GSK3b mediates convergence of protection signaling to limit mitochondrial damage

GSK3b 介导保护信号汇聚以限制线粒体损伤

基本信息

批准号：
7732328
负责人：
Steven J Sollott
金额：
$ 62.61万
依托单位：
NATIONAL INSTITUTE ON AGING
依托单位国家：
美国
项目类别：
财政年份：
资助国家：
美国
起止时间：
至
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/7732328
关键词：
70-kDa Ribosomal Protein S6 Kinases Age Area Biological Preservation Brain Calcium-Activated Potassium Channel Cardiac Cardiac Death Cardiac Myocytes Cell Death Cell Survival Cells Class Clinical Clinical Treatment Complex Cyclic AMP-Dependent Protein Kinases Cytoprotection Defect Development Diazoxide Exhibits Family member G-Protein-Coupled Receptors Glycogen Synthase Kinase 3 Goals Heart Hypoxia Infarction Injury Insulin Ischemic Preconditioning Lead Leucine-2-Alanine Enkephalin Mediating Memory Mitochondria Mitochondrial Swelling Mitotic Mus Muscle Cells Myocardial Infarction Nature Neurons Neuroprotective Agents Opioid Peptide Oxidants Oxidation-Reduction Pathway interactions Permeability Phosphorylation Phosphotransferases Potassium Channel Production Proteins Reactive Oxygen Species Receptor Activation Receptor Protein-Tyrosine Kinases Resistance Signal Pathway Signal Transduction Small Interfering RNA Stress Stroke Stroke prevention Swelling Transgenic Mice burden of illness cariporide design healthy aging human FRAP1 protein inhibitor/antagonist insight knock-down mitochondrial K(ATP) channel mitochondrial permeability transition pore novel oxidation preconditioning prevent response size

项目摘要

Environmental stresses converge on the mitochondria that can trigger or inhibit cell death. Excitable, post-mitotic cells (such as cardiac myocytes in heart, and neurons in brain), in response to sub-lethal noxious stress engage mechanisms affording protection from subsequent insults. These protection mechanisms involve activation of endogenous signaling which can confer significant resistance to oxidant and other stresses associated with hypoxia/reoxygenation (i.e., during a heart attack or stroke), which promotes the enhanced capacity for cell survival. However, the upstream signaling mechanisms have remained an area of active debate, and the end effector(s) have remained unsolved. We show that reoxygenation after prolonged hypoxia reduces the reactive oxygen species- (ROS-) threshold for the mitochondrial permeability transition (mPTP) in cardiac myocytes, and that cell survival is steeply negatively correlated with the fraction of depolarized mitochondria. We demonstrate that a wide variety of cardio/neuroprotective agents acting via distinct upstream mechanisms all promote cell survival by limiting mPTP induction. We found that protection can be triggered in 2 general ways dependent and independent of regulatory mitochondrial swelling which converge via inhibition of GSK-3b on the end effector, the permeability transition pore complex, preventing the mPTP. Cell protection exhibiting a memory (i.e., "preconditioning") results from triggered mitochondrial swelling (due to enhanced K+ accumulation via influx and/or retention) causing enhanced substrate oxidation and ROS production, leading to redox activation of PKC which in turn inhibits GSK-3b (via phosphorylation of ser-9). Both the diazoxide-activated mitochondrial ATP dependent K+ channel (mitoKATP, which we have identified and are now chararcterizing), and the Ca2+-activated K+ channel, for example, can serve as mitochondrial K+ influx mechanisms (that can mediate mitochondrial regulatory swelling-dependent protection, etc). The delta-opioid peptide, DADLE, and the NHE-inhibitors, HOE 642 (cariporide) and HOE 694, each produce mitochondrial regulatory swelling-dependent protection independently of the mitoKATP. Alternatively, receptor tyrosine kinase or certain G-protein coupled receptor activation elicits cell protection (without mitochondrial swelling or durable memory) by inhibiting GSK-3b, via either PKB/Akt and mTOR/p70s6k, PKC, or PKA pathways. Examples of this latter class include insulin (via Akt and mTOR/p70s6k) and the direct GSK-3 inhibitor, Li+. We found that siRNA knock-down of GSK-3b, but not GSK-3a, induced the protection state, and that transgenic mice expressing a cardiac-restricted, constitutively active, non-inhibitable form of GSK-3b (GSK-3b S9A) were resistant to a whole battery of upstream signals that were effective to induce the protected state in WT mice. We concluded that GSK-3b (and specifically its inactivation) is a major, required integration point for a multitude of upstream signals acting on an end-effector responsible for cardioprotection (the mitochondrial permeability transition pore). When cell protection signaling pathways are activated, we found that the Bcl-2 family members relay the signal from GSK-3b onto a target at or in close proximity to the pore. Thus, the effect of the convergence of these signaling pathways via inhibition of GSK-3b, relayed through Bcl-2 proteins, on the end effector, the permeability transition pore complex, to limit mPTP induction, is the general mechanism of cardiomyocyte protection. We propose that clinical treatment strategies designed to inhibit the master switch kinase, GSK-3b, to protect the permeability transition pore complex from mPTP induction, would be effective to reduce the size of infarction during episodes of heart attack or stroke by preventing the death of cardiac myocytes and neurons (respectively). Signaling defects underlying the age-assocciated loss of the capacity for ischemic preconditioning are being examined which could lead to testable clinical therapies relevant to the preservation of healthy aging.

