Quantification of stable isotope tracers after metabolic labeling provides a snapshot of the dynamic state of living cells and tissue. A form of imaging mass spectrometry quantifies isotope ratios with a lateral resolution <50 nm, using a methodology that we refer to as multi-isotope imaging mass spectrometry (MIMS). Despite lateral resolution exceeding diffraction-limited light microscopy, lack of contrast has largely limited use of MIMS to large or specialized subcellular structures, such as the nucleus and stereocilia. In this study, we repurpose the engineered peroxidase APEX2 as the first genetically encoded marker for MIMS. Coupling APEX2 labeling of lysosomes and metabolic labeling of protein, we identify that individual lysosomes exhibit substantial heterogeneity in protein age, which is lost in iPSC-derived neurons lacking the lysosomal protein progranulin. This study expands the practical use of MIMS for cell biology by enabling measurements of metabolic function from stable isotope labeling within individual organelles in situ.

中文翻译：

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将 APEX 标记与单个细胞器的成像质谱联用揭示了溶酶体蛋白质周转的异质性

代谢标记后稳定同位素示踪剂的定量提供了活细胞和组织动态状态的快照。成像质谱法的一种形式使用我们称为多同位素成像质谱法(MIMS)的方法来量化横向分辨率＜50nm的同位素比率。尽管横向分辨率超过了衍射极限光学显微镜，但对比度的缺乏在很大程度上限制了 MIMS 在大型或特殊亚细胞结构（例如细胞核和静纤毛）中的使用。在这项研究中，我们将工程过氧化物酶 APEX2 重新用作 MIMS 的第一个基因编码标记。将溶酶体的 APEX2 标记和蛋白质的代谢标记结合起来，我们发现各个溶酶体在蛋白质年龄方面表现出很大的异质性，而这种异质性在缺乏溶酶体蛋白颗粒体蛋白前体的 iPSC 衍生神经元中消失。这项研究通过对单个细胞器内的稳定同位素标记进行原位代谢功能测量，扩展了 MIMS 在细胞生物学中的实际应用。