Fibulin-1 Regulation of the DiGeorge Syndrome Pathogenesis Pathway

描述(申请人提供)：小鼠靶向基因失活研究的新发现表明，细胞外基质蛋白fiBulin-1(Fbln1)是心脏流出道(OFT)、咽腺、颅神经和颅骨正常发育所必需的。Fbln1缺陷小鼠表现出的发育异常概括了与22q11缺失/DiGeorge综合征(DGS)相关的许多异常，该综合征发生的频率为1：4000人出生。Fbln1缺陷小鼠和DGS人类在表型上的相似性表明，尽管Fbln1位于22q11缺失区域之外，但它是DGS中功能失调的途径的修饰者。DGS的分子途径尚未完全阐明。关键成分是转录因子Tbx1、Fgf8和Fgf8信号通路的中间产物。有几条证据支持Fbln1调节这一途径的假设。DNA微阵列分析表明，Fbln1是叉头盒转录因子表达所必需的，该转录因子调节包括次级心区(SHF)在内的咽弓区域TBX1和Fgf8的表达。定量聚合酶链式反应分析表明，在Fbln1缺失胚胎的咽弓组织中，TBX1的表达显著降低。我们还发现Fbln1与Fgf8相互作用，并抑制Fgf8介导的MAPK信号通路的激活。这些观察结果以及Fbln1沿着NCC迁移途径表达的发现，以及它可以作为细胞引导和运动的调节器，以及它相互作用和调节蛋白多糖的蛋白分解，NCC迁移的调节，导致了以下假设：Fbln1是Fgf8影响依赖NCC的咽弓发育和衍生结构形成的关键组成部分，该机制包括OFT，咽腺和颅面结构。为了验证这一假说并获得机制上的理解，有三个具体的目的：1)确定Fbln1在Fgf8信号通路中的作用；2)确定Fbln1与Tbx1之间以及Fbln1与Verscan之间的遗传相互作用；以及3)确定Fbln1在控制NCC增殖和引导中的作用。这项实验的发现有望为Fbln1引导心脏OFT、大血管、头盖骨和咽腺的正常发育的机制提供新的见解，并有助于理解涉及这些组织的先天性缺陷的病因。公共卫生相关性：这项研究试图从机制上了解与DiGeorge综合征(DGS)相关的发育异常(包括心脏缺陷)，DGS发生的频率为4000人出生中就有一例。这项拟议的研究旨在确定细胞外基质蛋白-1在调节细胞(神经脊细胞)行为中的作用，这些细胞行为有助于DGS受影响组织的形成。这项研究的一个新方向源于我们的发现，即纤维蛋白-1调节Fgf8的活性，Fgf8是一种生长因子，被认为是DGS中功能失调的途径的关键组成部分。

DESCRIPTION (provided by applicant): New findings from targeted gene inactivation studies in mice reveal that the extracellular matrix protein fibulin-1 (Fbln1) is required for normal development of the cardiac outflow tract (OFT), pharyngeal glands, cranial nerves and bones of the skull. The developmental anomalies displayed by mice deficient in Fbln1 recapitulate many of the anomalies associated with 22q11 deletion/DiGeorge syndrome (DGS), which occurs with a frequency of 1:4000 human births. The similarity in phenotypes between Fbln1-deficient mice and DGS humans suggests that Fbln1, although located outside of the 22q11 deletion region, is a modifier of the pathway that is dysfunctional in DGS. The molecular pathway of DGS has not been fully elucidated. Key components are the transcription factor Tbx1, Fgf8 and intermediates of the Fgf8 signaling pathway. Several lines of evidence support the hypothesis that Fbln1 regulates this pathway. DNA microarray analysis indicates that Fbln1 is required for expression of forkhead box transcription factors that regulate expression of both Tbx1 and Fgf8 in pharyngeal arch regions including the secondary heart field (SHF). qPCR analysis shows that Tbx1 expression is significantly decreased in pharyngeal arch-containing tissues from Fbln1 null embryos. We also find that Fbln1 interacts with Fgf8 and inhibits Fgf8-mediated activation of the Map kinase-signaling pathway. These observations together with findings that Fbln1 is expressed along pathways of NCC migration, that it can act as a regulator of cell guidance and motility and that it interacts and modulates proteolysis of the proteoglycan versican, a regulator of NCC migration, lead to the following hypothesis: Fbln1 is a key component of the mechanism by which Fgf8 influences NCC-dependent development of the pharyngeal arch and the formation of derivative structures including the OFT, pharyngeal glands and craniofacial structures. To test this hypothesis and gain mechanistic understanding there are three specific aims: 1) Define the effects of Fbln1 on the Fgf8 signaling pathway; 2) Identify genetic interactions between Fbln1 and Tbx1 and between Fbln1 and versican; and 3) Define the role of Fbln1 in controlling NCC proliferation and guidance. Findings from this experimentation are expected to lead to new insights into the mechanism by which Fbln1 guides the normal development of the cardiac OFT, great vessels, cranium and pharyngeal glands and to contribute to an understanding of the etiology of congenital defects involving these tissues. Public Health Relevance: This research seeks to gain a mechanistic understanding of the developmental anomalies (including heart defects) associated with DiGeorge syndrome (DGS), which occurs with a frequency of one in 4000 human births. The proposed studies are designed to determine the role of the extracellular matrix protein, fibulin-1, in regulating the behaviors of cells (neural crest cells) that contribute to the formation of tissues affected in DGS. A novel direction for the research stems from our discovery that fibulin-1 regulates the activity of Fgf8, a growth factor implicated as a key component of the pathway that is dysfunctional in DGS.