p62, Ref(2)P and ubiquitinated proteins are conserved markers of neuronal aging, aggregate formation and progressive autophagic defects
p62, Ref(2)P and ubiquitinated proteins are conserved markers of neuronal aging, aggregate formation and progressive autophagic defects
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DOI：
10.4161/auto.7.6.14943
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发表时间：
2011-06-01
期刊：
影响因子：
13.3
通讯作者：
Finley, Kim D.
中科院分区：
文献类型：
--
作者：
Bartlett, Bryan J.;Isakson, Pauline;Finley, Kim D.
由于突变或通过与衰老相关的过程而抑制大自噬，导致通常由该途径消除的细胞质底物的积累。这在长寿命细胞如神经元中是一个重要问题，其中通路缺陷可导致含有泛素化蛋白质的聚集体的积累。p62/Ref（2）P蛋白家族参与细胞质蛋白体或隔离体的自噬清除。这些独特的结构与含有泛素的蛋白质内含物以及自噬途径的关键组分密切相关。在这项研究中，我们表明，洗涤剂分级分离，然后进行蛋白质印迹分析的不溶性泛素化蛋白（IUP），哺乳动物p62及其果蝇同源物，参考（2）P可以用来定量评估活动水平的聚集体清除（aggrephagy）在复杂的组织。使用这种技术，我们表明，遗传或年龄依赖性的变化，修改的长期增强或抑制的aggrephagy可以确定。此外，使用果蝇模型系统，该方法可用于建立在广泛的生理条件下发生的自噬依赖性蛋白质清除谱，所述生理条件包括发育、禁食和改变的代谢途径。该技术还可用于检查与人类疾病相关的蛋白质病，例如额颞叶痴呆、亨廷顿和阿尔茨海默病。我们的研究结果表明，测量IUP概况连同p62/Ref（2）P蛋白的评估可用作筛选或诊断工具，以表征改变自噬的长期功能和复杂组织和细胞内发生的蛋白质聚集体的清除的遗传和年龄依赖性因素。
Suppression of macroautophagy, due to mutations or through processes linked to aging, results in the accumulation of cytoplasmic substrates that are normally eliminated by the pathway. This is a significant problem in long-lived cells like neurons, where pathway defects can result in the accumulation of aggregates containing ubiquitinated proteins. The p62/Ref(2)P family of proteins is involved in the autophagic clearance of cytoplasmic protein bodies or sequestosomes. These unique structures are closely associated with protein inclusions containing ubiquitin as well as key components of the autophagy pathway. In this study we show that detergent fractionation followed by western blot analysis of insoluble ubiquitinated proteins (IUP), mammalian p62 and its Drosophila homologue, Ref(2)P can be used to quantitatively assess the activity level of aggregate clearance (aggrephagy) in complex tissues. Using this technique we show that genetic or age-dependent changes that modify the long-term enhancement or suppression of aggrephagy can be identified. Moreover, using the Drosophila model system this method can be used to establish autophagy-dependent protein clearance profiles that are occurring under a wide range of physiological conditions including developmental, fasting and altered metabolic pathways. This technique can also be used to examine proteopathies that are associated with human disorders such as frontotemporal dementia, Huntington and Alzheimer disease. Our findings indicate that measuring IUP profiles together with an assessment of p62/Ref(2)P proteins can be used as a screening or diagnostic tool to characterize genetic and age-dependent factors that alter the long-term function of autophagy and the clearance of protein aggregates occurring within complex tissues and cells.