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Mass spectrometry based proteomic analysis of salivary glands of urban malaria vector Anopheles stephensi.

ABSTRACT: Salivary gland proteins of Anopheles mosquitoes offer attractive targets to understand interactions with sporozoites, blood feeding behavior, homeostasis, and immunological evaluation of malaria vectors and parasite interactions. To date limited studies have been carried out to elucidate salivary proteins of An. stephensi salivary glands. The aim of the present study was to provide detailed analytical attributives of functional salivary gland proteins of urban malaria vector An. stephensi. A proteomic approach combining one-dimensional electrophoresis (1DE), ion trap liquid chromatography mass spectrometry (LC/MS/MS), and computational bioinformatic analysis was adopted to provide the first direct insight into identification and functional characterization of known salivary proteins and novel salivary proteins of An. stephensi. Computational studies by online servers, namely, MASCOT and OMSSA algorithms, identified a total of 36 known salivary proteins and 123 novel proteins analysed by LC/MS/MS. This first report describes a baseline proteomic catalogue of 159 salivary proteins belonging to various categories of signal transduction, regulation of blood coagulation cascade, and various immune and energy pathways of An. stephensi sialotranscriptome by mass spectrometry. Our results may serve as basis to provide a putative functional role of proteins in concept of blood feeding, biting behavior, and other aspects of vector-parasite host interactions for parasite development in anopheline mosquitoes.

SUBMITTER: Vijay S

PROVIDER: S-EPMC4122192 | biostudies-literature | 2014

REPOSITORIES: biostudies-literature

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Mass spectrometry based proteomic analysis of salivary glands of urban malaria vector Anopheles stephensi.

Vijay Sonam S Rawat Manmeet M Sharma Arun A

BioMed research international 20140714

Salivary gland proteins of Anopheles mosquitoes offer attractive targets to understand interactions with sporozoites, blood feeding behavior, homeostasis, and immunological evaluation of malaria vectors and parasite interactions. To date limited studies have been carried out to elucidate salivary proteins of An. stephensi salivary glands. The aim of the present study was to provide detailed analytical attributives of functional salivary gland proteins of urban malaria vector An. stephensi. A pro ...[more]

PMID: 25126571

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Project description:In order to understand the importance of functional proteins in mosquito behavior, following blood meal, a baseline proteomic dataset is essential for providing insights into the physiology of blood feeding. Therefore, in this study as first step, in solution and 1-D electrophoresis digestion approach combined with tandem mass spectrometry (nano LC-MS/MS) and computational bioinformatics for data mining was used to prepare a baseline proteomic catalogue of salivary gland proteins of sugar fed An. culicifacies mosquitoes. A total of 106 proteins were identified and analyzed by SEQUEST algorithm against mosquito protein database from Uniprot/NCBI. Importantly, D7r1, D7r2, D7r4, salivary apyrase, anti-platelet protein, calreticulin, antigen 5 family proteins were identified and grouped on the basis of biological and functional roles. Secondly, differential protein expression and annotations between salivary glands of sugar fed vs blood fed mosquitoes was analyzed using 2-Delectrophoresis combined with MALDI-TOF mass spectrometry. The alterations in the differential expression of total 38 proteins was observed out of which 29 proteins like beclin-1, phosphorylating proteins, heme oxygenase 1, ferritin, apoptotic proteins, coagulation and immunity like, serine proteases, serpins, c-type lectin and protein in regulation of blood feeding behavior were found to be up regulated while 9 proteins related to blood feeding, juvenile hormone epoxide hydrolase ii, odorant binding proteins and energy metabolic enzymes were found to be down regulated. To our knowledge, this study provides a first time baseline proteomic dataset and functional annotations of An. culicifacies salivary gland proteins that may be involved during the blood feeding. Identification of differential salivary proteins between sugar fed and blood fed mosquitoes and their plausible role may provide insights into the physiological processes associated with feeding behavior and sporozoite transmission during the process of blood feeding.

Project description:Small laboratory cage trials of non-drive and gene-drive strains of the Asian malaria vector mosquito, Anopheles stephensi, were used to investigate release ratios and other strain properties for their impact on transgene spread during simulated population modification. We evaluated the effects of transgenes on survival, male contributions to next-generation populations, female reproductive success and the impact of accumulation of gene drive-resistant genomic target sites resulting from nonhomologous end-joining (NHEJ) mutagenesis during Cas9, guide RNA-mediated cleavage. Experiments with a non-drive, autosomally-linked malaria-resistance gene cassette showed 'full introduction' (100% of the insects have at least one copy of the transgene) within 8 weeks (? 3 generations) following weekly releases of 10:1 transgenic:wild-type males in an overlapping generation trial design. Male release ratios of 1:1 resulted in cages where mosquitoes with at least one copy of the transgene fluctuated around 50%. In comparison, two of three cages in which the malaria-resistance genes were linked to a gene-drive system in an overlapping generation, single 1:1 release reached full introduction in 6-8 generations with a third cage at ~80% within the same time. Release ratios of 0.1:1 failed to establish the transgenes. A non-overlapping generation, single-release trial of the same gene-drive strain resulted in two of three cages reaching 100% introduction within 6-12 generations following a 1:1 transgenic:wild-type male release. Two of three cages with 0.33:1 transgenic:wild-type male single releases achieved full introduction in 13-16 generations. All populations exhibiting full introduction went extinct within three generations due to a significant load on females having disruptions of both copies of the target gene, kynurenine hydroxylase. While repeated releases of high-ratio (10:1) non-drive constructs could achieve full introduction, results from the 1:1 release ratios across all experimental designs favor the use of gene drive, both for efficiency and anticipated cost of the control programs.

Dataset Information

Mass spectrometry based proteomic analysis of salivary glands of urban malaria vector Anopheles stephensi.

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Mass spectrometry based proteomic analysis of salivary glands of urban malaria vector Anopheles stephensi.

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