Short-term mild temperature-stress-induced alterations in C. elegans phosphoproteome

Exposure to mild early-life stresses can slow down the aging process, in which protein phosphorylation might be an essential regulator. Currently, the understanding of phosphorylation-based regulatory networks under mild early-life stress remains elusive. Herein, we systematically analyzed the phosphoproteomes of C. elegans, which were treated with three kinds of mild temperature (15°C, 20°C, and 25°C) from two different short-term groups (10 min and 60 min). By utilizing iTRAQ-based quantitative phosphoproteomic approach, a total of 18187 phosphorylation sites from 3330 proteins were identified. Volcano plots illustrated that the phosphorylation abundance of 374 proteins (15°C) and 347 proteins (25°C), were significantly changed, respectively. Gene ontology, KEGG pathway and protein-protein interaction network analysis revealed that these phosphoproteins were mainly involved in metabolism, translation, development, and lifespan determination. Motif analysis of kinase substrates suggested that MAPK, CK and CAMK were most likely involved in the adaption processes. Moreover, 16 and 14 aging-regulated proteins underwent phosphorylation modifications under the mild stress of 15°C and 25°C, respectively, indicating these proteins might be important for maintaining long-term health. Further experiments of lifespan confirmed that the candidate phosphoproteins, e.g.; EGL-27, XNP-1, and GTBP-1 could regulate lifespan at 15°C, 20°C, and 25°C, and showed increased tolerance to heat and oxidative stresses. In summary, our findings provided a wealth of datasets to better understand the phosphorylation mechanism of mild early-life stresses in C. elegans.