Molecular Characterization and Functional Analysis of the Nattectin-like Toxin from the Venomous Fish Thalassophryne maculosa.

TmC4-47.2 is a toxin with myotoxic activity found in the venom of Thalassophryne maculosa, a venomous fish commonly found in Latin America whose envenomation produces an injury characterized by delayed neutrophil migration, production of major pro-inflammatory cytokines, and necrosis at the wound site, as well as a specific systemic immune response. However, there are few studies on the protein structure and functions associated with it. Here, the toxin was identified from the crude venom by chromatography and protein purification systems. TmC4-47.2 shows high homology with the Nattectin from Thalassophryne nattereri venom, with 6 cysteines and QPD domain for binding to galactose. We confirm its hemagglutinating and microbicide abilities independent of carbohydrate binding, supporting its classification as a nattectin-like lectin. After performing the characterization of TmC4-47.2, we verified its ability to induce an increase in the rolling and adherence of leukocytes in cremaster post-capillary venules dependent on the α5ß1 integrin. Finally, we could observe the inflammatory activity of TmC4-47.2 through the production of IL-6 and eotaxin in the peritoneal cavity with sustained recruitment of eosinophils and neutrophils up to 24 h. Together, our study characterized a nattectin-like protein from T. maculosa, pointing to its role as a molecule involved in the carbohydrate-independent agglutination response and modulation of eosinophilic and neutrophilic inflammation.

Molecular characterization and functional analysis of the nattectin-like toxin from the venomous fish thalassophryne maculosa

TmC4-47.2 is a toxin with myotoxic activity found in the venom of Thalassophryne maculosa, a venomous fish commonly found in Latin America whose envenomation produces an injury characterized by delayed neutrophil migration, production of major pro-inflammatory cytokines, and necrosis at the wound site, as well as a specific systemic immune response. However, there are few studies on the protein structure and functions associated with it. Here, the toxin was identified from the crude venom by chromatography and protein purification systems. TmC4-47.2 shows high homology with the Nattectin from Thalassophryne nattereri venom, with 6 cysteines and QPD domain for binding to galactose. We confirm its hemagglutinating and microbicide abilities independent of carbohydrate binding, supporting its classification as a nattectin-like lectin. After performing the characterization of TmC4-47.2, we verified its ability to induce an increase in the rolling and adherence of leukocytes in cremaster post-capillary venules dependent on the α5β1 integrin. Finally, we could observe the inflammatory activity of TmC4-47.2 through the production of IL-6 and eotaxin in the peritoneal cavity with sustained recruitment of eosinophils and neutrophils up to 24 h. Together, our study characterized a nattectin-like protein from T. maculosa, pointing to its role as a molecule involved in the carbohydrate-independent agglutination response and modulation of eosinophilic and neutrophilic inflammation.

Analysis of the intersexual variation in Thalassophryne maculosa fish venoms.

Gender related variation in the molecular composition of venoms and secretions have been described for some animal species, and there are some evidences that the difference in the toxin (s) profile among males and females may be related to different physiopathological effects caused by the envenomation by either gender. In order to investigate whether this same phenomenon occurs to the toadfish Thalassophryne maculosa, we have compared some biological and biochemical properties of female and male venoms. Twenty females and males were collected in deep waters of the La Restinga lagoon (Venezuela) and, after protein concentration assessed, the induction of toxic activities in mice and the biochemical properties were analyzed. Protein content is higher in males than in females, which may be associated to a higher size and weight of the male body. In vivo studies showed that mice injected with male venoms presented higher nociception when compared to those injected with female venoms, and both venoms induced migration of macrophages into the paw of mice. On the other hand, mice injected with female venoms had more paw edema and extravasation of Evans blue in peritoneal cavity than mice injected with male venoms. We observed that the female venoms had more capacity for necrosis induction when compared with male venoms. The female samples present a higher proteolytic activity then the male venom when gelatin, casein and FRETs were used as substrates. Evaluation of the venoms of females and males by SDS-PAGE and chromatographic profile showed that, at least three components (present in two peaks) are only present in males. Although the severity of the lesion, characterized by necrosis development, is related with the poisoning by female specimens, the presence of exclusive toxins in the male venoms could be associated with the largest capacity of nociception induction by this sample.