Abstract
Aedes aegypti mosquitoes do not have a typical functional urea cycle for ammonia disposal such as the one present in most terrestrial vertebrates. However, they can synthesize urea by two different pathways, argininolysis and uricolysis. We investigated how formation of urea by these two pathways is regulated in females of A. aegypti. The expression of arginase (AR) and urate oxidase (UO), either separately or simultaneously (ARUO) was silenced by RNAi. The amounts of several nitrogen compounds were quantified in excreta using mass spectrometry. Injection of mosquitoes with either dsRNA-AR or dsRNA-UO significantly decreased the expressions of AR or UO in the fat body (FB) and Malpighian tubules (MT). Surprisingly, the expression level of AR was increased when UO was silenced and vice versa, suggesting a cross-talk regulation between pathways. In agreement with these data, the amount of urea measured 48 h after blood feeding remained unchanged in those mosquitoes injected with dsRNA-AR or dsRNA-UO. However, allantoin significantly increased in the excreta of dsRNA-AR-injected females. The knockdown of ARUO mainly led to a decrease in urea and allantoin excretion, and an increase in arginine excretion. In addition, dsRNA-AR-injected mosquitoes treated with a specific nitric oxide synthase inhibitor showed an increase of UO expression in FB and MT and a significant increase in the excretion of nitrogen compounds. Interestingly, both a temporary delay in the digestion of a blood meal and a significant reduction in the expression of several genes involved in ammonia metabolism were observed in dsRNA-AR, UO or ARUO-injected females. These results reveal that urea synthesis and excretion in A. aegypti are tightly regulated by a unique cross-talk signaling mechanism. This process allows blood-fed mosquitoes to regulate the synthesis and/or excretion of nitrogen waste products, and avoid toxic effects that could result from a lethal concentration of ammonia in their tissues.
Highlights
Mosquitoes constitute a severe scourge for the world population since they are vectors of etiological agents that cause more than one million deaths annually [1,2,3,4,5]
To assess the efficiency of Double-stranded RNA (dsRNA)-mediated knockdown, arginase and urate oxidase expression was first evaluated by qRT-PCR in the fat body (FB) and Malpighian tubules (MT) of individual dsRNA-injected mosquitoes at 24 (Fig. 1 A–D) and 48 h after blood feeding (Fig. 1 E–H)
A. aegypti females were first injected with dsRNA-firefly luciferase control, dsRNA-arginase, dsRNA-urate oxidase or both dsRNA-AR and dsRNA-UO, and fed a blood meal
Summary
Mosquitoes constitute a severe scourge for the world population since they are vectors of etiological agents that cause more than one million deaths annually [1,2,3,4,5]. The populations of Aedes aegypti, the main vector of dengue and yellow fever viruses, as well as the number of cases of people infected with viruses transmitted by these mosquitoes have increased dramatically [6,7]. How the female mosquitoes can overcome this metabolic challenge during blood meal digestion is poorly understood. Metabolic labeling studies using 14C-proteins revealed that in A. aegypti most of the amino acids generated after the digestion of a blood meal (,70%) are oxidized for immediate energy needs and excreted as CO2 or waste; 20% is retained in the female as a mixture of protein (,10%), lipid (,8%), and sugar (,2%); whereas only 10% is allocated for oogenesis (,4% for protein and ,6% for lipid) [8]
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have