CELL STRETCH AND CARDIAC HYPERTROPHY

细胞拉伸和心脏肥大

• 批准号: 2220429
• 负责人: PETER A WATSON
• 金额: $ 3.51万
• 依托单位: GEISINGER CLINIC
• 依托单位国家: 美国
• 财政年份: 1990
• 资助国家: 美国
• 起止时间: 1990-01-01 至 1995-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2220429
• 关键词: adenylate cyclase; biological signal transduction; cyclic AMP; hormone regulation /control mechanism; isolation perfusion; laboratory rat; mechanical stress; mechanoreceptors; molecular pathology; muscle stress; polymerase chain reaction; protein biosynthesis; stretch reflex; tissue /cell culture; ventricular hypertrophy

Pressure-induced stretch in the heart, both in vivo and in vitro, has been shown to stimulate anabolic processes including increased rates of ribosome and protein synthesis. In perfused rat hearts, stretch-induced anabolic responses occur concurrent with and are dependent upon accumulation of the intracellular second messenger adenosine 3', 5'- cyclic monophosphate (cAMP) and activation of cAMP-dependent protein kinase. It has been the continuing goal of this project to identify the molecular requirements for accumulation of the intracellular second messenger cAMP in response to cell deformation. Prior to the results from work funded by this grant, no information was available regarding the isolation and purification of the specific mechanosensitive element of any mechanoresponsive signal transduction pathway. Work in a simple cell model, hypotonic swelling of S49 mouse lymphoma cells, has delineated the components of a signal transduction pathway responsible for cAMP accumulation following cell deformation. Mechanical forces stimulate cAMP accumulation through direct acceleration of adenylyl cyclase activity. This stimulation appears to occur independent of the action of the guanine nucleotide-binding regulatory proteins involved in hormonal regulation of adenylyl cyclase activity. Currently no information is available as to the structure/function relationships which impart the capacity to transduce deformation stimuli into the generation of an intracellular signal. It is the goal of the work proposed in this continuation application to obtain structure/function information for adenylyl cyclase. Recent work has shown that adenylyl cyclase exists in multiple isoforms in mammalian cells, with no specific isoform thus far being exclusively found in mechanoresponsive tissues or cells. Initial work on the continuation of this project has involved identifying, cloning, and sequencing of isoforms from mechanoresponsive cells. Once these and other isoforms are cloned, they are expressed in cells which lack mechanoresponsive cAMP accumulation and tested to determine the conservation of the mechanoresponsive phenotype in the adenylyl cyclase family. Thus far, Type I and Type III isoforms have been tested, with only Type I demonstrating mechanoresponsive behavior. Once isoforms are sequenced and tested for mechanoresponsiveness, conserved and divergent structures will be identified and domains selected which correlate to this property. Genetic manipulation of these conserved domains and subsequent expression of mutated adenylyl cyclases in the cell systems described above will be used to test the participation of these domains in mechanoresponsive behavior.

在体内和体外，心脏中的压力诱导拉伸具有 被证明可以刺激合成代谢过程，包括增加 核糖体和蛋白质合成。 在灌流的大鼠心脏中，牵拉诱导 合成代谢反应同时发生并依赖于 细胞内第二信使腺苷3 '，5'- 环磷酸（cAMP）和cAMP依赖性蛋白的活化 激酶。 本项目的持续目标是确定 细胞内第二信使积累的分子要求 信使cAMP对细胞变形的反应。 在结果出来之前 从这项赠款资助的工作中，没有关于 特异性机械敏感元件的分离纯化 任何机械反应信号转导途径的一部分。 工作在一个简单的 细胞模型，S49小鼠淋巴瘤细胞的低渗肿胀， 描绘了一个信号转导通路的组成部分， 细胞变形后cAMP的积累。 机械力 通过直接加速腺苷酸来刺激cAMP积累 环化酶活性 这种刺激似乎独立于 鸟嘌呤核苷酸结合调节蛋白参与 腺苷酸环化酶活性的激素调节。 目前没有 关于结构/功能关系的信息是可用的， 将变形刺激的能力赋予一代， 一种细胞内信号。 这是本文提出的工作目标， 继续申请，以获得结构/功能信息， 腺苷酸环化酶 最近的研究表明，腺苷酸环化酶存在于 哺乳动物细胞中存在多种亚型，迄今为止没有特异性亚型 仅存在于机械反应组织或细胞中。 初始 继续开展这一项目的工作包括确定， 克隆和对来自机械应答细胞的同种型进行测序。 一旦 克隆这些和其它同种型，它们在细胞中表达， 缺乏机械反应性cAMP积累，并进行测试以确定 腺苷酸环化酶机械反应表型的保守性 家人 到目前为止，已经测试了I型和III型同种型， 只有I型表现出机械反应行为。 一旦同种型 测序并测试机械反应性、保守性和差异性 将识别结构并选择与以下相关的域： 这个财产。 这些保守结构域的遗传操作， 随后在细胞系统中表达突变的腺苷酸环化酶 将使用上述方法测试这些域的参与情况 机械反应行为。