MODULATION OF AMYLOID PRECURSOR PROTEIN METABOLISM BY X11ALPHA/MINT-1

神经元衔接蛋白X11 alpha/mint-1在体外与淀粉样前体蛋白（APP）相互作用以调节其运输和加工。在转染的非神经元细胞中，X11 alpha抑制APP的α-和γ-裂解，但不抑制β-裂解。由于X11 alpha的磷酸酪氨酸结合（PTB）结构域与APP家族特异性相互作用，X11 alpha似乎特异性抑制APP的γ-裂解，同时保留Notch和其他受调节的膜内蛋白水解底物的γ-裂解。为了确定这些体外数据的体内意义，我们将产生和表征新型hX 11 α转基因小鼠和X11 α敲除小鼠，并检查小鼠APP代谢。将这些小鼠与Tg 2576小鼠（hAPP的瑞典突变或hAPP swe的转基因）杂交，将探测Xi la对hAPP swe代谢和对老化脑中部分AD样表型的发展的影响。最近的一项人类全基因组分析显示，散发性AD与9号染色体上的单核苷酸多态性（SNP）存在显着联系，可能包括X11 α区域。 我们将通过使用从AD受试者和仔细匹配的对照受试者的样本中提取的基因组DNA来探测X11 α区域的SNP与散发性AD的遗传连锁。具体目标是：1）产生X11 α敲除和hX 11 α转基因小鼠，并确定对脑和原代神经元培养物中的鼠APP代谢的影响，2）将X11 α敲除和hX 11 α转基因小鼠与Tg 2576小鼠杂交，以确定a）对脑和神经元培养物中的hAPP代谢的调节作用，和B）AD样表型随年龄的发展，和3）阐明AD病例和年龄、性别和ApoE匹配的对照的X11 α基因内或附近的SNP和单倍型，以确定是否存在与散发性AD的统计学显著联系。具体目标3将研究从病理核心和其他ADRC获得的DNA样本。这些结果将1）通知正常功能 2）为在hAPP转基因小鼠模型中使用X11 α或其PTB结构域进行基于病毒载体的AD基因治疗奠定基础; 3）探索散发性AD的潜在遗传危险因素。

The neuronal adaptor protein X11alpha/mint-1 interacts with amyloid precursor protein (APP) to regulate its trafficking and processing in vitro. In transfected non-neuronal cells, X11alpha inhibits alpha- and gamma- but not beta- cleavage of APP. Because the phosphotyrosine binding (PTB) domain of X11alpha interacts specifically with the APP family, X11alpha appears to inhibit gamma-cleavage of APP specifically while sparing gamma-cleavage of Notch and other substrates of regulated intramembranous proteolysis. To determine the in vivo significance of these in vitro data, we will generate and characterize novel hX11alpha transgenic mice and X11alpha knockout mice and examine murine APP metabolism. Crosses of these mice to Tg2576 mice (transgenic for the Swedish mutation of hAPP, or hAPPswe) will probe the effects of XI la on hAPPswe metabolism and on the development of partial AD-like phenotypes in aging brain. A recent human genome-wide analysis revealed significant linkage of sporadic AD to single nucleotide polymorphisms (SNPs) on chromosome 9, perhaps including the X11alpha region. We will probe genetic linkage of SNPs in the X11alpha region to sporadic AD by using genomic DNA extracted from samples from AD subjects versus carefully-matched control subjects. The specific aims are to: 1) generate X11alpha knockout and hX11alpha transgenic mice and determine effects on murine APP metabolism in brain and in primary neuronal cultures, 2) cross X11alpha knockout and hX11alpha transgenic mice with Tg2576 mice to determine a) modulatory effects on hAPPswe metabolism in brain and in neuronal cultures, and b) development of AD-like phenotypes with aging, and 3) elucidate the SNPs and haplotypes within or adjacent to the X11alpha gene of AD cases and age-, gender-, and ApoE-matched controls to determine if there is a statistically significant link to sporadic AD. Specific aim 3 will study DNA samples obtained from the Pathology Core and from other ADRCs. These results will 1) inform the normal functions of X11alpha as well as APP and its derivatives in CNS neurons, 2) lay the groundwork for viral-vector based gene therapy of AD using either X11alpha or its PTB domain in hAPP transgenic mouse models, and 3) probe a potential genetic risk factor of sporadic AD.