Buck Institute for Research on Aging, Novato, CA 94945, USA.
Free Radic Biol Med. 2012 Sep 1;53(5):1048-60. doi: 10.1016/j.freeradbiomed.2012.07.004. Epub 2012 Jul 13.
Oxidative stress is frequently implicated in the pathology of neurodegenerative disease. The chief source of this stress is mitochondrial respiration, via the passage of reducing equivalents through the respiratory chain resulting in a small but potentially pathological production of superoxide. The superoxide that is produced during normal respiration is primarily detoxified within the mitochondria by superoxide dismutase 2 (Sod2), a key protein for maintaining mitochondrial function. Mitochondria are distributed throughout the soma of neurons, as well as along neuronal processes and at the synaptic terminus. This distribution of potentially independent mitochondria throughout the neuron, at distinct subcellular locations, allows for the possibility of regional subcellular deficits in mitochondrial function. There has been increasing interest in the quantification and characterization of messages and proteins at the synapse, because of its importance in neurodegenerative disease, most notably Alzheimer disease. Here, we report the transcriptomic and proteomic changes that occur in synaptosomes from frontal cortices of Sod2 null mice. Constitutively Sod2 null mice were differentially dosed with the synthetic catalytic antioxidant EUK-189, which can extend the life span of these mice, as well as uncovering or preventing neurodegeneration due to endogenous oxidative stress. This approach facilitated insight into the quantification of trafficked messages and proteins to the synaptosome. We used two complementary methods to investigate the nature of the synaptosome under oxidative stress: either whole-genome gene expression microarrays or mass spectrometry-based proteomics using isobaric tagging for relative and absolute quantitation of proteins. We characterized the relative enrichment of gene ontologies at both gene and protein expression levels that occurs from mitochondrial oxidative stress in the synaptosome, which may lead to new avenues of investigation in understanding the regulation of synaptic function in normal and diseased states. As a result of using these approaches, we report for the first time an activation of the mTOR pathway in synaptosomes isolated from Sod2 null mice, confirmed by an upregulation of the phosphorylation of 4E-BP1.
氧化应激在神经退行性疾病的病理机制中经常被涉及。这种应激的主要来源是线粒体呼吸，通过还原当量穿过呼吸链传递，导致超氧化物的产生虽小但具有潜在的病理意义。在线粒体呼吸过程中产生的超氧化物主要通过超氧化物歧化酶 2（Sod2）在内部进行解毒，Sod2 是维持线粒体功能的关键蛋白。线粒体分布在神经元的胞体以及神经元突起和突触末端。这种分布在神经元中独立的线粒体的可能性，分布在不同的亚细胞位置，为线粒体功能在局部亚细胞水平上的缺陷提供了可能。由于其在神经退行性疾病（尤其是阿尔茨海默病）中的重要性，人们对突触中信息和蛋白质的定量和特征描述越来越感兴趣。在这里，我们报告了来自 Sod2 基因敲除小鼠额皮质突触体的转录组和蛋白质组的变化。Sod2 基因敲除小鼠被持续给予合成的催化抗氧化剂 EUK-189 进行不同剂量处理，EUK-189 可以延长这些小鼠的寿命，并揭示或预防由于内源性氧化应激引起的神经退行性病变。这种方法有助于深入了解突触体中运输信息和蛋白质的定量。我们使用两种互补的方法来研究氧化应激下的突触体的性质：一种是全基因组基因表达微阵列，另一种是基于质谱的蛋白质组学，使用等重标记相对和绝对定量蛋白质。我们在基因和蛋白质表达水平上描述了线粒体氧化应激引起的突触体中基因本体论的相对富集，这可能为理解正常和疾病状态下突触功能的调节开辟新的途径。由于使用了这些方法，我们首次报道了来自 Sod2 基因敲除小鼠的突触体中 mTOR 途径的激活，这一结果通过 4E-BP1 磷酸化的上调得到了证实。
