Mass spectrometry-based stable-isotope tracing uncovers metabolic alterations in pyruvate kinase-deficient Aedes aegypti mosquitoes.
Mass spectrometry-based stable-isotope tracing uncovers metabolic alterations in pyruvate kinase-deficient Aedes aegypti mosquitoes.
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DOI：
10.1016/j.ibmb.2020.103366
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发表时间：
2020-04
影响因子：
3.8
通讯作者：
N. Petchampai;J. Isoe;Thomas D. Horvath;S. Dagan;L. Tan;P. Lorenzi;D. Hawke;P. Y. Scaraffia
中科院分区：
文献类型：
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作者：
N. Petchampai;J. Isoe;Thomas D. Horvath;S. Dagan;L. Tan;P. Lorenzi;D. Hawke;P. Y. Scaraffia
最近对埃及伊蚊重组丙酮酸激酶(PK)的体外鉴定表明，该酶受多种变构效应物的独特调节。在这里，我们进一步研究了在不同营养条件下保持的蚊子关键代谢组织中PK基因和蛋白的表达以及酶活性。我们还使用了几种技术，包括RNA干扰和基于稳定同位素示踪的质谱仪，研究了PK耗竭对代谢的影响。转录分析显示，喂食后PK mRNA在蚊子组织内和跨组织中的动态表达模式，而相应的蛋白质水平在分析的整个时间过程中保持稳定。然而，在吸食蔗糖、吸血和饥饿的蚊子的脂肪体中，PK活性显著不同。PK的基因沉默并没有改变靠蔗糖维持血液喂养的雌性动物的存活率。然而，观察到在不同营养方案下维持的PK缺乏的雌性动物的存活率提高。我们的结果表明，蚊子通过上调编码NADP-苹果酸酶-1、磷酸烯醇式丙酮酸羧激酶-1、磷酸甘油脱氢酶和谷氨酸脱氢酶基因的表达，以及通过减少葡萄糖氧化和与氨解毒相关的代谢途径，克服了PK缺乏。综上所述，我们的数据表明，PK赋予了ToA。埃及式代谢可塑性，严密地调节碳和氮的代谢。
A recentin vitrocharacterization of a recombinant pyruvate kinase (PK) fromAedes aegyptimosquitoes 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 toA. aegyptia metabolic plasticity to tightly regulate both carbon and nitrogen metabolism.