AggreCount: an unbiased image analysis tool for identifying and quantifying cellular aggregates in a spatially defined manner.
AggreCount: an unbiased image analysis tool for identifying and quantifying cellular aggregates in a spatially defined manner.
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DOI：
10.1074/jbc.ra120.015398
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发表时间：
2020-12-18
期刊：
影响因子：
--
通讯作者：
Raman M
中科院分区：
文献类型：
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作者：
Klickstein JA;Mukkavalli S;Raman M
蛋白质的质量控制是由许多整合的细胞途径来维持的，这些途径监测细胞蛋白质组的折叠和功能。这些途径中的缺陷导致错误折叠或有缺陷的蛋白质的积累，这些蛋白质可能随着时间的推移变得不可溶并聚集。蛋白质聚集体显著促进许多人类疾病的发展，例如肌萎缩性侧索硬化症、亨廷顿病和阿尔茨海默病。在体外，基于成像的细胞研究已经定义了识别和清除聚集体的关键生物分子组分;然而，没有统一的方法可用于量化细胞聚集体，限制了我们重复和准确量化这些结构的能力。在这里，我们描述了一个名为AggreCount的ImageJ宏，用于识别和测量细胞中的蛋白质聚集体。AggreCount被设计为直观，易于使用，并可针对细胞中观察到的不同类型的聚集体进行定制。使用脚本需要最少的编码经验。基于用户定义的图像，AggreCount将报告许多指标：（i）细胞聚集体的总数，（ii）具有聚集体的细胞的百分比，（iii）每个细胞的聚集体，（iv）聚集体的面积，以及（v）聚集体的定位（胞质溶胶，核周或核）。为进一步的数据分析，提供了一个以每个单元格为基础的聚合信息数据表以及一个汇总表。我们证明了AggreCount的多功能性，通过分析一些不同的细胞聚集体，包括侵略，应力颗粒，和包涵体引起的亨廷顿多聚谷氨酰胺的扩张。
Protein quality control is maintained by a number of integrated cellular pathways that monitor the folding and functionality of the cellular proteome. Defects in these pathways lead to the accumulation of misfolded or faulty proteins that may become insoluble and aggregate over time. Protein aggregates significantly contribute to the development of a number of human diseases such as amyotrophic lateral sclerosis, Huntington's disease, and Alzheimer's disease. In vitro, imaging-based, cellular studies have defined key biomolecular components that recognize and clear aggregates; however, no unifying method is available to quantify cellular aggregates, limiting our ability to reproducibly and accurately quantify these structures. Here we describe an ImageJ macro called AggreCount to identify and measure protein aggregates in cells. AggreCount is designed to be intuitive, easy to use, and customizable for different types of aggregates observed in cells. Minimal experience in coding is required to utilize the script. Based on a user-defined image, AggreCount will report a number of metrics: (i) total number of cellular aggregates, (ii) percentage of cells with aggregates, (iii) aggregates per cell, (iv) area of aggregates, and (v) localization of aggregates (cytosol, perinuclear, or nuclear). A data table of aggregate information on a per cell basis, as well as a summary table, is provided for further data analysis. We demonstrate the versatility of AggreCount by analyzing a number of different cellular aggregates including aggresomes, stress granules, and inclusion bodies caused by huntingtin polyglutamine expansion.