Transcriptome and venom proteome of the box jellyfish Chironex fleckeri

Brinkman et al. BMC Genomics (2015) 16:407 DOI 10.1186/s12864-015-1568-3 RESEARCH ARTICLE Open Access Transcriptome and venom proteome of the box jellyfish Chironex fleckeri Diane L Brinkman1*, Xinying Jia2, Jeremy Potriquet2, Dhirendra Kumar2,3, Debasis Dash3, David Kvaskoff4 and Jason Mulvenna2,5* Abstract Background: The box jellyfish, Chironex fleckeri, is the largest and most dangerous cubozoan jellyfish to humans. It produces potent and rapid-acting venom and its sting causes severe localized and systemic effects that are potentially life-threatening. In this study, a combined transcriptomic and proteomic approach was used to identify C. fleckeri proteins that elicit toxic effects in envenoming. Results: More than 40,000,000 Illumina reads were used to de novo assemble ∼ 34,000 contiguous cDNA sequences and ∼ 20,000 proteins were predicted based on homology searches, protein motifs, gene ontology and biological pathway mapping. More than 170 potential toxin proteins were identified from the transcriptome on the basis of homology to known toxins in publicly available sequence databases. MS/MS analysis of C. fleckeri venom identified over 250 proteins, including a subset of the toxins predicted from analysis of the transcriptome. Potential toxins identified using MS/MS included metalloproteinases, an alpha-macroglobulin domain containing protein, two CRISP proteins and a turripeptide-like protease inhibitor. Nine novel examples of a taxonomically restricted family of potent cnidarian pore-forming toxins were also identified. Members of this toxin family are potently haemolytic and cause pain, inflammation, dermonecrosis, cardiovascular collapse and death in experimental animals, suggesting that these toxins are responsible for many of the symptoms of C. fleckeri envenomation. Conclusions: This study provides the first overview of a box jellyfish transcriptome which, coupled with venom proteomics data, enhances our current understanding of box jellyfish venom composition and the molecular structure and function of cnidarian toxins. The generated data represent a useful resource to guide future comparative studies, novel protein/peptide discovery and the development of more effective treatments for jellyfish stings in humans. (Length: 300). Keywords: Chironex fleckeri, Venom, Transcriptome, Proteome Background Box jellyfish (Class Cubozoa) produce venoms that are designed to swiftly incapacitate prey and deter predators, but they also cause adverse effects in envenomed humans. Cubozoan venoms are stored within complex intracellular structures (nematocysts) that are housed within specialized cells (nematocytes) located mainly in the tentacles of the jellyfish. When triggered to discharge, each nematocyst explosively releases a harpoon-like tubule * Correspondence: ; jason.mulvenna@qimrberghofer. edu.au 1 Australian Institute of Marine Science, Townsville, QLD, Australia 2 Infectious Diseases Program, QIMR Berghofer Medical Research Institute, Brisbane, QLD, Australia Full list of author information is available at the end of the article that injects a toxic cocktail of venom components into the victim or prey. Chironex fleckeri is the largest and most venomous box jellyfish species. It inhabits the tropical coastal waters of Australia and is renowned for its ability to inflict extremely painful and potentially life threatening stings to humans. Symptoms of C. fleckeri envenoming can include the rapid onset of severe cutaneous pain and inflammation, dermonecrosis, dyspnoea, transient hypertension, hypotension, cardiovascular collapse and cardiac arrest (reviewed in [1]). Due to its clinical importance, C. fleckeri has remained one of the most intensively researched box jellyfish species. Over five decades of research on whole or fractionated C. fleckeri tentacle extracts and nematocystderived venom has established that C. fleckeri toxins elicit © 2015 Brinkman et al.; licensee BioMed Central. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly credited. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated. Brinkman et al. BMC Genomics (2015) 16:407 Page 2 of 15 a diverse range of bioactivities including nociception, in vitro cytotoxicity in cultured myocytes (cardiac, skeletal and smooth muscle) and hepatocytes, haemolytic activity and pore formation in mammalian cell membranes, neurotoxicity and myotoxicity in nerve and muscle preparations, and in vivo dermonecrotic, cardiovascular and lethal effects in a variety of experimental animals [1-5]. In recent studies, the potent in vitro haemolytic and in vivo cardiovascular activities of C. fleckeri venom have been attributed primarily to the action of a subset of C. fleckeri toxins (CfTXs) that are members of a taxonomically restricted family of cnidarian pore-forming toxins [2,5]. A single proteomics study of C. fleckeri venom revealed that several isoforms of the CfTXs are highly abundant in the venom proteome [6], but due to the lack of genomic and transcriptomic data for cubozoans, few other potential toxins were identified [6]. However, the diversity of biological activities associated with C. fleckeri venom and the complexity of its venom composition, suggest that other biologically important venom components are yet to be identified. These novel cubozoan venoms could represent a source of potentially useful bioactive compounds for the development of novel therapeutics. Advances in computational techniques for the assembly and annotation of sequence data have enabled the rapid characterization of biologically important protein mixtures from a range of organisms [7,8]. In this work we utilized Illumina sequencing in concert with tandem mass spectroscopy (MS/MS) to conduct a large-scale exploration of the transcriptome and venom proteome of C. fleckeri. The newly obtained transcriptomic data facilitated the detection of several new CfTX isoforms and other putative toxin families, including metalloproteinases, that have not been previously identified in cubozoan venoms. This study not only provides extensive information on the molecular diversity of toxins in C. fleckeri venom, but also provides the first overview of a box jellyfish transcriptome; thus representing a valuable resource for future comparative genomic, transcriptomic and proteomic studies or novel protein/peptide discovery. Table 1 Summary of assembly and annotation of nucleotide sequence data from Chironex fleckeri tentacle tissue Results using blastx — SwissProt, Cnidaria protein sequences from the GenBank non-redundant protein database and predicted protein sets from the Hydra magnipapillata and Nemato (...truncated)