Department of Tropical Medicine, Vector-Borne Infectious Disease Research Center, School of Public Health and Tropical Medicine, Tulane University, New Orleans, LA, 70112, USA.
Department of Chemistry and Biochemistry, The University of Arizona, Tucson, AZ, 85721, USA.
Insect Biochem Mol Biol. 2020 Jun;121:103366. doi: 10.1016/j.ibmb.2020.103366. Epub 2020 Apr 7.
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.
最近对埃及伊蚊重组丙酮酸激酶（PK）的体外特性分析表明，该酶受到多种别构效应物的独特调控。在这里，我们进一步研究了在不同营养条件下维持的蚊子的关键代谢组织中的 PK 基因和蛋白表达以及酶活性。我们还使用包括 RNA 干扰和基于质谱的稳定同位素示踪在内的几种技术研究了 PK 耗竭的代谢效应。转录分析显示，在蚊子组织内和组织间存在动态的 PK mRNA 表达模式，而相应的蛋白水平在整个分析过程中保持稳定。尽管如此，在蔗糖、吸血和饥饿的蚊子的脂肪体中，PK 活性差异显著。在以蔗糖为食的吸血雌性蚊子中，PK 的基因沉默并未改变其存活率。然而，在不同营养方案下维持的 PK 缺陷雌性蚊子中观察到存活率提高。我们的结果表明，蚊子通过上调编码 NADP-苹果酸酶-1、磷酸烯醇丙酮酸羧激酶-1、磷酸甘油酸脱氢酶和谷氨酸脱氢酶的基因表达，以及通过降低葡萄糖氧化和与氨解毒相关的代谢途径，克服了 PK 缺陷。总的来说，我们的数据表明，PK 赋予埃及伊蚊一种代谢灵活性，以紧密调节碳和氮代谢。
