Abstract
Conotoxins are peptide neurotoxins produced by predatory cone snails. They are mostly cysteine-rich short peptides with remarkable structural diversity. The conserved signal peptide sequences of their mRNA-encoded precursors have enabled the grouping of known conotoxins into a limited number of superfamilies. However, the conotoxins within each superfamily often present variable sequences, cysteine frameworks, and post-translational modifications. To understand better how conotoxins are diversified, we performed a venomic study with C. flavidus, an uninvestigated vermivorous Conus species, by combining transcriptomic and proteomic analyses. In order to obtain the full-length conotoxin sequences, protease digestion was not performed with the venom extraction prior to spectra acquisition via tandem mass spectrometry (MS/MS). Because conotoxins are produced from mRNA-encoded precursors by means of proteolytic cleavage, nonspecific digestion of precursors was applied during the database search. Special attention was also paid in interpreting the MS/MS spectra. All together, these analyses identified 69 nonredundant cDNA sequences and 31 conotoxin components with confident MS/MS spectra. A new Q-superfamily was also identified. More importantly, this study revealed that conotoxin-encoding transcripts are diversified by hypermutation, fragment insertion/deletion, and mutation-induced premature termination, and that a single mRNA species can produce multiple toxin products through alternative post-translational modifications and alternative cleavages of the translated precursor. These multiple diversification strategies at different levels may explain, at least in part, the diversity of conotoxins, and provide the basis for further investigation.
Highlights
Cone snails (Conus species) are predatory mollusks that inhabit tropical and subtropical shallow seawater
We studied the venom from Conus flavidus, a previously uninvestigated vermivorous Conus snail collected in the South China Sea, using a combined transcriptomic and proteomic approach
414 sequences were cloned with cis mRNA sequences marked with poly-A tags, and five sequences were cloned with trans mRNA sequences marked with poly-T tags
Summary
Cone snails (Conus species) are predatory mollusks that inhabit tropical and subtropical shallow seawater Their venom ducts produce a mixture of peptides, generally known as conotoxins, that have exquisite specificity for different ion channels, receptors, and transporters. Because fewer than 2000 conotoxins have been conclusively identified in 92 different Conus species [7] and 99.8% of conotoxins are yet to be discovered, the mechanisms of conotoxin diversity are far from clear Despite this conundrum, it has been established that conotoxin peptides are produced from corresponding mRNA-encoded precursors that typically contain a highly conserved N-terminal signal peptide, a more variable propeptide, and a hypervariable mature peptide [8, 9]. Conotoxins are considered to Molecular & Cellular Proteomics 13.1
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