Neuropetides are a diverse class of molecules, with a wide range of physiological effects, making them an interesting target to study. They are produced as inactive prepropeptides that can contain one or more copies of the active peptide, as well as signal sequences, cleavage sites, and spacer regions. Due to this unique composition and low in vivo concentration, they present a challenge to researchers wanting to examine these molecules using LC-MS/MS. One strategy, which was recently used by Realis-Doyelle and colleagues, is to focus on a specific tissue, visceral ganglia in this case, and to use advanced MS methods that maximise the identification of low abundance peptides. This study incorporates trapped ion mobility separation to time-of-flight (tims-TOF) mass analysis with parallel accumulation serial fragmentation (PASEF) technology. These technologies are powerful methods that produce complex datasets that PEAKS platforms can take advantage of.
How was PEAKS used?
The database search workflow was used to identify peptides isolated from visceral ganglia over an annual reproductive cycle. The authors used a custom database of known neuropeptide precursors from the Oyster species used in the study. Settings available in the project setup allow for the specification of timsTOF as the instrument type, making sure the data is correctly handled. For more information on the use of PEAKS platforms with timsTOF data please see the webinars available on our website: timsTOF Data Analysis With PEAKS Xpro.
Réalis-Doyelle, Emilie, et al. “Transcriptome Profiling of the Pacific Oyster Crassostrea Gigas Visceral Ganglia over a Reproduction Cycle Identifies Novel Regulatory Peptides.” Marine Drugs, no. 8, MDPI AG, Aug. 2021, p. 452. Crossref, doi:10.3390/md19080452.
The neuropeptides involved in the regulation of reproduction in the Pacific oyster (Crassostrea gigas) are quite diverse. To investigate this diversity, a transcriptomic survey of the visceral ganglia (VG) was carried out over an annual reproductive cycle. RNA-seq data from 26 samples corresponding to VG at different stages of reproduction were de novo assembled to generate a specific reference transcriptome of the oyster nervous system and used to identify differentially expressed transcripts. Transcriptome mining led to the identification of novel neuropeptide precursors (NPPs) related to the bilaterian Eclosion Hormone (EH), crustacean female sex hormone/Interleukin 17, Nesfatin, neuroparsin/IGFBP, prokineticins, and urotensin I; to the protostome GNQQN, pleurin, prohormones 3 and 4, prothoracotropic hormones (PTTH), and QSamide/PXXXamide; to the lophotrochozoan CCWamide, CLCCY, HFAamide, and LXRX; and to the mollusk-specific NPPs CCCGS, clionin, FYFY, GNamide, GRWRN, GSWN, GWE, IWMPxxGYxx, LXRYamide, RTLFamide, SLRFamide, and WGAGamide. Among the complete repertoire of NPPs, no sex-biased expression was observed. However, 25 NPPs displayed reproduction stage-specific expression, supporting their involvement in the control of gametogenesis or associated metabolisms.