SynthSys-Synthetic and Systems Biology, School of Biological Sciences, University of Edinburgh, Edinburgh, United Kingdom.
Centre for Immunity, Infection and Evolution, University of Edinburgh, Edinburgh, United Kingdom.
PLoS One. 2019 Jul 29;14(7):e0220225. doi: 10.1371/journal.pone.0220225. eCollection 2019.
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.
按蚊和其他蚊子会根据时间来调节自身的生物学特性。在本研究中,我们采用了一种非假设驱动的方法(非靶向蛋白质组学)来鉴定蚊子组织中的蛋白质,并对斯蒂芬斯按蚊体内鉴定出的蛋白质的相对丰度进行定量分析。利用这些定量的蛋白质水平,我们分析了仅在特定时间出现的蛋白质数据,这突出表明在实验设计中需要考虑时间因素。此外,我们将时间分析扩展到寻找以 24 小时为周期(“昼夜节律”)循环的蛋白质,共鉴定出 31 种节律蛋白。最后,为了最大限度地利用我们的数据,我们进行了蛋白质基因组学分析以改进斯蒂芬斯按蚊的基因组注释。我们将通过质谱检测到但在斯蒂芬斯按蚊预测的蛋白质组中“缺失”的肽与 38 种主要的人类疾病载体物种的参考蛋白质组进行比较。我们发现了 239 个这样的肽匹配,并表明使用来自分类上多样化的参考蛋白质组的蛋白质基因组学分析可以改进基因组注释。对“缺失”肽的检查揭示了读码框错误、基因调用错误、重叠的基因模型以及基因组组装中疑似存在的空白。