Revisiting toxins with transcriptomics-informed proteomics of venom glands and crude venom from Centruroides bicolor...

The sting of the scorpion Centruroides bicolor causes a large morbidity in Panama. To characterize its venom, transcriptomic and proteomic analyses of the venom glands and the crude venom were performed. These two approaches utilized high-throughput sequencing to enhance the likelihood of detecting a wide range of venom proteins correlated with the venom proteome. After RNA venom gland extraction, a cDNA library was constructed and sequenced by RNA-seq. Also, the crude venom was digested using trypsin and chymotrypsin, and the resulting peptides were analyzed using a nano-LC-MS/MS. Notably, transcriptomic and proteomic venom approaches identified a hyaluronidase, alpha- and beta-neurotoxins that affect Na⁺ channels, CRISP proteins, metalloproteinases, transferrin, monooxygenase alpha-peptidyl-glycine, serine proteases, alpha pancreatic amylase, lysozyme, neurotoxins targeting K⁺ channels, neprilysin, scorpine, angiotensin-converting enzyme, insulin-like growth factor-binding domain proteins, nucleobindin-like proteins, and uncharacterized proteins. Interestingly, some of the venom proteins such as nucleobindin and angiotensin-converting enzymes have been not reported in the proteome, their predicted presence has only been previously derived from the genomic sequence of Centruroides sculpturatus and C. vittatus. These newly identified components enhance the understanding of the venomous nature of C. bicolor. SIGNIFICANCE: The proteins and peptides found in Centruroides bicolor venom by transcriptomic and proteomic analyses were assessed according to the protein and toxin databases available on public domains. Notably, some of the venom proteins such as nucleobindin and angiotensin-converting enzymes have been not reported in the proteome, their predicted presence has only been previously derived from the genomic sequence of Centruroides sculpturatus and C. vittatus. Moreover, enzymatic assays, including hyaluronidase, phospholipase A2, and proteolytic activity were conducted to confirm the presence or absence of those enzymes. Interestingly, neurotoxins from C. limbatus, a related species in the region, were found in the proteome but no mRNAs were identified in the transcriptome. These newly identified components enhance the understanding of the venomous nature of Centruroides bicolor.