Genome-wide survey and molecular characterization of vacuolar-ATPase subunit genes in the yellow fever mosquito Aedes aegypti (Diptera: Culicidae)

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Show simple item record Coskun, Basak 2017-06-21T16:51:01Z 2017-06-21T16:51:01Z 2017-08-01 en_US
dc.description.abstract The yellow fever mosquito, Aedes aegypti, is a significant vector of several viral diseases, including Zika, dengue fever, yellow fever, and chikungunya. Since vaccines are not currently available for these viruses, control of the disease vectors by using insecticides is the most common practice for preventing disease. As a result, Ae. aegypti has developed resistance against many of the most commonly used insecticides, including organophosphates and pyrethroids. The rise in resistance in vector mosquitoes requires the search for new control strategies, such as RNA interference (RNAi), to manage mosquito populations. Vacuolar H[sup plus]+-ATPase (V-ATPase), a multi-subunit enzyme involved in many cellular processes, including membrane energization, acidification of organelles, and entry of dengue virus into the cytoplasm, is a potential target for RNAi, though little is known about its genetic structure or expression patterns in Ae. aegypti. In this study, I performed genome-wide surveys to identify the genes encoding different subunits of the V-ATPase protein complex, partially characterized the molecular properties and expression patterns of selected V-ATPase subunit genes, and tested the feasibility of using oral-based delivery of nanoparticles formed from double-stranded RNA (dsRNA) and chitosan to suppress the expression of selected V-ATPase subunit genes in Ae. aegypti. My genome-wide surveys revealed that Ae. aegypti V-ATPase consists of 13 different subunits (A, B, C, D, E, F, G, H, a, c, c”, d, e) encoded by 14 genes. Analysis of exon-intron arrangements for each gene demonstrated that each V-ATPase subunit gene has between one (subunit c) and 12 (subunit C) exons, with most genes (11) having 3 to 6 exons. Subsequent phylogenetic analysis of the deduced amino acid sequences of each subunit showed that V-ATPase subunits A, B, C, F, G, H, and a exhibited high levels of conservation among all the examined species, but subunits D, E, c, c”, d, and e showed high conservation only among dipteran species. Analysis of the expression profiles in different tissues and developmental stages of three specific V-ATPase subunits (A, D, and H) showed that whereas the expression of these genes varied between tissues and developmental stages, the patterns of expression of subunits A, D, and H were very similar. The highest mRNA expression level was observed in Malpighian tubules in fourth-instar larvae. Interestingly, expression of subunits A, D, or H in different tissues of adults was highest in male hindgut versus Malpighian tubules in females. Feeding mosquito larvae with chitosan nanoparticles made with dsRNA complementary to subunits A, D, or H resulted in significant suppression of mRNA transcript levels of each of these subunits. Peak suppression of V-ATPase A, D, or H transcripts occurred on the fifth day, where the gene transcript level was suppressed by 66.0, 27.3, or 70.4%, respectively, as compared with those of the control. Additionally, feeding of dsRNA/chitosan nanoparticles targeting subunit D caused mortality starting on day 3, with cumulative larval mortality reaching 14.8% on the sixth day. These results suggest that oral delivery of dsRNA/chitosan nanoparticles can substantially suppress target gene expression in Ae. aegypti larvae. However, increasing RNAi efficiency in targeting V-ATPase subunit genes in mosquito larvae appears to be necessary in order to obtain higher larval mortality using oral delivery of dsRNA/chitosan nanoparticles. en_US
dc.description.sponsorship Ministry of National Education of Turkey en_US
dc.language.iso en_US en_US
dc.publisher Kansas State University en
dc.subject Aedes aegypti en_US
dc.subject Vacuolar H+-ATPase en_US
dc.subject Stage-specific expression en_US
dc.subject Tissue-specific expresion en_US
dc.subject Chitosan nanoparticles en_US
dc.subject RNA interference en_US
dc.title Genome-wide survey and molecular characterization of vacuolar-ATPase subunit genes in the yellow fever mosquito Aedes aegypti (Diptera: Culicidae) en_US
dc.type Thesis en_US Master of Science en_US
dc.description.level Masters en_US
dc.description.department Department of Entomology en_US
dc.description.advisor Kristopher S. Silver en_US
dc.description.advisor Kun Yan Zhu en_US 2017 en_US August en_US

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