HuR Regulation of HSP 70 in Brain Reperfusion

描述(申请人提供)：德格雷西亚实验室的长期目标是研究脑缺血/再灌注(L/R)损伤的基本细胞病理学，为开发有效的临床治疗方法提供基础。L/R损伤是卒中、心脏骤停和复苏相关的发病率和死亡率的基础。目前还没有有效的治疗方法来阻止脑L再灌注后神经元的死亡。在这里，我们提出了一种新的机制来调节70 kDa可诱导的热休克蛋白HSP70。L耐药神经元(如海马区CA3)在再灌流早期合成HSP70蛋白。L/R易损神经元(海马CA1区)转录丰富的HSP70mRNA，但不能合成HSP70蛋白。我们的初步数据显示，mRNA稳定蛋白HUR的核输出与HSP70的翻译相关。HUR结合含有ARE序列的mRNAs，HSP70的mRNA含有ARE序列。为了评价Hur在调节HSP70合成中的作用，我们提出了以下目标：1.确定Hur和HSP70在再灌流后海马区的亚细胞定位。这一目标将决定HSP70 mRNA的亚细胞位置是否与HUR的位置相关，以及HSP70 mRNA在抗缺血的CA1神经元和易缺血的CAS神经元中的定位是否不同。2.确定HUR核出口是否与HSP70翻译存在因果关系。5‘-腺苷一磷酸活化蛋白激酶(AMPK)的激活会导致HUR的核滞留，并恶化中风的预后。HUR亚细胞定位、HSP70翻译和细胞存活将在药物激活或抑制AMPK的情况下在缺血和再灌流后确定。 公共卫生相关性：每年有数千人死于缺血性脑损伤。这些研究将增加我们对中风或心脏骤停后大脑受损原因的理解，并可能有助于治疗方法的开发。

DESCRIPTION (provided by applicant): The long-term goal of the DeGracia laboratory is to investigate the basic cellular pathology of ischemia/reperfusion (l/R) brain injury to provide a foundation to develop effective clinical therapies. l/R injury underlies the morbidity and mortality associated with stroke and cardiac arrest and resuscitation. There are currently no effective therapies to halt neuronal death following brain l/R. Here, we propose a novel mechanism of regulation of the 70 kDa inducible heat shock protein HSP70. l/R-resistant neurons (e.g. hippocampal CA3), synthesize HSP70 protein early in reperfusion. l/R-vulnerable neurons (hippocampal CAI), transcribe ample hsp70 mRNA, but fail to synthesize the HSP70 protein. Our preliminary data shows nuclear export of the mRNA-stabilizing protein HuR correlates with HSP70 translation. HuR binds ARE-containing mRNAs, and hsp70 mRNA contains an ARE sequence. To assess the role of HuR in regulating HSP70 synthesis, we propose the following aims: 1. To determine the subcellular localization of HuR and hsp70 mRNA in the reperfused hippocampus. This aim will determine whether the subcellular location of hsp70 mRNA correlates with the location of HuR and whether the location of hsp70 mRNA differs between ischemia-resistant CA1 and ischemia-vulnerable CAS neurons. 2. To determine if HuR nuclear export is causally linked to HSP70 translation. 5' adenosine monophosphate- activated protein kinase (AMPK) activation causes nuclear retention of HuR and worsens stroke outcome. HuR subcellular localization, HSP70 translation, and cell survival will be determined following ischemia and reperfusion in the presence of pharmacologic activation or inhibition of AMPK. Public Heath Relevance: Thousands die from ischemic brain injury annually. These studies will increase our understanding of why the brain is damaged following stroke or cardiac arrest and potentially contribute to development of treatments.