环境应力会在线粒体上汇聚，该线粒体会触发或抑制细胞死亡。可激发的，可激发的有丝分裂后细胞（例如心脏中的心肌细胞和大脑中的神经元），以响应亚洲性有害压力的响应，从而使机制具有保护，从而可以保护免受随后的侮辱。这些保护机制涉及激活内源性信号传导，该信号传导可以赋予对氧化剂的显着抗性和与缺氧/重氧相关的其他应力（即心脏病发作或中风期间），从而促进了细胞存活的增强能力。但是，上游信号传导机制仍然是主动争论的领域，末端效应子仍未解决。我们表明，长时间缺氧后的重氧降低了心肌细胞中线粒体通透性过渡（MPTP）的活性氧（ROS-）阈值，并且该细胞的存活与地下细胞体的脱极层的分数陡峭地呈负相关。我们证明，通过不同上游机制作用的各种心脏/神经保护剂都通过限制MPTP诱导来促进细胞存活。我们发现，可以以两种一般方式触发保护，并独立于调节线粒体肿胀，这些线粒体肿胀通过抑制GSK-3B对末端效应器的抑制（渗透性过渡孔复合物）融合，从而防止MPTP。细胞保护表现出记忆（即“预处理”）是由触发的线粒体肿胀引起的（由于涌入和/或保留的K+积累增强而导致底物氧化和ROS产生增强，从而导致PKC的氧化还原激活，从而导致PKC的氧化还原激活，从而通过Ser-3b抑制了GSK-3B（通过Ser-9的磷酸化）。二氮氧化物激活的线粒体ATP依赖性K+通道（MitokATP，我们已经确定并正在炭化），例如，Ca2+激活的K+通道（例如，可以用作线粒体K+涌入机制）（可以介导线粒体调节性膨胀依赖性依赖性依赖性依赖性依赖性依赖）。 Delta-Apeid肽，Dadle和NHE抑制剂，HOE 642（Cariporide）和HOE 694，每种都会产生线粒体调节性肿胀依赖性依赖性依赖性保护，独立于MitokATP。另外，通过抑制PKB/AKT和MTOR/P70S6K，PKC，PKC或PKA通路，通过抑制GSK-3B，受体酪氨酸激酶或某些G蛋白偶联受体激活会引起细胞保护（无线粒体肿胀或耐用记忆）。后一类的例子包括胰岛素（通过AKT和MTOR/P70S6K）和直接的GSK-3抑制剂LI+。 We found that siRNA knock-down of GSK-3b, but not GSK-3a, induced the protection state, and that transgenic mice expressing a cardiac-restricted, constitutively active, non-inhibitable form of GSK-3b (GSK-3b S9A) were resistant to a whole battery of upstream signals that were effective to induce the protected state in WT mice. 我们得出的结论是，GSK-3B（尤其是其失活的）是主要的，所需的集成点，用于作用于负责心脏保护的最终效应器（线粒体渗透性过渡孔）的众多上游信号。当激活细胞保护信号通路时，我们发现Bcl-2家族成员将信号从GSK-3B传递到孔或近距离接近孔的目标。因此，通过抑制GSK-3B的抑制，通过Bcl-2蛋白继电器对末端效应子（渗透性过渡孔复合物）限制MPTP诱导的融合的影响，是心肌细胞保护的一般机制。我们建议旨在抑制主开关激酶GSK-3B的临床治疗策略，以保护渗透性过渡孔复合物免受MPTP诱导的影响，将有效地减少心脏病发作或中风期间梗塞的大小，从而通过防止心肌细胞和神经元的死亡（分别可以防止心脏肌细胞和神经元）（分别防止）。正在检查与年龄相关的缺血性预处理损失背后的信号缺陷，这可能会导致与保存健康衰老有关的可测试临床疗法。