Comprehensive characterization of the Hsp70 interactome reveals novel client proteins and interactions mediated by posttranslational modifications.
Comprehensive characterization of the Hsp70 interactome reveals novel client proteins and interactions mediated by posttranslational modifications.
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DOI：
10.1371/journal.pbio.3001839
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发表时间：
2022-10
期刊：
影响因子：
9.8
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中科院分区：
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热休克蛋白70的相互作用是至关重要的细胞活力和应激反应。以前的尝试，表征热休克蛋白70相互作用的限制，其短暂的性质和目前的技术无法区分直接与桥接的相互作用。我们报告的新用途的交联质谱（XL-MS），全面表征酿酒酵母（芽殖酵母）Hsp 70蛋白相互作用组。使用这种方法，我们已经获得了对Hsp 70功能的基本新见解，包括Hsp 70自缔合以及与其客户蛋白的多点相互作用的明确证据。除了确定一组新的直接热休克蛋白70相互作用，可用于探测分子伴侣在细胞中的功能，我们还确定了一套翻译后修饰（PTM）相关的热休克蛋白70的相互作用。这些PTM中的大多数以前没有报道过，并且似乎在客户蛋白质功能的调节中至关重要。这些数据表明，PTM有助于蛋白质功能的机制之一是通过促进与分子伴侣的相互作用。两者合计，我们建议，XL-MS分析伴侣复合物可作为一种独特的方式来确定客户端蛋白的生物重要的PTM。分子伴侣Hsp 70与其客户蛋白之间的相互作用对于细胞活力和应激反应至关重要。本蛋白质组学研究使用交联质谱法来鉴定酵母中Hsp 70的新型直接相互作用物，并表明翻译后修饰在这些相互作用中起重要作用。
Hsp70 interactions are critical for cellular viability and the response to stress. Previous attempts to characterize Hsp70 interactions have been limited by their transient nature and the inability of current technologies to distinguish direct versus bridged interactions. We report the novel use of cross-linking mass spectrometry (XL-MS) to comprehensively characterize the Saccharomyces cerevisiae (budding yeast) Hsp70 protein interactome. Using this approach, we have gained fundamental new insights into Hsp70 function, including definitive evidence of Hsp70 self-association as well as multipoint interaction with its client proteins. In addition to identifying a novel set of direct Hsp70 interactors that can be used to probe chaperone function in cells, we have also identified a suite of posttranslational modification (PTM)-associated Hsp70 interactions. The majority of these PTMs have not been previously reported and appear to be critical in the regulation of client protein function. These data indicate that one of the mechanisms by which PTMs contribute to protein function is by facilitating interaction with chaperones. Taken together, we propose that XL-MS analysis of chaperone complexes may be used as a unique way to identify biologically important PTMs on client proteins. Interactions between the molecular chaperone Hsp70 and its client proteins are critical for cellular viability and the response to stress. This proteomics study uses cross-linking mass spectrometry to identify novel direct interactors of Hsp70 in yeast and suggests that post-translational modifications play an important role in these interactions.