A New Non-Canonical Role for an Alternatively Translated Ion Channel Protein

离子通道有太多的功能，通常不能通过它们的能力来理解 插入质膜并传导离子电流。缝隙连接蛋白43(Cx43) 其母基因为GJA1的Channel被认为在细胞周期进程中发挥作用， 转移转化，伤口愈合，发育，迁移，上皮-间充质转化， 和新陈代谢。GJA1只有一个编码外显子，因此不能进行剪接和 可供选择的转录。然而，我们已经确认，这已经得到了几个人的确认 实验室认为，GJA1 mRNA可以进行替代翻译，在这种翻译中，核糖体的起始 翻译不是发生在第一个AUG(蛋氨酸)起始密码子，而是在下游的蛋氨酸。这个 结果是另外六个递增的N-末端截断蛋白(都包含远端的C- 末端)可以从相同的mRNA分子中内源产生，我们通过蛋白质来识别 大小(全长蛋白GJA1-43k，GJA1-32k，GJA1-29k，GJA1-26k，GJA1-20k，GJA1-11k， 和GJA1-7K)。 在过去的三年里，我的团队一直在探索这些较小的异构体的作用。这 该提案是基于一项令人惊讶的、坦率地令人震惊的关于GJA1-20k的发现 它通常是较小的异构体中翻译程度最高的。我们发现GJA1-20k有一个 对线粒体有很强的趋向性，可以作为线粒体的标志物。其他内容 初步数据表明，GJA1-20k对心肌缺血应激具有保护作用。我们的 中心假说是氧化应激和缺血性损伤之后，交替转换 内源性GJA1-20k增加，并以线粒体为靶点作为心肌生存信号。 此外，我们还将测试外源性GJA1-20k是否能减轻缺血所致的心肌损伤 侮辱。提出了三个具体目标。 目的#1-GJA1-20k在心肌损伤中的表达、定位和功能是什么？ 目的#2-外源性GJA1-20k是否能改善氧化应激后心肌细胞的存活？ 目的#3-GJA1-20k基因转移是否能减少心肌缺血小鼠模型中的梗塞范围？ 当三个目标中的工作完成时，我们预计已经确定了 GJA1-20k在应激过程中的调节，并推出了一种新的治疗缺血性损伤的多肽。 此外，我们还将开发替代翻译的一个重要方面的范式 心脏电生理学，并应应用于多个通道作为一种手段，以了解 它们的非正则功能的机制。

Ion channels have a plethora of functions than, frequently, can not be understood by their ability to insert in the plasma membrane and conduct ionic current. The Connexin43 (Cx43) gap junction channel, whose parent gene is GJA1, has been implicated in having a role in cell cycle progression, metastatic transformation, wound healing, development, migration, epithelial-mesenchymal transition, and metabolism. GJA1 has a single coding exon, and therefore it can not be subject to splicing and alternative transcription. However we have identified, and it has been confirmed by several laboratories, that GJA1 mRNA is subject to alternative translation in which initiation of ribosomal translation occurs not at the first AUG (Methionine) start codon, but at downstream Methionines. The result is that six additional progressive N-terminal truncation proteins (all containing the distal C- terminus) can be endogenously generated from the same mRNA molecule, which we identify by protein size (GJA1-43k for the full length protein, then GJA1-32k, GJA1-29k, GJA1-26k, GJA1-20k, GJA1-11k, and GJA1-7k). For the last three years, my group has explored the role of these smaller isoforms. This proposal is based on the application of a surprising, and frankly startling, finding regarding GJA1-20k which is usually the most highly translated of the smaller isoforms. We found that GJA1-20k has a strong tropism for mitochondria, so strong that it could function as a mitochondrial marker. Additional preliminary data indicate that GJA1-20k is protective against the stress of myocardial ischemia. Our central hypothesis is that following oxidative stress and ischemic injury, alternatively translated endogenous GJA1-20k is increased and targets to mitochondria as a myocardial survival signal. Furthermore, we will test whether exogenous GJA1-20k can reduce myocardial injury due to ischemic insult. Three specific aims are proposed. Aim #1- What is the expression, localization, and function of GJA1-20k during myocardial injury? Aim #2- Does exogenous GJA1-20k improves cardiomyocyte survival following oxidative stress? Aim #3- Does gene transfer of GJA1-20k reduce infarct size in a mouse model of myocardial ischemia? When the work in the three aims is complete, we expect to have identified critical pathways of GJA1-20k regulation during stress and have introduced a new therapeutic peptide for ischemic injury. Furthermore, we will develop the paradigm by which alternative translation is an important aspect of cardiac electrophysiology and should be applied to multiple channels as a means for understanding the mechanism of their non-canonical function.