Australian funnel-web spiders evolved human-lethal δ-hexatoxins for defense against vertebrate predators

Australian funnel-web spiders are infamous for causing human fatalities, which are induced by venom peptides known as delta-hexatoxins (delta-HXTXs). Humans and other primates did not fea-ture in the prey or predator spectrum during evolution of these spiders, and consequently the primate lethality of delta-HXTXs remains enigmatic. Funnel-web envenomations are mostly inflicted by male spiders that wander from their burrow in search of females during the mating season, which suggests a role for 5-HXTXs in self-defense since male spiders rarely feed during this period. Although 35 species of Australian funnel-web spiders have been described, only nine delta-HXTXs from four species have been characterized, resulting in a lack of understanding of the ecological roles and molecular evolution of delta-HXTXs. Here, by profiling venom-gland transcriptomes of 10 funnel-web species, we report 22 delta-HXTXs. Phylogenetic and evolutionary assessments reveal a remarkable sequence conservation of delta-HXTXs despite their deep evolutionary origin within funnel-web spiders, consistent with a defensive role. We demonstrate that delta-HXTX-Ar1a, the lethal toxin from the Sydney funnel-web spider Atrax robustus, induces pain in mice by inhibiting inactivation of voltage-gated sodium (NaV) channels involved in nociceptive signaling. delta-HXTX-Ar1a also inhibited inactivation of cockroach NaV channels and was insecticidal to sheep blowflies. Considering their algogenic effects in mice, potent insecticidal effects, and high levels of sequence conservation, we propose that the delta-HXTXs were repurposed from an initial insecticidal predatory function to a role in defending against nonhuman vertebrate predators by male spiders, with their lethal effects on humans being an unfortunate evolutionary coincidence. tain sodium the victims, out cur the other sensitive during