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

替代翻译的离子通道蛋白的新非典型作用

• 批准号: 9376618
• 负责人: Robin M Shaw
• 金额: $ 43.75万
• 依托单位: CEDARS-SINAI MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-07-01 至 2021-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9376618
• 关键词: Accounting; Actins; Adenoviruses; Adult; Arrhythmia; Biological; Biological Preservation; Biological Process; Biology; C-terminal; Cardiac; Cardiac Electrophysiologic Techniques; Cardiac Myocytes; Cardiovascular Diseases; Cardiovascular system; Cause of Death; Cell Communication; Cell Cycle Progression; Cell Maintenance; Cell Survival; Cell membrane; Cells; Cessation of life; Charge; Code; Connexin 43; Connexins; Critical Pathways; Data; Development; Disease; Distal; Epithelial; Exons; Failure; Gene Transfer; Genes; Genetic Transcription; Goals; Heart; Heart failure; Hydrogen Peroxide; Infarction; Initiator Codon; Injury; Intercalated disc; Ion Channel; Ion Channel Protein; Ischemia; Laboratories; Length; Mediating; Mesenchymal; Messenger RNA; Metabolism; Methionine; Mitochondria; Modeling; Morbidity - disease rate; Movement; Mus; Myocardial; Myocardial Ischemia; Myocardium; N-terminal; Organelles; Outcome; Oxidative Stress; Parents; Pathogenesis; Pathologic; Peptides; Phenotype; Process; Protein Isoforms; Protein Truncation; Proteins; RNA Splicing; Regulation; Reperfusion Therapy; Research; Ribosomes; Role; Science; Signal Transduction; Stress; Tail; Testing; Therapeutic; Transcript; Translating; Translations; Tropism; United States; United States National Institutes of Health; Ventricular; Work; Wound Healing; base; gap junction channel; gene therapy; improved; in vivo; migration; mortality; mouse model; new therapeutic target; novel therapeutic intervention; novel therapeutics; peptide drug; sudden cardiac death; targeted delivery; therapeutic development; tool; trafficking

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.

离子通道有过多的功能，往往不能理解他们的能力 插入质膜并传导离子电流。缝隙连接蛋白43（Cx43） 通道，其亲本基因是GJA 1，已被牵连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。GJA 1具有单个编码外显子，因此它不能进行剪接， 替代转录。但是我们已经确认，并且已经被几个 在实验室中，GJA 1 mRNA进行选择性翻译，其中核糖体 翻译不是发生在第一个AUG（蛋氨酸）起始密码子，而是发生在下游蛋氨酸。的 结果是，六个额外的进行性N-末端截短蛋白（均含有远端C-末端截短蛋白）被发现， 末端）可以从相同的mRNA分子内源性产生，我们通过蛋白质鉴定 大小（对于全长蛋白质，GJA 1 - 43 k，然后是GJA 1 - 32 k，GJA 1 - 29 k，GJA 1 - 26 k，GJA 1 - 20 k，GJA 1 - 11 k， 和GJA 1 - 7 k）。 在过去的三年里，我的团队一直在探索这些较小亚型的作用。这 该建议是基于一个令人惊讶的应用，坦率地说，惊人的发现，关于GJA 1 - 20 k 其通常是较小同种型中翻译最高的。我们发现GJA 1 - 20 k具有 对线粒体的强烈趋向性，如此强烈以至于它可以作为线粒体标记物。额外 初步数据表明GJA 1 - 20 k对心肌缺血的应激具有保护作用。我们 中心假设是，在氧化应激和缺血性损伤之后， 内源性GJA 1 - 20 k增加并靶向线粒体作为心肌存活信号。 此外，我们将测试外源性GJA 1 - 20 k是否可以减少缺血性心肌损伤， 侮辱。提出了三个具体目标。 目的#1-心肌损伤过程中GJA 1 - 20 k的表达、定位和功能是什么？ 目的#2-外源性GJA 1 - 20 k是否改善氧化应激后的心肌细胞存活？ 目的#3-GJA 1 - 20 k的基因转移是否减少心肌缺血小鼠模型中的梗死面积？ 当这三个目标的工作完成后，我们希望已经确定了关键的途径， GJA 1 - 20 k在应激过程中的调节，并为缺血性损伤引入了一种新的治疗肽。 此外，我们将发展一种范式，使替代翻译成为翻译的一个重要方面， 心脏电生理学，并应适用于多个通道作为一种手段，了解 其非典型功能的机制。