NON-APOPTOTIC ACTION OF CASPASES IN PLURIPOTENT EMBRYONIC STEM CELLS

半胱天冬酶在多能胚胎干细胞中的非凋亡作用

基本信息

批准号：
8269736
负责人：
Thomas P. Zwaka
金额：
$ 30.09万
依托单位：
BAYLOR COLLEGE OF MEDICINE
依托单位国家：
美国
项目类别：
财政年份：
2008
资助国家：
美国
起止时间：
2008-08-01 至 2014-05-31
项目状态：
已结题

项目摘要

Embryonic stem (ES) cells are pluripotent and can expand in vitro without any apparent limits, while retaining their ability to become any type of cell in the body. The long-term goal of this proposal is to link the molecular mechanisms of programmed cell death to those underlying ES cell renewal and differentiation, with a view toward accelerating the clinical introduction of ES cell regenerative medicine. In my preliminary studies, I found that caspase-3, an important mediator of programmed cell death, has an unexpected role in controlling ES cell fate. I demonstrate an increase of caspase-3 activity upon induction of differentiation and show that caspase-3 can directly cleave the Nanog transcription factor, leading to rapid loss of this core pluripotency- related protein and subsequent ES cell differentiation it typically mediates. These results suggest that caspase- 3 and perhaps other key components of the programmed cell death pathway may have an integral role in the regulation of ES cell renewal/differentiation. The central hypothesis of this proposed work is that classical mediators of programmed cell death, especially caspase-3, also mediate the fate decisions affecting pluripotent stem cells. In Aim 1 I will dissect the functional roles of caspase-3 and its activating caspase in the fate of ES cells. In Aim 2 I will modulate caspase activity in ES cells and assess the effects on self-renewal, differentiation and programmed cell death. I will also address the question of whether the differentiation-promoting activity of caspase-3 in ES cells is due to an instructive or selective signaling and elucidate if caspase activity provides a specific signal to differentiate or simply promotes differentiation in general. In Aim 3 I will assess the importance of caspase-3-mediated cleavage of Nanog in ES cell differentiation. I consider the transcription factor Nanog to be a paradigm for other potential caspase targets in ES cells, so that my findings for this regulatory protein could well extend to other transcriptional pathways involved in ES cell differentiation. Results of the experiments described in this proposal are expected to provide insight into the pleiotropic effects of caspases in pluripotent stem cells. Thus, specific pharmacological alteration of caspases may be useful not only for modulating apoptosis, but also for directing stem cell fate. The involvement of caspases in nonapoptotic pathways suggests that efforts to block apoptosis via caspase inhibition could have much broader consequences than initially thought. The ability of embryonic stem cells to remain undifferentiated in culture while retaining the ability to become any cell within the human body make them an invaluable tool for use in transplant medicine, drug discovery, and understanding basic developmental biology. Results of the experiments described in this proposal are expected to provide insight into the pleiotropic effects of the cell death enzyme caspase on the differentiation process of embryonic stem cells. Specific pharmacological alteration of caspases may be useful not only for modulating programmed cell death, but also for directing stem cell fate.

胚胎茎（ES）细胞具有多能，可以在体外扩展而无明显限制，同时保持它们成为体内任何类型的细胞的能力。该提议的长期目标是将分子联系起来对基础ES细胞更新和分化的人的编程细胞死亡机制，为了加速ES细胞再生医学的临床引入。在我的初步研究中，我发现caspase-3是程序性细胞死亡的重要介体，在控制中具有意想不到的作用 ES细胞命运。我证明了caspase-3活性在诱导后的增加，并表明 caspase-3可以直接裂解纳米转录因子，从而导致这种核心多能的迅速丧失 - 相关蛋白质和随后的ES细胞分化通常会介导。这些结果表明caspase- 3也许是程序性细胞死亡途径的其他关键组成部分可能在 ES细胞更新/分化的调节。这项拟议工作的核心假设是，编程细胞死亡的经典介体，尤其是 caspase-3，还介导影响多能干细胞的命运决定。在AIM 1中，我将剖析功能 caspase-3的作用及其在ES细胞命运中的激活caspase。在AIM 2中，我将调节caspase活动在ES细胞中，评估对自我更新，分化和程序性细胞死亡的影响。我也会解决了ES细胞中caspase-3的分化促进活性的问题启发性或选择性信号传导并阐明caspase活动是否提供了特定的信号来区分或一般而言，简单地促进分化。在AIM 3中，我将评估caspase-3介导的重要性 ES细胞分化中纳米的切割。我认为转录因子nanog是 ES细胞中其他潜在的caspase靶标，以便我对这种调节蛋白的发现可以很好地延伸到 ES细胞分化涉及的其他转录途径。预计本提案中描述的实验结果将提供对多效效应的见解多能干细胞中的胱天蛋白酶。因此，胱天蛋白酶的特定药理改变可能是有用的仅用于调节细胞凋亡，还用于指导干细胞命运。胱天蛋白酶参与非凋亡途径表明，通过caspase抑制阻止凋亡的努力可能具有很多比最初想象的更广泛的后果。胚胎干细胞在培养中保持未分化的能力，同时保持变为人体内的任何细胞都使它们成为用于移植药物，药物发现的宝贵工具，并了解基本的发展生物学。本提案中描述的实验结果是预计将深入了解细胞死亡酶caspase对分化的多效性影响胚胎干细胞的过程。 caspases的特定药理改变不仅对调节程序性细胞死亡，但也用于指导干细胞命运。