title: Deep venomics of the Pseudonaja genus reveals inter- and intra-specific variation.
abstract: Australian elapid venom remains an under-investigated resource of novel bioactive peptides. In this study, the venom gland transcriptomes and proteomes of the Australian western brown snakes, Pseudonaja aspidorhyncha and Pseudonaja nuchalis, were compared to Pseudonaja textilis. A deep venomics strategy incorporating high throughput 454 pyrosequencing gave a total of 200,911 raw reads for the three venoms. Subsequent annotation identified 5716 transcripts from 20 different toxin families with inter-specific variation between species observed in eight of the less abundant families. Integration of each venom proteome with the corresponding annotated reads identified 65 isoforms from six toxin families; high sequence coverage highlighted subtle differences between sequences and intra and inter-specific variation between species. High quality MS/MS data identified unusual glycoforms with natriuretic peptides from P. aspidorhyncha and P. nuchaliscontaining O-linked trisaccharides with high homology to the glycosylated region of TNPc. Molecular evolutionary assessments indicated the accelerated evolution of all toxin families with the exception of both natriuretic peptides and P. aspidorhyncha PLA2s that were found to be evolutionarily constrained under purifying selection pressures. This study has revealed a wide range of novel peptide sequences from six bioactive peptide families and highlights the subtle differences between toxins in these closely related species.
  
  
  CONCLUSIONS
  Mining Australia's vastly untapped source of toxins from its venomous creatures has been significantly advanced by employing deep venomics methodology. Technological advances in transcriptome analysis using next generation sequencing platforms and proteome analysis by highly sensitive tandem mass spectrometry allowed a more comprehensive interrogation of three underinvestigated brown snake (Pseudonaja) venoms uncovering many novel peptide sequences that are unique to these closely related species. This generic strategy will provide invaluable information when applied to other venomous snakes for a deeper understanding of venom composition, envenomation, venom evolution, as well as identifying research tools and drug leads.
authors.affiliation: Institute for Molecular Bioscience, The University of Queensland, QLD 4072, Australia.
authors.forename: Timothy
authors.initials: T
authors.lastname: Reeks
authors.affiliation: Institute for Molecular Bioscience, The University of Queensland, QLD 4072, Australia.
authors.forename: Vincent
authors.initials: V
authors.lastname: Lavergne
authors.affiliation: Department of Ecology, Evolution and Behavior The Alexander Silberman Institute for Life Sciences, Hebrew University of Jerusalem, Israel.
authors.forename: Kartik
authors.initials: K
authors.lastname: Sunagar
authors.affiliation: Institute for Molecular Bioscience, The University of Queensland, QLD 4072, Australia.
authors.forename: Alun
authors.initials: A
authors.lastname: Jones
authors.affiliation: Institute for Molecular Bioscience, The University of Queensland, QLD 4072, Australia.
authors.forename: Eivind
authors.initials: E
authors.lastname: Undheim
authors.affiliation: Venom Supplies Pty Ltd., Australia.
authors.forename: Nathan
authors.initials: N
authors.lastname: Dunstan
authors.affiliation: Institute for Molecular Bioscience, The University of Queensland, QLD 4072, Australia; Venom Evolution Lab, School of Biological Sciences, The University of Queensland, QLD 4072, Australia.
authors.forename: Bryan
authors.initials: B
authors.lastname: Fry
authors.affiliation: Institute for Molecular Bioscience, The University of Queensland, QLD 4072, Australia. Electronic address: p.alewood@imb.uq.edu.au.
authors.forename: Paul F
authors.initials: PF
authors.lastname: Alewood
mesh_terms: D000818:Animals; D004546:Elapid Venoms / Q000235:genetics / Q000378:metabolism; D017815:Elapidae / Q000235:genetics / Q000378:metabolism; D013045:Species Specificity
chemical_list: D004546:Elapid Venoms
keywords: Glycopeptides; Proteome; Pseudonaja genus; Snake toxins; Venom gland transcriptome; Venomics