The Role of Alternative Splicing of Fgf8 in Mouse Development

Fgf8 选择性剪接在小鼠发育中的作用

• 批准号: 7572933
• 负责人: JAMES Y.H LI
• 金额: $ 31.55万
• 依托单位: UNIVERSITY OF CONNECTICUT SCH OF MED/DNT
• 依托单位国家: 美国
• 财政年份: 2007
• 资助国家: 美国
• 起止时间: 2007-02-10 至 2012-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7572933
• 关键词: Adult; Affinity; Alternative Splicing; Binding; Biological; Biological Models; Brain; Brain region; Cells; Cerebellum; Chick Embryo; Development; Developmental Process; Diffusion; Embryo; Embryonic Development; FGF8 gene; Fibroblast Growth Factor 8; Fibroblast Growth Factor Receptor 1; Genes; Human; Individual; Light; Malignant Neoplasms; Mediating; Midbrain structure; Molecular; Mus; Physiological; Play; Production; Property; Protein Isoforms; Proto-Oncogene Proteins c-akt; RNA Splicing; Ras Signaling Pathway; Receptor Protein-Tyrosine Kinases; Regulation; Relative (related person); Role; Signal Transduction; Testing; Variant; base; combinatorial; developmental disease; falls; functional group; gain of function; hindbrain; insight; loss of function; mouse development; overexpression; prevent; receptor binding; response; tumor

DESCRIPTION (provided by applicant): Fibroblast growth factor 8 (Fgf8) plays crucial roles in controlling various developmental processes. Fgf8 binds and activates transmembrane receptor tyrosine kinases (FGFR1-4). Two major signaling pathways, Ras-ERK and PI3K-Akt, are known to mediate Fgf8 signaling. The single Fgf8 gene in both tiuman and mouse produces multiple isoforms of variable N-termini by alternative splicing. The biological significance of Fgf8 splice variants remains unanswered. Using the developing midbrain and hindbrain as a model system, we have demonstrated the distinct inductive activities of two Fgf8 isoforms, Fgf8a and Fgf8b. We recently discovered that all eight Fgf8 isoforms in mice fall into two functional groups, a-like (a, c, e and g) and b-like (b, d, f and h), based on their similar activities to Fgf8a and Fgf8b. The overall hypothesis to be tested is that the function of Fgf8 is orchestrated by the different activities of these two groups of Fgf8 isoforms. We postulate further that the alternative splicing of Fgf8 is an important mechanism in regulating both the levels and range of Fgf8 signaling. Focusing on Fgf8 function in development of the midbrain and cerebellum in the chick and mouse, we propose to dissect the functional roles of Fgf8a-like and Fgf8b-like isoforms, and the mechanisms underlying distinct activities of these isoforms and their mutual interactions. First, to determine the combinatorial activities of different Fgf8 isoforms in the developing mid/hindbrain, we will use a gain-of-function approach, and examine the activity of individual or combinatorial Fgf8 isoforms in chick embryos. Second, to ascertain the relative contributions of Fgf8a-like and Fgf8b-like isoforms to normal development, we will use a loss-of-function approach, and determine the requirements for Fgf8a-like and Fgf8b-like isoforms during development. Third, to determine the molecular mechanism underlying the distinct functions of Fgf8 isoforms, we will examine the range of action, diffusion and functional properties of Fgf8a and Fgf8b in brain explants. Furthermore, to determine the mechanism behind the different functions of Fgf8a and Fgf8b in the mid/hindbrain, we will examine expression of subtypes of Fgfrl and Fgfr2 in this brain region. Finally, we will examine activation of ERK and AKT in response to alterations of Fgf8a-like or Fgf8b-like isoforms in chick and mouse embryos generated in the first two aims. These studies will provide new insights into the mechanisms for the regulation of Fgf8 function during development, which should have important implications for preventing human developmental disorders. Specific increase of FGF8 expression has been implicated in the onset and progression of various human cancers. Therefore, our proposed studies should shed light on abnormal regulation of Fgf8 splicing on pathological conditions, including tumor formation in human.

描述(申请人提供)：成纤维细胞生长因子8(FGF8)在控制各种发育过程中起着关键作用。FGF8结合并激活跨膜受体酪氨酸激酶(FGFR1-4)。两条主要的信号通路RAS-ERK和PI3K-Akt是已知的介导Fgf8信号的途径。在牛曼和小鼠中，单一的Fgf8基因通过选择性剪接产生多种异构体的可变N末端。Fgf8剪接变异体的生物学意义尚不清楚。以发育中的中脑和后脑为模型系统，研究了Fgf8的两种亚型Fgf8a和Fgf8b的诱导活性。我们最近发现，小鼠体内的所有八种Fgf8亚型都具有与Fgf8a和Fgf8b相似的活性，可分为两个功能基团，即类a(a，c，e和g)和类b(b，d，f和h)。需要检验的总体假设是，Fgf8的功能是由这两组Fgf8亚型的不同活动协调的。我们进一步推测，Fgf8的选择性剪接是调节Fgf8信号水平和范围的重要机制。围绕Fgf8在鸡和小鼠中脑和小脑发育中的作用，我们建议剖析Fgf8a和Fgf8b亚型的功能作用，以及这些亚型不同活动的机制和它们之间的相互作用。首先，为了确定不同Fgf8亚型在发育中/后脑中的组合活性，我们将使用功能增益方法，并检测单个或组合Fgf8亚型在鸡胚胎中的活性。其次，为了确定Fgf8a和Fgf8b亚型对正常发育的相对贡献，我们将使用功能丧失的方法，并确定Fgf8a和Fgf8b亚型在发育过程中的需求。第三，为了确定Fgf8亚型不同功能的分子机制，我们将研究Fgf8a和Fgf8b在脑外植体中的作用范围、扩散和功能特性。此外，为了确定Fgf8a和Fgf8b在中脑/后脑不同功能的机制，我们将检测Fgfr1和FGFR2亚型在该脑区的表达。最后，我们将检测ERK和AKT在前两个目标产生的鸡和小鼠胚胎中对类Fgf8a或类Fgf8b亚型改变的反应中ERK和AKT的激活。这些研究将为Fgf8在发育过程中的功能调节机制提供新的见解，这将对预防人类发育障碍具有重要意义。FGF8表达的特异性增加与多种人类癌症的发生和发展有关。因此，我们提出的研究将有助于揭示FGF8剪接在包括人类肿瘤形成在内的病理条件下的异常调控。