Auditory Neurogenetics

描述（由申请人提供）：听觉转导主要依赖于Atp2b2基因（编码PMCA2蛋白）。我们的实验室和其他实验室已经在小鼠和人类基因中发现了几种突变（Tempel and Shilling, 2007）。我们已经开发了一个突变的Atp2b2突变的聋哑小鼠等位基因系列，使我们能够研究耳蜗中功能性PMCA2的逐步减少的影响。我们使用这个等位基因序列来证明杂合性部分功能突变缺失是单倍不足的（McCullough and Tempel, 2004），并提供了一个年龄相关性听力损失（AHL）的模型（McCullough and Tempel, 2005; Walker et al., 2008）。在这篇文章中，我们将通过检测聋哑人等位基因中的噪声性听力损失（NIHL）来增加我们对PMCA2在耳蜗中的作用的了解。我们将开发一种过表达PMCA2的新小鼠系，看看这些小鼠是否对AHL或NIHL具有抗性。我们还将研究PMCA2在听觉脑干神经元中的作用。我们假设PMCA2在调节这些快速放电神经元中的钙（Ca2+）中起关键作用；这是一个以前未被发现的作用，因为研究主要集中在PMCA2在毛细胞中的功能上。最后，我们将开发一个细胞表达系统来研究PMCA2的功能。我们将使用PMCA2的半胱氨酸扫描诱变来确定巯基试剂在细胞外应用时可以快速阻断Ca2+泵活性的位点。除了提供药物靶向的潜在位点外，这应该为研究PMCA2在毛细胞（HC）转导和神经元传递中的细胞作用提供了一种方法。在一个日益嘈杂和寿命延长的社会中，了解与年龄相关的听力损失和噪音引起的听力损失的基因是很重要的。我们对质膜钙泵的研究表明，它定位于听觉毛细胞的静纤毛，当其功能因突变而降低时，小鼠或人类都有明显的听力损失。我们正在研究钙泵是如何工作的，希望能找到使它保护毛细胞免受损害的方法，从而防止听力丧失。

DESCRIPTION (provided by applicant): Auditory transduction depends critically on the Atp2b2 gene (encoding the PMCA2 protein). Our lab and others have identified several mutations in mouse and human genes (reviewed in Tempel and Shilling, 2007). We have developed an allelic series of deafwaddler mouse mutants with altered Atp2b2 that allow us to study the effects of stepwise reductions of functional PMCA2 in the cochlea. We have used this allelic series to show that heterozygous partial loss of function mutations are haplo-insufficient (McCullough and Tempel, 2004) and provide a model of age-related hearing loss (AHL) (McCullough and Tempel, 2005; Walker et al., 2008). In this proposal we will add to our knowledge of the role of PMCA2 in the cochlea by examining noise-induced hearing loss (NIHL) in alleles of deafwaddler. We will develop a new line of mice that over-express PMCA2 to see if these mice are resistant to AHL or NIHL. We will also study the role of PMCA2 in neurons of the auditory brainstem. We hypothesize that PMCA2 plays a critical role in regulating calcium (Ca2+) in these fast-firing neurons; a role previously unexplored because studies have focused on PMCA2 function in hair cells. Finally, we will develop a cellular expression system for studying PMCA2 function. We will use cysteine-scanning mutagenesis of PMCA2 to define sites where thiol reagents, when applied extracellularly can rapidly block Ca2+ pump activity. Besides providing insight into potential sites for drug targeting, this should provide a way to study the cellular role of PMCA2 in hair cell (HC) transduction and neuronal transmission. In an increasingly noisy and longer-lived society, knowledge of the genes contributing to age-related hearing loss and noise induced hearing loss is important. Our work on the plasma membrane calcium pump shows that it is localized to the stereocilia of the auditory hair cells and that when its function is reduced by mutations in mouse or human, they have significant hearing loss. We are studying how the calcium pump works, hoping to learn ways to make it protect hair cells from damage, thereby preventing hearing loss.