Early evolution of the venom system in lizards and snakes

Abstract

Among extant reptiles only two lineages are known to have evolved venom delivery systems, the advanced snakes and helodermatid lizards (Gila Monster and Beaded Lizard). Evolution of the venom system is thought to underlie the impressive radiation of the advanced snakes (2,500 of 3,000 snake species). In contrast, the lizard venom system is thought to be restricted to just two species and to have evolved independently from the snake venom system. Here we report the presence of venom toxins in two additional lizard lineages (Monitor Lizards and Iguania) and show that all lineages possessing toxin-secreting oral glands form a clade, demonstrating a single early origin of the venom system in lizards and snakes. Construction of gland complementary-DNA libraries and phylogenetic analysis of transcripts revealed that nine toxin types are shared between lizards and snakes. Toxinological analyses of venom components from the Lace Monitor Varanus varius showed potent effects on blood pressure and clotting ability, bioactivities associated with a rapid loss of consciousness and extensive bleeding in prey. The iguanian lizard Pogona barbata retains characteristics of the ancestral venom system, namely serial, lobular non-compound venom-secreting glands on both the upper and lower jaws, whereas the advanced snakes and anguimorph lizards (including Monitor Lizards, Gila Monster and Beaded Lizard) have more derived venom systems characterized by the loss of the mandibular (lower) or maxillary (upper) glands. Demonstration that the snakes, iguanians and anguimorphs form a single clade provides overwhelming support for a single, early origin of the venom system in lizards and snakes. These results provide new insights into the evolution of the venom system in squamate reptiles and open new avenues for biomedical research and drug design using hitherto unexplored venom proteins.

Dynamic evolution of venom proteins in squamate reptiles

“…Here, we use these new sequences of physiological proteins sourced from non-venom gland tissues to rigorously test the hypothesis of an early recruitment of nine toxin families into the venom arsenal of squamate reptiles. Evidence of toxin monophyly following the inclusion of sampled non-toxin gene homologues will provide strong support for the SEO hypothesis proposed by Fry et al 10 In contrast, evidence of toxin non-monophyly (that is, non-toxins nesting within toxin clades) would indicate that either multiple origins of venom have occurred in the squamate reptiles or that the recruitment of proteins into the venom gland may not be a one-way process, but also involve reverse recruitment of toxins into nonvenom functions outside the venom gland. To explicitly test these hypotheses, we utilised rigorous phylogenetic analyses alongside ancestral character state reconstructions to investigate the origin of venom, and the nature and frequency of recruitments into and from a toxin function in squamate venom protein families.…”

“…Both databases were interrogated using representative venom sequences utilised by Fry et al [10][11][12]29 . Sequence searches were undertaken for 12 toxin families identified as basal to the Toxicofera (AVIT, cystatin, cysteine-rich secretory protein, CVF, crotamine, hyaluronidase, kallikrein, lectin, natriuretic peptides, NGF, veficolin and vespryn) [10][11][12] . Contigs exhibiting a BLAST e-value cut-off of 1e − 05 were retained.…”

“…As the venom apparatus of Heloderma is confined to the lower jaw, whereas that of snakes is restricted to the upper jaw, the two systems had always been assumed to be non-homologous. However, this assumption was challenged by Fry et al 10 , who identified toxin-secreting glands in the lower jaws of additional lizards of the families Varanidae and Anguidae, and in both upper and lower jaws of representatives of the Iguania. As these three groups, together with the Helodermatidae and Serpentes (snakes), form a monophyletic group, Fry et al 10 postulated a single early origin (SEO) of venom at the base of that clade, termed the Toxicofera (see Supplementary Fig.…”

“…S1 for a phylogeny of the Toxicoferan reptiles). However, in the absence of strong morphological or developmental evidence of homology between the upper jaw glands of iguanian lizards and snakes, the key piece of evidence for the single origin of venom rested with toxin gene phylogenies, which showed the monophyly of lizard and snake toxin genes to the exclusion of non-venom homologues, and in some cases the lack of reciprocal monophyly of snake and lizard toxins 10 .…”

“…In the absence of available genomic sequences, non-toxin homologues of the venom toxins in these studies were derived from a variety of other vertebrate taxa, most commonly mammals or birds 7,[9][10][11][12] . This is potentially problematic, because failure to sample non-toxin homologues from the focal clade can lead to gene trees that falsely depict the toxin genes as monophyletic, leading to the erroneous conclusion that they are the result of a single recruitment event (Fig.…”