Department of Pharmacology, University of California at San Diego, La Jolla, CA 92093, USA.
Proc Natl Acad Sci U S A. 2013 Feb 19;110(8):E726-35. doi: 10.1073/pnas.1300424110. Epub 2013 Feb 4.
Postnatal bilateral whisker trimming was used as a model system to test how synaptic proteomes are altered in barrel cortex by sensory deprivation during synaptogenesis. Using quantitative mass spectrometry, we quantified more than 7,000 synaptic proteins and identified 89 significantly reduced and 161 significantly elevated proteins in sensory-deprived synapses, 22 of which were validated by immunoblotting. More than 95% of quantified proteins, including abundant synaptic proteins such as PSD-95 and gephyrin, exhibited no significant difference under high- and low-activity rearing conditions, suggesting no tissue-wide changes in excitatory or inhibitory synaptic density. In contrast, several proteins that promote mature spine morphology and synaptic strength, such as excitatory glutamate receptors and known accessory factors, were reduced significantly in deprived synapses. Immunohistochemistry revealed that the reduction in SynGAP1, a postsynaptic scaffolding protein, was restricted largely to layer I of barrel cortex in sensory-deprived rats. In addition, protein-degradation machinery such as proteasome subunits, E2 ligases, and E3 ligases, accumulated significantly in deprived synapses, suggesting targeted synaptic protein degradation under sensory deprivation. Importantly, this screen identified synaptic proteins whose levels were affected by sensory deprivation but whose synaptic roles have not yet been characterized in mammalian neurons. These data demonstrate the feasibility of defining synaptic proteomes under different sensory rearing conditions and could be applied to elucidate further molecular mechanisms of sensory development.
产后双侧胡须修剪被用作模型系统,以测试在突触发生期间感觉剥夺如何改变桶状皮层中的突触蛋白组。使用定量质谱法,我们定量了超过 7000 种突触蛋白,并在感觉剥夺的突触中鉴定出 89 种显著减少的和 161 种显著增加的蛋白质,其中 22 种通过免疫印迹进行了验证。超过 95%的定量蛋白,包括 PSD-95 和 gephyrin 等丰富的突触蛋白,在高和低活性饲养条件下没有显著差异,这表明兴奋性或抑制性突触密度没有全组织范围的变化。相比之下,几种促进成熟棘突形态和突触强度的蛋白质,如兴奋性谷氨酸受体和已知的辅助因子,在剥夺的突触中显著减少。免疫组织化学显示,突触后支架蛋白 SynGAP1 的减少主要局限于感觉剥夺大鼠的桶状皮层 I 层。此外,蛋白降解机制,如蛋白酶体亚基、E2 连接酶和 E3 连接酶,在剥夺的突触中显著积累,表明在感觉剥夺下靶向突触蛋白降解。重要的是,该筛选鉴定了其水平受感觉剥夺影响但在哺乳动物神经元中其突触作用尚未被表征的突触蛋白。这些数据证明了在不同感觉饲养条件下定义突触蛋白组的可行性,并且可以应用于进一步阐明感觉发育的分子机制。
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