REGULATION OF ION CHANNELS BY APOPTOSIS TRIGGER PROTEINS

细胞凋亡或程序性细胞死亡可由多种刺激诱导。细胞凋亡被认为与许多生理和临床现象有关，包括缺血性损伤、多种形式的癌症和神经退行性疾病，如阿尔茨海默病、肌萎缩侧索硬化症和一些普恩相关疾病。已鉴定出许多诱导细胞凋亡的刺激物和基因产物。然而，这些刺激触发细胞死亡程序的分子机制还知之甚少。这里提出的研究计划将集中在凋亡触发基因Reaper如何促进电刺激细胞中的细胞死亡。在细胞死亡程序开始前1到2小时，死神基因在注定要经历细胞凋亡的细胞中表达。通过对Reaper蛋白和“A”型电压依赖性钾通道(SHB)N端序列的比较发现，Reaper蛋白可以作为一个失活球来阻断钾通道。我们的初步结果确实表明，全长的Reaper蛋白通过诱导非常稳定的失活来阻断电压依赖的钾通道。我们的初步结果还表明，收割机减缓了电压依赖性钠通道的失活过程。这项拟议的研究计划将研究收割机蛋白如何改变电压依赖性离子通道的功能特性，以促进细胞凋亡。中心假设是Reaper改变了质膜离子通道的性质以促进去极化，这种去极化促进了细胞凋亡。我们将使用非洲爪哇卵母细胞中异源表达的离子通道以及天然哺乳动物的离子通道来验证这一假设。我们将研究诱导表达收割机如何影响哺乳动物细胞的离子通道属性和膜电位。这项拟议研究的预期结果将确定一些凋亡触发蛋白如何促进细胞死亡，这些结果可能有助于设计治疗措施来治疗涉及细胞凋亡的疾病。

Apoptosis or programmed cell death can be induced by a variety of stimuli. Apoptosis is considered to be involved in many physiological and clinical phenomena, including ischemic injury, many forms of cancer, and neurodegenerative diseases, such as Alzheimer's disease, amyotrophic lateral sclerosis, and some prion-related disorders. Many apoptosis-inducing stimuli and gene products have been identified. The molecular mechanisms by which these stimuli trigger the cell-death program are, however, only poorly understood. The research program proposed here will focus on how an apoptosis-trigger gene, reaper, promotes cell death in electrically excitable cells. The reaper gene is expressed in cells destined to undergo apoptosis 1 to 2 hours before the cell death program starts. A sequence comparison of the N-termini of Reaper and a "A-type" voltage-dependent potassium channel (ShB) shows that the Reaper protein could work as an inactivation ball to block potassium channels. Our preliminary results indeed show that the full-length Reaper protein blocks voltage-dependent potassium channels by inducing very stable inactivation. Our preliminary results also show that Reaper slows down the inactivation process of voltage-dependent sodium channels. The proposed research program will examine how the Reaper protein alters functional properties of voltage-dependent ion channels to promote apoptosis. The central hypothesis is that Reaper alters properties of the plasma membrane ion channels to promote depolarization and that this depolarization facilitates apoptosis. We will test this hypothesis using heterologously expressed ion channels in Xenopus oocytes as well as native mammalian channels. We will examine how induced expression Reaper affects the ion channel properties and membrane potential in mammalian cells. The results expected from the proposed research will establish how some apoptosis-trigger proteins promote cell-death and the results may be useful in designing therapeutic interventions to treat the diseases that involve apoptosis.