Abstract
BackgroundFemale Aedes aegypti mosquitoes are vectors of arboviruses that cause diverse diseases of public health significance. Blood protein digestion by midgut proteases provides anautogenous mosquitoes with the nutrients essential for oocyte maturation and egg production. Midgut-specific miR-1174 affects the functions of the midgut through its target gene serine hydroxymethyltransferase (SHMT). However, less is known about SHMT-regulated processes in blood digestion by mosquitoes.MethodsRNAi of SHMT was realized by injection of the double-stranded RNA at 16 h post-eclosion. The expression of SHMT at mRNA level and protein level was assayed by real-time PCR and Western blotting, respectively. Statistical analyses were performed with GraphPad7 using Student’s t-test.ResultsHere, we confirmed that digestion of blood was inhibited in SHMT RNAi-silenced female A. aegypti mosquitoes. Evidence is also presented that all SHMT-depleted female mosquitoes lost their flight ability and died within 48 h of a blood meal. Furthermore, most examined digestive enzymes responded differently in their transcriptional expression to RNAi depletion of SHMT, with some downregulated, some upregulated and some remaining stable. Phylogenetic analysis showed that transcriptional expression responses to SHMT silence were largely unrelated to the sequence similarity between these enzymes.ConclusionsOverall, this research shows that SHMT was expressed at a low level in the midgut of Aedes aegypti mosquitoes, but blood-meal digestion was inhibited when SHMT was silenced. Transcriptional expressions of different digestive enzymes were affected in response to SHMT depletion, suggesting that SHMT is required for the blood-meal digestion in the midgut and targeting SHMT could provide an effective strategy for vector mosquito population control.
Highlights
Female Aedes aegypti mosquitoes are vectors of arboviruses that cause diverse diseases of public health significance
We performed quantitative reverse transcriptase polymerase chain reaction (qRT-PCR) to reveal the transcriptional expression patterns of serine hydroxymethyltransferase (SHMT) in diverse tissues of female adults at different time points and found that it was highly expressed in the head, fat body, ovary and leftover tissues but expressed at very low levels in the midgut (Fig. 2a–f ); the latter is consistent with our previous evidence [22]
We examined the spatial expression of twelve digestive enzymes at 24 hours postblood-meal (h PBM) by qRT-PCR, including three trypsins (AAEL013712, AAEL013284, AAEL006425), five chymotrypsins (AAEL002347, AAEL011929, AAEL003060, AAEL001703, AAEL022646) and four serine proteases (AAEL010196, AAEL010202, AAEL007432, AAEL000028) and found that the expression of eleven enzymes was limited to the midgut, while AAEL000028 was exclusively absent from the midgut (Fig. 4a)
Summary
Female Aedes aegypti mosquitoes are vectors of arboviruses that cause diverse diseases of public health significance. A chymotrypsin-like protease gene, JHA15, is transcriptionally activated by juvenile hormone (JH) in the newly emerged female adults but its silencing resulted in no clear phenotype in blood-meal digestion [18]. It is known that the digestion of blood in the midgut of A. aegypti mosquitoes can be divided into two phases, early phase one to three hours postblood-meal (h PBM) and late phase 8–36 h PBM [19]. During the early phase of digestion, the Trypsin 3A1 Precursor AaET (AAEL007818) and female-specific chymotrypsin AaCHYMO (AAEL003060) accumulate [11, 17, 18], but Trypsin-like serine protease AaLT (AAEL013284) and Trypsin 5G1 Precursor Aa5G1 (AAEL013712) begin to be translated by 6–8 h PBM and reach maximal concentrations 24–36 h PBM [20, 21]
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