Characterization of the interaction between Mmp2 Frac and during axon guidance

描述(申请人提供)：癌症和神经退行性疾病通常以基质金属蛋白酶(MMPs)表达的剧烈变化为标志。这些蛋白水解酶已被研究为治疗多种疾病的潜在靶点。不幸的是，24种不同的脊椎动物MMPs的临床试验一直受到副作用的阻碍，这表明MMPs对正常的生理功能很重要。果蝇体内只有两个MMPs，在很大程度上消除了冗余和补偿问题，是研究MMPs功能的一个很好的模型系统。我们最近报道了果蝇MMPs在胚胎发育过程中对轴突的束/束起重要作用。要了解Frac在神经元发育过程中的功能，关键是要确定它的蛋白分解底物(S)。在酵母双杂交筛选中，已经确定了一个候选靶标，致命吸引(FRAC)。在这个方案中，我们建议表征Frac在轴突引导中的功能，并确定它是否受MMP2的调节。Frc蛋白与脊椎动物Matrlin和纤维蛋白同源。人类中纤维蛋白的丢失会导致马凡或比尔综合征，这表明这些研究可能与临床相关。初步结果表明，Frc RNA定位于胚胎肌肉，这一位置与轴突引导的作用一致。在肌肉中Frc的错误表达导致轴突导向缺陷，这与MMP2功能丧失(LOF)突变体中报告的去卷曲表型无法区分，有力地支持了Frac和MMP2之间的功能联系。我们假设Frac在肌肉中扮演着粘连分子的角色，将轴突吸引到他们的目标肌肉上，远离神经分支。然后，MMP2将负责切割和灭活Frac。在特定的目标1中，我们将通过分析FRAC LOF突变表型和FRAC与MMP2之间的遗传相互作用来表征FRAC在轴突引导中的功能。特定目的2测试Frac是否被处理，如果是，则测试该切割是否依赖于MMP2。由于这项提议涉及一种可能参与轴突引导决策的新的MMP2底物的特征，它可能有助于揭示MMPs及其底物在脊椎动物神经元发育中的正常生理功能。

DESCRIPTION (provided by applicant): Cancers and neurodegenerative disorders are often marked by dramatic changes in the expression of matrix metalloproteinases (MMPs). These proteases have been investigated as potential therapeutic targets to treat a variety of diseases. Unfortunately, with 24 different vertebrate MMPs, clinical trials have been hindered by side effects, suggesting that MMPs are important for normal physiological functions. There are only two MMPs in Drosophila, making it an excellent model system to study MMP function as the problems of redundancy and compensation are largely eliminated. We recently reported that Drosophila MMPs are important for the fasciculation/bundling of axons during embryonic development. To understand Frac function during neuronal development, it is critical to identify its proteolytic substrate(s). A candidate target, Fatal Attraction (Frac), has been identified in a yeast two-hybrid screen. In this proposal, we propose to characterize the function of Frac in axon guidance and determine whether it is regulated by Mmp2. Frac protein is homologous to the vertebrate Matrilin and Fibrillin protein. Loss of fibrillin in humans leads to Marfan or Beals syndrome indicating that these studies are likely to be relevant clinically. Preliminary results indicate that frac RNA is localized to embryonic muscle, a position consistent with a role in axon guidance. Frac misexpression in muscle results in axon guidance defects that are indistinguishable from the defasciculation phenotype reported in Mmp2 loss-of-function (LOF) mutants strongly supporting a functional link between Frac and Mmp2. We hypothesize that Frac acts as an adhesive molecule in muscle to attract axons to their target muscles and away from nerve branches. Mmp2 would then be responsible for cleaving and inactivating Frac. In Specific Aim 1, we will characterize the function of Frac in axon guidance by assaying the frac LOF mutant phenotype and genetic interactions between frac and Mmp2. Specific Aim 2 tests whether Frac is processed and, if so, whether this cleavge is Mmp2-dependent. As this proposal addresses the characterization of a novel Mmp2 substrate likely involved in axon guidance decisions, it may shed light on the normal physiological functions of MMPs and their substrates in vertebrate neuronal development.