Metabolic and transcriptome responses of RNAi-mediated AMPKα knockdown in Tribolium castaneum

Abstract

Background The AMP-activated protein kinase (AMPK) is an intracellular fuel sensor for lipid and glucose metabolism. In addition to the short-term regulation of metabolic enzymes by phosphorylation, AMPK may also exert long-term effects on the transcription of downstream genes through the regulation of transcription factors and coactivators. In this study, RNA interference (RNAi) was conducted to investigate the effects of knockdown of TcAMPKα on lipid and carbohydrate metabolism in the red flour beetle, Tribolium castaneum, and the transcriptome profiles of dsTcAMPKα-injected and dsEGFP-injected beetles under normal conditions were compared by RNA-sequencing. Results RNAi-mediated suppression of TcAMPKα increased whole-body triglyceride (TG) level and the ratio between glucose and trehalose, as was confirmed by in vivo treatment with the AMPK-activating compound, 5-Aminoimidazole-4-carboxamide1-β-D-ribofuranoside (AICAR). A total of 1184 differentially expressed genes (DEGs) were identified between dsTcAMPKα-injected and dsEGFP-injected beetles. These include genes involved in lipid and carbohydrate metabolism as well as insulin/insulin-like growth factor signaling (IIS). Real-time quantitative polymerase chain reaction analysis confirmed the differential expression of selected genes. Interestingly, metabolism-related transcription factors such as sterol regulatory element-binding protein 1 (SREBP1) and carbohydrate response element-binding protein (ChREBP) were also significantly upregulated in dsTcAMPKα-injected beetles. Conclusions AMPK plays a critical role in the regulation of beetle metabolism. The findings of DEGs involved in lipid and carbohydrate metabolism provide valuable insight into the role of AMPK signaling in the transcriptional regulation of insect metabolism.

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“…The above harvested PGs were subjected to quantitative determination of the concentrations of trehalose, pyruvic acid, and acyl-CoA by using the concentration determination kits (A149-1-1, A076-1-1, and A081 kit from Nanjing Jiancheng Bioengineering Institute, respectively). The detailed steps followed the manufacturer's suggestions 1 and the previous reports (Du et al, 2017;Jiang et al, 2020;Xu et al, 2020).…”

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“…Furthermore, AMPK directly interacts with phosphorylates SREBP1 and represses SREBP1 target gene, leading to reduced lipogenesis. 34–36 The phosphorylation of AMPK suppresses the cleavage and nuclear translocation of SREBP1c, further repressing the expression of SREBP1c-mediated target genes, and reducing lipogenesis and lipid accumulation. 37…”