To understand the diversity of scorpion venom, RNA from venomous glands from a sawfinger scorpion, Serradigitus gertschi, of the family Vaejovidae, was extracted and used for transcriptomic analysis. A total of 84,835 transcripts were assembled after Illumina sequencing. From those, 119 transcripts were annotated and found to putatively code for peptides or proteins that share sequence similarities with the previously reported venom components of other species. In accordance with sequence similarity, the transcripts were classified as potentially coding for 37 ion channel toxins; 17 host defense peptides; 28 enzymes, including phospholipases, hyaluronidases, metalloproteases, and serine proteases; nine protease inhibitor-like peptides; 10 peptides of the cysteine-rich secretory proteins, antigen 5, and pathogenesis-related 1 protein superfamily; seven La1-like peptides; and 11 sequences classified as “other venom components”. A mass fingerprint performed by mass spectrometry identified 204 components with molecular masses varying from 444.26 Da to 12,432.80 Da, plus several higher molecular weight proteins whose precise masses were not determined. The LC-MS/MS analysis of a tryptic digestion of the soluble venom resulted in the de novo determination of 16,840 peptide sequences, 24 of which matched sequences predicted from the translated transcriptome. The database presented here increases our general knowledge of the biodiversity of venom components from neglected non-buthid scorpions.

In T cells, the Kv1.3 and the KCa3.1 potassium channels regulate the membrane potential and calcium homeostasis. Notably, during TEM cell activation, the number of Kv1.3 channels on the cell membrane dramatically increases. Kv1.3 blockade results in inhibition of Ca2+ signaling in TEM cells, thus eliciting an immunomodulatory effect. Among the naturally occurring peptides, the Vm24 toxin from the Mexican scorpion Vaejovis mexicanus is the most potent and selective Kv1.3 channel blocker known, which makes it a promissory candidate for its use in the clinic. We have shown that addition of Vm24 to TCR-activated human T cells inhibits CD25 expression, cell proliferation and reduces delayed-type hypersensitivity reactions in a chronic inflammation model. Here, we used the Vm24 toxin as a tool to investigate the molecular events that follow Kv1.3 blockade specifically on human CD4+ TEM cells as they are actively involved in inflammation and are key mediators of autoimmune diseases. We combined cell viability, activation, and multiplex cytokine assays with a proteomic analysis to identify the biological processes affected by Kv1.3 blockade on healthy donors CD4+ TEM cells, following TCR activation in the presence or absence of the Vm24 toxin. The peptide completely blocked Kv1.3 channels currents without impairing TEM cell viability, and in response to TCR stimulation, it inhibited the expression of the activation markers CD25 and CD40L (but not that of CD69), as well as the secretion of the pro-inflammatory cytokines IFN-γ and TNF and the anti-inflammatory cytokines IL-4, IL-5, IL-9, IL-10, and IL-13. These results, in combination with data from the proteomic analysis, indicate that the biological processes most affected by the blockade of Kv1.3 channels in a T cell activation context were cytokine-cytokine receptor interaction, mRNA processing via spliceosome, response to unfolded proteins and intracellular vesicle transport, targeting the cell protein synthesis machinery. The Vm24 toxin, a highly specific inhibitor of Kv1.3 channels allowed us to define downstream functions of the Kv1.3 channels in human CD4+ TEM lymphocytes. Blocking Kv1.3 channels profoundly affects the mRNA synthesis machinery, the unfolded protein response and the intracellular vesicle transport, impairing the synthesis and secretion of cytokines in response to TCR engagement, underscoring the role of Kv1.3 channels in regulating TEM lymphocyte function.

Centruroides possanii is a recently discovered species of “striped scorpion” found in Mexico. Certain species of Centruroides are known to be toxic to mammals, leading to numerous cases of human intoxications in the country. Venom components are thought to possess therapeutic potential and/or biotechnological applications. Hence, obtaining and analyzing the secretory gland transcriptome and venom proteome of C. possanii is relevant, and that is what is described in this communication. Since this is a newly described species, first, its LD₅₀ to mice was determined and estimated to be 659 ng/g mouse weight. Using RNA extracted from this species and preparing their corresponding cDNA fragments, a transcriptome analysis was obtained on a Genome Analyzer (Illumina) using the 76-base pair-end sequencing protocol. Via high-throughput sequencing, 19,158,736 reads were obtained and ensembled in 835,204 sequences. Of them, 28,399 transcripts were annotated with Pfam. A total of 244 complete transcripts were identified in the transcriptome of C. possanii. Of these, 109 sequences showed identity to toxins that act on ion channels, 47 enzymes, 17 protease inhibitors (PINs), 11 defense peptides (HDPs), and 60 in other components. In addition, a sample of the soluble venom obtained from this scorpion was analyzed using an Orbitrap Velos apparatus, which allowed for identification by liquid chromatography followed by mass spectrometry (LC-MS/MS) of 70 peptides and proteins: 23 toxins, 27 enzymes, 6 PINs, 3 HDPs, and 11 other components. Until now, this work has the highest number of scorpion venom components identified through omics technologies. The main novel findings described here were analyzed in comparison with the known data from the literature, and this process permitted some new insights in this field.