Cell-type-specific metabolic labeling of nascent proteomes in vivo
Cell-type-specific metabolic labeling of nascent proteomes in vivo
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DOI：
10.1038/nbt.4016
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发表时间：
2017-12-01
影响因子：
46.9
通讯作者：
Schuman, Erin M.
中科院分区：
文献类型：
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作者：
Alvarez-Castelao, Beatriz;Schanzenbaecher, Christoph T.;Schuman, Erin M.
虽然蛋白质标记方法的进步已经使测量混合细胞群体的蛋白质组成为可能，但在体内分离特定细胞类型的蛋白质组是不可能的。这是因为现有的体内代谢蛋白标记方法可以接触到所有类型的细胞。我们报道了一种转基因小鼠系的发展，其中Crerecombinase诱导突变的蛋氨酰-tRNA合成酶(L274G)的表达使得能够用非规范的氨基酸和点击化学对新生蛋白进行细胞类型特异性标记。利用免疫印迹、成像和质谱仪，我们使用我们的转基因小鼠在体外(脑片)和体内标记和分析兴奋性主神经元和浦肯野神经元中的蛋白质。我们通过将小鼠暴露在具有丰富感觉线索的环境中，在海马区兴奋性神经元中发现了200多种差异调节的蛋白质。我们的方法可以用于分离、分析和定量健康和疾病组织中特定细胞类型的蛋白质组及其动力学。
Although advances in protein labeling methods have made it possible to measure the proteome of mixed cell populations, it has not been possible to isolate cell-type-specific proteomes in vivo. This is because the existing methods for metabolic protein labeling in vivo access all cell types. We report the development of a transgenic mouse line where Crerecombinase-induced expression of a mutant methionyl-tRNA synthetase (L274G) enables the cell-type-specific labeling of nascent proteins with a non-canonical amino-acid and click chemistry. Using immunoblotting, imaging and mass spectrometry, we use our transgenic mouse to label and analyze proteins in excitatory principal neurons and Purkinje neurons in vitro (brain slices) and in vivo. We discover more than 200 proteins that are differentially regulated in hippocampal excitatory neurons by exposing mice to an environment with enriched sensory cues. Our approach can be used to isolate, analyze and quantitate cell-type-specific proteomes and their dynamics in healthy and diseased tissues.