Unknown,Transcriptomics,Genomics,Proteomics

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Bulk RNAseq of Anopheles gambiae hemocytes upon Toll pathway overactivation

ABSTRACT: Toll pathway is a key mediator of antiplasmodial immunity and its mechanism of action is dependent on hemocytes (mosquito white blood cells). Anopheles gambiae were injected with dsRNA for Cactus, an inhibitor of the Toll pathway. Cactus silencing over activates the Toll pathway. The goal of this experiment is to evaluate hemocyte transcriptional changes between control mosquitoes injected with a non-related dsRNA (dsLacZ and dsCactus injected mosquitoes. We also divide hemocytes from control and experimental conditions into 2 groups: bound (hemocytes that attach to glass - granulocyte populations) and unbound (hemocytes that remain in suspension - prohemocytes and oenocytoids). Files from Unbound fractions are labelled: UB and from bound fractions are labelled B. Some of the samples were run twice and have 2 gz files.

INSTRUMENT(S): Illumina HiSeq 4000

ORGANISM(S): Anopheles gambiae

SUBMITTER: Ana Beatriz Barletta Ferreira

PROVIDER: E-MTAB-11252 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Project description:The squid, Euprymna scolopes, and the luminescent bacterium, Vibrio fischeri, form a highly specific beneficial light organ symbiosis. Not only does the host have to select V. fischeri from the environment, but it must also prevent subsequent colonization by non-symbiotic microorganisms. Host macrophage-like hemocytes are believed to play a role in mediating the symbiosis with V. fischeri. Previous studies have shown that the colonization state of the light organ influences the host’s hemocyte response to the symbiont. To further understand the molecular mechanisms behind this process, two quantitative mass spectrometry-based proteomic techniques, isobaric tags for relative and absolute quantification (iTRAQ) and label-free spectral counting, were used to compare and quantify the adult hemocyte proteomes from colonized (sym) and uncolonized (antibiotic-treated/cured) squid. Overall, iTRAQ allowed for the quantification of 1,024 proteins with 2 or more peptides. Thirty-seven unique proteins were determined to be significantly different between sym and cured hemocytes (p-value < 0.05), with 20 more abundant proteins and 17 less abundant in sym hemocytes. The label-free approach resulted in 1,241 proteins that were identified in all replicates. Of 185 unique proteins present at significantly different amounts in sym hemocytes (as determined by spectral counting), 92 were more abundant and 93 were less abundant. Comparisons between iTRAQ and spectral counting revealed that 30 of the 37 proteins quantified by iTRAQ exhibited similar trends identified by the label-free method. Both proteomic techniques mutually identified 16 proteins that were significantly different between the two groups of hemocytes (p-value <0.05). The presence of V. fischeri in the host light organ influenced the abundance of proteins associated with the cytoskeleton, adhesion, lysosomes, proteolysis, and the innate immune response. These data provide evidence that colonization by V. fischeri alters the hemocyte proteome and reveals proteins that may be important for maintaining host-symbiont specificity.

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Bulk RNAseq of Anopheles gambiae hemocytes upon Toll pathway overactivation

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