Missiles of Mass Disruption: Composition and Glandular Origin of Venom Used as a Projectile Defensive Weapon by the Assassin Bug Platymeris rhadamanthus

Abstract

Assassin bugs (Reduviidae) produce venoms that are insecticidal, and which induce pain in predators, but the composition and function of their individual venom components is poorly understood. We report findings on the venom system of the red-spotted assassin bug Platymeris rhadamanthus, a large species of African origin that is unique in propelling venom as a projectile weapon when threatened. We performed RNA sequencing experiments on venom glands (separate transcriptomes of the posterior main gland, PMG, and the anterior main gland, AMG), and proteomic experiments on venom that was either defensively propelled or collected from the proboscis in response to electrostimulation. We resolved a venom proteome comprising 166 polypeptides. Both defensively propelled venom and most venom samples collected in response to electrostimulation show a protein profile similar to the predicted secretory products of the PMG, with a smaller contribution from the AMG. Pooled venom samples induce calcium influx via membrane lysis when applied to mammalian neuronal cells, consistent with their ability to cause pain when propelled into the eyes or mucus membranes of potential predators. The same venom induces rapid paralysis and death when injected into fruit flies. These data suggest that the cytolytic, insecticidal venom used by reduviids to capture prey is also a highly effective defensive weapon when propelled at predators.

Context‐dependent venom deployment and protein composition in two assassin bugs

“…In zoophagous Heteroptera, the salivary glands (also called venom glands) not only secrete enzymes for the digestion of animal tissue (Cohen, 1995(Cohen, , 1998 but also proteins and peptides that facilitate the capture of prey (Edwards, 1961;Walker et al, 2017Walker et al, , 2019. The rapid paralysis of insects attacked by predatory assassin bugs such as Rhinocoris carmelita Stål and Platymeris rhadamanthus Gerstaecker was initially attributed to the disruption of cell membranes by digestive enzymes rather than the action of neurotoxins (Edwards, 1961).…”

Anthelminthic Activity of Assassin Bug Venom against the Blood Fluke Schistosoma mansoni

“…In addition, venom necrotoxins and cytotoxins might be involved, both of which typically kill cells [ 44 ]. Redulysins have been found in the venoms of other assassin bugs and were defined as putative pore-forming proteins with a cytolytic motif [ 45 , 46 ]. Therefore, we assume that R. iracundus redulysins may play a role for the observed cytotoxic effects against schistosome stem cells, with support from other compounds.…”

Deadly Proteomes: A Practical Guide to Proteotranscriptomics of Animal Venoms

“…LC-MS/MS using DDA is the most common method for obtaining venom peptide fragmentation data in many laboratories, [22,25,[48][49][50][51][52]57,[65][66][67][68][69][70][71][72][73][74][75][76][77][78][79] including ours. [29,42,43,53,61,[80][81][82][83] These data are typically searched using an automated algorithm against a database of protein sequences to identify peptide primary structures. However the non-matched data from these experiments are also well suited to de novo sequencing, particularly for mass spectra collected from high mass resolving instruments.…”

“…For example, we have used this method to demonstrate that assassin bugs (Insecta: Reduviidae) produce two different complex venoms in separate gland lumens that can be injected separately, [80] and to observe the variations in venom composition in a time series of venom milked from a single individual. [81] Various software is available for more nuanced calculations of abundance based on spectral counts, ion chromatograms, and sequence features, such as FreeQuant [124] and Pep-C. [125] If spectral counts are used for quantification, it is important to note that data-dependent acquisition (DDA) methods may bias results, whereas data-independent acquisition (DIA) detection methods such as sequential window acquisition of all theoretical mass spectra MS (SWATH-MS) may yield more accurate and reproducible results. [126,127] Another label-free quantitation method that is highly suited to venom proteomes is the exponential modified protein abundance index (emPAI), [128] which estimates relative protein abundances based on the fraction of proteolytic fragments observed originating from each polypeptide compared to the total number of possible fragments (PAI).…”

“…[159] Similar methods have been used to examine the site of toxin production in wasps, [77] ants, [6] and assassin bugs. [80,81] Since the venom apparatus of assassin bugs comprises multiple chambers with separate secretory tissues and lumens, we analyzed protein and poly(A) + RNA extracted from each gland region to establish the source of venom, using LC-MS/MS and transcriptomics, respectively. Mass profiles of peptide toxins measured by MALDI-TOF-MS, and quantitation of larger toxins from spectral counts, indicated that two different gland regions, the anterior main gland and the posterior main gland, are responsible for the production of two different complex venoms (Figure 7).…”