Mass spectrometry-based stable-isotope tracing uncovers metabolic alterations in pyruvate kinase-deficient Aedes aegypti mosquitoes. (June 2020)

• Record Type: Journal Article
• Authors: Petchampai, Natthida; Isoe, Jun; Horvath, Thomas D.; Dagan, Shai; Tan, Lin; Lorenzi, Philip L.; Hawke, David H.; Scaraffia, Patricia Y.
• Is Part Of: Insect biochemistry and molecular biology. Volume 121(2020)
• Publication Date: 2020-06
• Subjects: Ammonia metabolism -- Gene regulation -- Stable-isotope labeled compounds -- Mass spectrometry -- Metabolomics; AA amino acid -- AaPK Aedes aegypti pyruvate kinase -- Aa5G1 A. aegypti serine protease -- ACN acetonitrile -- ALT1 alanine aminotransferase-1 -- dsRNA double-stranded RNA -- FA formic acid -- FL firefly luciferase -- GDH glutamate dehydrogenase -- G6PDH glucose-6-phosphate dehydrogenase -- HRAM high-resolution accurate-mass -- IC/MS ion chromatography-mass spectrometry -- LC/MS liquid chromatography-mass spectrometry -- NADP-ME1 NADP-malic enzyme-1 -- OA organic acid -- ODC ornithine decarboxylase -- PBM post blood meal -- PEPCK1 phosphoenolpyruvate carboxykinase-1 -- PPP pentose phosphate pathway -- PHGDH phosphoglycerate dehydrogenase -- PK pyruvate kinase -- PKM1, PKM2, PKR and PKL human PK isoforms -- qPCR quantitative real-time PCR -- RNAi RNA interference -- XDH1 xanthine dehydrogenase-1; Insect biochemistry -- Periodicals; Insects -- Physiology -- Periodicals; Insects -- Molecular aspects -- Periodicals; Biochemistry -- Periodicals; Insectes -- Biochimie -- Périodiques; Insectes -- Composition -- Périodiques; Insectes -- Physiologie -- Périodiques; Insectes -- Aspect moléculaire -- Périodiques; Biochimie -- Périodiques; Biochemistry; Insect biochemistry; Insects -- Molecular aspects; Insects -- Physiology; Periodicals; 572.8157
• DOI: 10.1016/j.ibmb.2020.103366
• Languages: English
• ISSNs: 0965-1748

Abstract

Abstract: A recent in vitro characterization of a recombinant pyruvate kinase (PK) from Aedes aegypti mosquitoes demonstrated that the enzyme is uniquely regulated by multiple allosteric effectors. Here, we further explored PK gene and protein expression, and enzymatic activity in key metabolic tissues of mosquitoes maintained under different nutritional conditions. We also studied the metabolic effects of PK depletion using several techniques including RNA interference and mass spectrometry-based stable-isotope tracing. Transcriptional analysis showed a dynamic post-feeding PK mRNA expression pattern within and across mosquito tissues, whereas corresponding protein levels remained stable throughout the time course analyzed. Nevertheless, PK activity significantly differed in the fat body of sucrose-, blood-fed, and starved mosquitoes. Genetic silencing of PK did not alter survival in blood-fed females maintained on sucrose. However, an enhanced survivorship was observed in PK-deficient females maintained under different nutritional regimens. Our results indicate that mosquitoes overcame PK deficiency by up-regulating the expression of genes encoding NADP-malic enzyme-1, phosphoenolpyruvate carboxykinase-1, phosphoglycerate dehydrogenase and glutamate dehydrogenase, and by decreasing glucose oxidation and metabolic pathways associated with ammonia detoxification. Taken together, our data demonstrate that PK confers to A. aegypti a metabolic plasticity to tightly regulate both carbon and nitrogen metabolism. Graphical abstract: Image 1 Highlights: Pyruvate kinase (PK) activity in fat body varies in response to nutritional changes. PK silencing enhances survival in females maintained under specific nutritional regimens. PK-deficient females increase transcript levels of specific genes. PK knockdown reduces the abundance of metabolites involved in several pathways.