Role of Fibulin-1 in APP Processing

描述（由申请人提供）：目前许多对抗阿尔茨海默病（AD）的药理学方法试图通过抑制淀粉样蛋白生成酶（称为β-和β-）来阻断A β的产生。分泌酶另一种方法是激活淀粉样前体蛋白（APP）的β-分泌酶加工，这是由金属蛋白酶解整合素家族的几个成员，ADAM 9，ADAM 10和ADAM 17介导的。这些β-分泌酶对APP的加工被认为对AD有益，因为它限制了A β的产生，并产生了神经保护性可溶性APP β（sAPP β）产物。Fibulin-1（Fbln 1）是一种细胞外基质蛋白，在脑中由神经元表达， APP和sAPP β的氨基末端。这种相互作用的意义尚未确定，然而，我们发现Fbln 1也结合其他膜锚定β-分泌酶底物，肝素结合表皮生长因子（HB-EGF）和神经调节蛋白-1（NRG 1）。我们还表明，Fbln 1的行为，以抑制HB-EGF和NRG 1的可溶性形式的蛋白水解释放。此外，我们发现与野生型MEFs相比，Fbln 1缺失小鼠胚胎成纤维细胞（MEFs）的条件培养基中sAPP β水平增加。基于这些发现，假设Fbln 1作为APP的β-分泌酶加工的抑制剂，因此可能代表一种治疗靶点，如果被抑制，可能导致APP的β-分泌酶加工增强和病理性APP切割减少。为了解决这一假设，有三个具体的目标：1）确定在Fbln 1缺陷小鼠中脑APP蛋白水解切割是否改变，2）确定Fbln 1的转基因过表达是否加速A β产生并加剧AD发病机制，3）确定Fbln 1是否抑制培养细胞中APP的β分泌酶加工。

DESCRIPTION (provided by applicant): Many current pharmacological approaches to combating Alzheimer's disease (AD) seek to block A� production through inhibition of the amyloidogenic enzymes known as �- and ?-secretases. An alternative approach is to activate the �-secretase processing of amyloid precursor protein (APP), which is mediated by several members of the disintegrin family of metalloproteases, ADAM9, ADAM10 and ADAM17. Processing of APP by these �-secretases is thought to be beneficial with respect to AD since it limits production of A� and generates the neuroprotective soluble APP� (sAPP�) product. Fibulin-1 (Fbln1) is an extracellular matrix protein, expressed in the brain by neurons, that binds the amino terminus of APP and sAPP�. The significance of this interaction is not yet established however, we have found that Fbln1 also binds to other membrane anchored �-secretase substrates, heparin binding-epidermal growth factor (HB-EGF) and neuregulin-1 (NRG1). We also show that Fbln1 acts to inhibit the proteolytic release of soluble forms of HB-EGF and NRG1. Furthermore, we have found increased levels of sAPP� in the conditioned culture medium of Fbln1 null mouse embryo fibroblasts (MEFs) as compared to wildtype MEFs. Based on these findings it is hypothesized that Fbln1 serves as an inhibitor of �-secretase processing of APP and therefore may represent a therapeutic target that if inhibited might lead to augmented �-secretase processing of APP and reduced pathological APP cleavage. To address this hypothesis there are three specific aims: 1) Determine whether brain APP proteolytic cleavage is altered in Fbln1-deficient mice, 2) determine whether transgenic overexpression of Fbln1 accelerates A� production and exacerbates AD pathogenesis, and 3) determine whether Fbln1 inhibits �-secretase processing of APP in cultured cells.