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Proteomics analysis of muscles from long lived O1/O3 flies in young and old age (Circadian gene variants and the skeletal muscle circadian clock contribute to the evolutionary divergence in longevity across Drosophila populations) Liam C. Hunt, Jianqin Jiao, Yong-Dong Wang, David Finkelstein, Deepti Rao, Michelle Curley, Maricela Robles-Murguia, Abbas Shirinifard, Vishwajeeth R. Pagala, Junmin Peng, Yiping Fan, and Fabio Demontis. Genome Research In press 2019.

ABSTRACT: Protein expression profiles in the whole tissue lysate extracted from the thorax of O1 and O3 long-lived flies with the control B3 strain flies and analyzed by tandem mass tag (TMT)–based mass spectrometry.

INSTRUMENT(S): Orbitrap Fusion

ORGANISM(S): Drosophila Melanogaster (fruit Fly)

TISSUE(S): Aorta Thoracica Smooth Muscle

SUBMITTER: Vishwajeeth Pagala

LAB HEAD: Fabio Demontis

PROVIDER: PXD014223 | Pride | 2019-06-13

REPOSITORIES: Pride

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Circadian gene variants and the skeletal muscle circadian clock contribute to the evolutionary divergence in longevity across <i>Drosophila</i> populations.

Hunt Liam C LC Jiao Jianqin J Wang Yong-Dong YD Finkelstein David D Rao Deepti D Curley Michelle M Robles-Murguia Maricela M Shirinifard Abbas A Pagala Vishwajeeth R VR Peng Junmin J Fan Yiping Y Demontis Fabio F

Genome research 20190627 8

Organisms use endogenous clocks to adapt to the rhythmicity of the environment and to synchronize social activities. Although the circadian cycle is implicated in aging, it is unknown whether natural variation in its function contributes to differences in lifespan between populations and whether the circadian clock of specific tissues is key for longevity. We have sequenced the genomes of <i>Drosophila melanogaster</i> strains with exceptional longevity that were obtained via multiple rounds of ...[more]

PMID: 31249065

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Project description:Tsetse flies (Glossina spp.) are major vectors of African trypanosomes, causing either Human or Animal African Trypanosomiasis (HAT or AAT). Several approaches are developed to control the disease among which the anti-vector Sterile Insect Technique. Another approach in the frame of anti-vector strategies could consist in controlling the fly’s vector competence which needs identifying factors (genes, proteins, biological pathways, …) involved in this process. The present work aims to verify whether protein candidates identified under experimental controlled conditions on insectary-reared tsetse flies have their counterpart in field-collected flies. Glossina palpalis palpalis flies naturally infected with Trypanosoma congolense were sampled in two HAT/AAT foci in Southern Cameroon. After dissection, the proteome from guts of parasite-infected flies were compared to that from uninfected flies in order to identify quantitative and/or qualitative changes associated to infection. A total of 3291 proteins were identified of which 1818 could be quantified. The comparative analysis allowed identifying 175 proteins with significant decreased abundance in infected as compared to uninfected flies, while 61 proteins displayed increased abundance. Among the former are RNA binding proteins, kinases, actin, ribosomal proteins, endocytosis proteins, oxido-reductases, as well as proteins that are unusually found such as tsetse salivary proteins (Tsal) or Yolk proteins. Among the proteins with increased abundance are fructose-1,6-biphosphatase, serine proteases, membrane trafficking proteins, death proteins (or apoptosis proteins), and SERPINs (inhibitor of serine proteases, enzymes considered as trypanosome virulence factors) that displayed highest increased abundance. Sodalis, Wiggleswothia and Wolbachia proteins are strongly under-represented, particularly when compared to data from similar experimentation conducted under controlled conditions on T. brucei gambiense infected (or uninfected) G. palpalis gambiensis insectary reared flies. Comparing the overall recorded data, 364 proteins identified in gut extracts from field flies were shown to have a homologue in insectary flies. Discrepancies between the two studies may arise from differences in the species of studied flies and trypanosomes as well as in differences in environmental conditions in which the two experiments were carried out. Finally, the present study together with former proteomic and transcriptomic studies on the secretome of trypanosomes, on the gut extracts from insectary reared and on field collected tsetse flies, provide a pool of data and information on which to draw in order to perform further investigations on, for example, mammal host immunization or on fly vector competence modification via para-transgenic approaches.

Project description:To test whether other genes were being silenced in the Cyp6g1 knockdown strain due to off-target RNAi effects, and whether other gene expression changes were contributing to the altered susceptibility to imidacloprid in these knockdown flies. A comparison between w;Act5C-GAL4/CyO; UAS:RNAi_Cyp6g1Hp2/TM3Sb and the genetic control w;Act-GAL4/CyO;+/TM3Sb was performed. Ten 2-3 day old adult males or females were transferred to sugar-agar plates and then collected at various time points (0, 2, 5, 8 hours). The RNA samples for up to three independent experiments per timepoint for each genotype were then pooled, in equal concentrations, before hybridisation to the Affymetrix Drosophila GeneChip 1. As a preliminary analysis provided evidence for assuming that there is no interaction between genotype and duration in the sugar-agar plate, the knockdown and control chips for each of the five timepoints provided some degree of replication for the comparison between the knockdown flies and control flies. Differential expression that was consistently observed between knockdown and control over each of the five timepoints was regarded as providing evidence of a genuine difference. 2-3 day old adult flies of the appropriate genotype (Cyp6g1 knockdown or its genetic control) were transferred to bioassay plates and collected after specified timepoints in the assay plate, i.e. 0, 2, 5 or 8 hours. Each experiment contained ?3 replicates of each genotype/timepoint, and a different number of independent experiments (1-3) were pooled per microarray sample as described below. An experiment is defined as treatments set up from the same cohort of flies on a single day. Independent experiments are set up using distinct cohorts of flies, on distinct days. We included one pair of tests to compare samples from single experiments with pooled samples from independent experiments (DS003001 to DS003004, DS003002 to DS003005), and also a single pair of samples to compare males to females (DS003002 to DS003003). As it turned out - in our analysis - the timepoints selected were not discriminative, in that there were no differences detected that could be attributed to an effect of time in the bioassay.

			Action	DRS
	1WkTMT_F1.mzXML	Mzxml
	1WkTMT_F1.pepXML	Pepxml
	1WkTMT_F1.raw	Raw
	1WkTMT_F10.mzXML	Mzxml
	1WkTMT_F10.pepXML	Pepxml

Dataset Information

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Publications

Circadian gene variants and the skeletal muscle circadian clock contribute to the evolutionary divergence in longevity across <i>Drosophila</i> populations.

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