NEUROLIGIN AND AUTISM

这个子项目是许多利用 由NIH/NCRR资助的中心赠款提供的资源。子项目和 研究者（PI）可能从另一个NIH来源获得了主要资金， 因此可以在其他CRISP条目中表示。所列机构为 研究中心，而研究中心不一定是研究者所在的机构。 孤独症是一种广泛性障碍，与形成社会互动和交流的缺陷，刻板行为和不寻常的兴趣的发展有关。Jamain等人（2003）最近的一项研究报道，在患有自闭症谱系障碍（ASD）的同胞中发现了编码神经配蛋白-3（NL 3）和神经配蛋白-4（NL 4）的两个X连锁同源基因的突变。神经连接素，a/b-水解酶折叠家族的蛋白质，是突触后跨膜蛋白，与突触前伴侣b-神经毒素相关联。Neurexin和neuroligin似乎在突触连接处形成异源细胞接触，使它们成为控制早期脑发育过程中突触识别模式的合适候选者。本研究的目的是确定与ASD相关的人类NL 3-R451 C突变的生化作用。Arg 451在所有物种（从果蝇到人类）中高度保守，并且它位于预测的EF-手区域附近，怀疑该区域干扰神经胶质素-neurexin关联的Ca 2+依赖性。然而，使用FLAG-NL 3的截短的可溶形式，我们发现这两种突变体蛋白在我们的哺乳动物细胞系统中不表达为可溶性实体。我们推测突变体Cys 471的引入可能通过与7个天然存在的Cys之一产生新的二硫键来破坏蛋白质折叠。我们计划在稳定转染的HEK 293细胞中使用免疫荧光观察野生型和突变全长蛋白质的细胞内运输，以确定两种蛋白质（天然靶向细胞膜）是否遵循相同的运输途径。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Autism is a pervasive disorder associated with deficits in forming social reciprocal interaction and communications, development of stereotyped behavior and unusual interests. A recent study by Jamain et al. (2003) reports that mutations of two X-linked homologous genes encoding neuroligin-3 (NL3) and neuroligin-4 (NL4) were found in siblings with autism spectrum disorders (ASD). Neuroligins, proteins of the a/b-hydrolase fold family, are postsynaptic transmembrane proteins that associate with presynaptic partners, the b-neurexins. Neurexin and neuroligin appear to form heterologous cell contacts at synaptic connections, making them suitable candidates for controlling synaptic recognition patterns during early brain development.The aim of this study is to determine the biochemical role of the human NL3-R451C mutation associated with the ASD. Arg451 is highly conserved among all species (from fruit fly to human) and it is located near a predicted EF-hand region, where is suspected to perturb the Ca2+ dependence of the neuroligin-neurexin association. However, using a truncated soluble form of FLAG-NL3 we found that both mutant proteins were not expressed as soluble entities by our mammalian cell system. We hypothesize that the introduction of mutant Cys471 might disrupt the protein folding by creating a new disulfide bond with one of the 7 naturally occurring Cys. We plan to visualize the intracellular trafficking of both wild type and mutant full length proteins using immunofluorescence in stably transfected HEK293 cells in order to determine if both proteins, naturally targeted to the cell membrane, follow the same trafficking pathway.