Sharma, Kiran Bala and Sharma, Manish and Aggarwal, Suruchi and Yadav, Amit Kumar and Bhatnagar, Shinjini and Vrati, Sudhanshu and Kalia, Manjula and Cristea, Ileana M. (2019) Quantitative Proteome Analysis of Atg5-Deficient Mouse Embryonic Fibroblasts Reveals the Range of the Autophagy-Modulated Basal Cellular Proteome. mSystems, 4 (6). ISSN 2379-5077

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Abstract

Basal autophagy is crucial for maintenance of cellular homeostasis. ATG5 is an essential protein for autophagosome formation, and its depletion has been extensively used as a tool to disrupt autophagy. Here, we characterize the impact of Atg5 deficiency on the cellular proteome of mouse embryonic fibroblasts (MEFs). Using a tandem mass tagging (TMT)-based quantitative proteomics analysis, we observe that 14% of identified proteins show dysregulated levels in atg5/ MEFs. These proteins were distributed across diverse biological processes, such as cell adhesion, development, differentiation, transport, metabolism, and immune responses. Several of the upregulated proteins were receptors involved in transforming growth factor (TGF-) signaling, JAK-STAT signaling, junction adhesion, and interferon/cytokine-receptor interactions and were validated as autophagy substrates. Nearly equal numbers of proteins, including several lysosomal proteins and enzymes, were downregulated, suggesting a complex role of autophagy/ATG5 in regulating their levels. The atg5/ MEFs had lower levels of key immune sensors and effectors, including Toll-like receptor 2 (TLR2), interferon regulatory factor 3 (IRF3), IRF7, MLKL, and STAT1/3/5/6, which were restored by reexpression of ATG5. While these cells could efficiently mount a type I interferon response to the doublestranded RNA (dsRNA) mimic poly(I·C), they were compromised in their inflammatory response to the bacterial pathogen-associated molecular patterns (PAMPs) lipopolysaccharide (LPS) and Pam3CSK4. Transcriptional activation and secretion of interleukin-6 (IL-6) in these cells could be recovered by ATG5 expression, supporting the role of autophagy in the TLR2-induced inflammatory response. This study provides a key resource for understanding the effect of autophagy/ATG5 deficiency on the fibroblast proteome.

Item Type:	Article
Subjects:	Biomedical Science
Depositing User:	RCB Library
Date Deposited:	14 Mar 2023 09:55
Last Modified:	14 Mar 2023 09:55
URI:	http://rcb.sciencecentral.in/id/eprint/257

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