caused scoliosis and dysregulation of gene expression in fibrocartilaginous tissues of the spine. Here, we will test the hypothesis that Adgrg6 and Sox9 are functionally linked for regulation homeostasis and alignment of the spine. To add breadth to our program goals, we continued a forward genetic screen to isolate a collection of spine disorder mutant zebrafish. We recently identified two zebrafish mutants that fail to complement a novel thoracic scoliosis phenotype, suggesting a new pathway controlling spine morphogenesis. The characterization of this unique thoracic scoliosis phenotype will expand our knowledge into the cellular and molecular heterogeneity of spine disorders. Here, we will test the hypothesis that thoracic scoliosis in zebrafish is caused by a disruption of purinergic signaling leading to defects in notochord biogenesis. We will test these hypotheses via studies divided into three Specific Aims. Specific Aim 1 will deepen our mechanistic understanding of effectors of Adgrg6 signaling in the spine and test whether stimulation of cAMP can restore homeostasis to fibrocartilaginous tissues of the spine and halt the onset and progression of scoliosis. Specific Aim 2 will characterize the cellular and molecular causes of scoliosis in a novel Sox9 mutant mouse and use this model to test whether genetic interactions between Adgrg6 and Sox9 variants increase the susceptibility to scoliosis. Specific Aim 3 will characterize novel thoracic scoliosis mutant zebrafish and test a model that purinergic signaling is essential for notochord biogenesis and spine morphogenesis in zebrafish. Our results will provide new insights into the molecular genetics and biological processes necessary for the development and homeostasis of the spine. These studies may provide fundamental insights into the biological processes and pathways associated with human skeletal dysplasia and scoliosis.