RNAseq and quantitative proteomic analysis of Dictyostelium knock-out cells lacking the core autophagy proteins ATG9 and/or ATG16

BMC Genomics

Gernot Glöckner¹  Ludwig Eichinger¹  Roman Konertz¹  Sarah Fischer¹  Prerana Wagle²  Changxin Wu³  Ping Li³  Qiuhong Xiong³  Ning Song³ 

[1] Center for Biochemistry, Medical Faculty, University of Cologne, Joseph-Stelzmann-Str. 52, 50931, Cologne, Germany;Cologne Excellence Cluster on Cellular Stress Responses in Aging Associated Diseases, University of Cologne, Joseph-Stelzmann-Str. 26, 50931, Cologne, Germany;Institutes of Biomedical Sciences, Key Laboratory of Chemical Biology and Molecular Engineering of Ministry of Education, Shanxi University, No. 92 Wucheng Road, 030006, Taiyuan, China;

关键词: Autophagy;    ATG9;    ATG16;    RNA;    proteome;    tandem mass tag (TMT);    parallel reaction monitoring (PRM);    Dictyostelium discoideum;   

DOI  :  10.1186/s12864-021-07756-2

来源: Springer

【 摘 要 】

BackgroundAutophagy is an evolutionary ancient mechanism that sequesters substrates for degradation within autolysosomes. The process is driven by many autophagy-related (ATG) proteins, including the core members ATG9 and ATG16. However, the functions of these two core ATG proteins still need further elucidation. Here, we applied RNAseq and tandem mass tag (TMT) proteomic approaches to identify differentially expressed genes (DEGs) and proteins (DEPs) in Dictyostelium discoideum ATG9‾, ATG16‾ and ATG9‾/16‾ strains in comparison to AX2 wild-type cells.ResultIn total, we identified 332 (279 up and 53 down), 639 (487 up and 152 down) and 260 (114 up and 146 down) DEGs and 124 (83 up and 41 down), 431 (238 up and 193 down) and 677 (347 up and 330 down) DEPs in ATG9‾, ATG16‾ and ATG9‾/16‾ strains, respectively. Thus, in the single knock-out strains, the number of DEGs was higher than the number of DEPs while in the double knock-out strain the number of DEPs was higher. Comparison of RNAseq and proteomic data further revealed, that only a small proportion of the transcriptional changes were reflected on the protein level. Gene ontology (GO) analysis revealed an enrichment of DEPs involved in lipid metabolism and oxidative phosphorylation. Furthermore, we found increased expression of the anti-oxidant enzymes glutathione reductase (gsr) and catalase A (catA) in ATG16‾ and ATG9‾/16‾ cells, respectively, indicating adaptation to excess reactive oxygen species (ROS).ConclusionsOur study provides the first combined transcriptome and proteome analysis of ATG9‾, ATG16‾ and ATG9‾/16‾ cells. Our results suggest, that most changes in protein abundance were not caused by transcriptional changes, but were rather due to changes in protein homeostasis. In particular, knock-out of atg9 and/or atg16 appears to cause dysregulation of lipid metabolism and oxidative phosphorylation.

【 授权许可】

CC BY