Abstract
The malaria mosquito, Anopheles stephensi, and other mosquitoes modulate their biology to match the time-of-day. In the present work, we used a non-hypothesis driven approach (untargeted proteomics) to identify proteins in mosquito tissue, and then quantified the relative abundance of the identified proteins from An. stephensi bodies. Using these quantified protein levels, we then analyzed the data for proteins that were only detectable at certain times-of-the day, highlighting the need to consider time-of-day in experimental design. Further, we extended our time-of-day analysis to look for proteins which cycle in a rhythmic 24-hour (“circadian”) manner, identifying 31 rhythmic proteins. Finally, to maximize the utility of our data, we performed a proteogenomic analysis to improve the genome annotation of An. stephensi. We compare peptides that were detected using mass spectrometry but are ‘missing’ from the An. stephensi predicted proteome, to reference proteomes from 38 other primarily human disease vector species. We found 239 such peptide matches and reveal that genome annotation can be improved using proteogenomic analysis from taxonomically diverse reference proteomes. Examination of ‘missing’ peptides revealed reading frame errors, errors in gene-calling, overlapping gene models, and suspected gaps in the genome assembly.
Highlights
Anopheles stephensi is a major malaria vector in southern Asia where its geographic range extends across the Indian subcontinent [1]
Research on the African Anopheles gambiae mosquito has demonstrated that the behavior and physiology of the mosquito is highly dependent on circadian biology and time-of-day
We reveal that genome annotation can be improved using proteogenomic analysis from taxonomically diverse reference proteomes
Summary
Anopheles stephensi is a major malaria vector in southern Asia where its geographic range extends across the Indian subcontinent [1]. Research on the African Anopheles gambiae mosquito has demonstrated that the behavior and physiology of the mosquito is highly dependent on circadian biology and time-of-day. Wellcome Trust (No 095831) for the Centre for Immunity, Infection and Evolution. The LC-MS QExactive equipment was purchased by a Wellcome Trust Institutional Strategic Support Fund and a strategic award from the Wellcome Trust for the Centre for Immunity, Infection and Evolution (095831/Z/11/Z). Rapid Novor provided support in the form of a salary for author TLB, but did not have any additional role in the study design, data collection and analysis, decision to publish, or preparation of the manuscript. The remaining funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript
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