Restriction and recruitment-gene duplication and the origin and evolution of snake venom toxins

Abstract

Snake venom has been hypothesized to have originated and diversified through a process that involves duplication of genes encoding body proteins with subsequent recruitment of the copy to the venom gland, where natural selection acts to develop or increase toxicity. However, gene duplication is known to be a rare event in vertebrate genomes, and the recruitment of duplicated genes to a novel expression domain (neofunctionalization) is an even rarer process that requires the evolution of novel combinations of transcription factor binding sites in upstream regulatory regions. Therefore, although this hypothesis concerning the evolution of snake venom is very unlikely and should be regarded with caution, it is nonetheless often assumed to be established fact, hindering research into the true origins of snake venom toxins. To critically evaluate this hypothesis, we have generated transcriptomic data for body tissues and salivary and venom glands from five species of venomous and nonvenomous reptiles. Our comparative transcriptomic analysis of these data reveals that snake venom does not evolve through the hypothesized process of duplication and recruitment of genes encoding body proteins. Indeed, our results show that many proposed venom toxins are in fact expressed in a wide variety of body tissues, including the salivary gland of nonvenomous reptiles and that these genes have therefore been restricted to the venom gland following duplication, not recruited. Thus, snake venom evolves through the duplication and subfunctionalization of genes encoding existing salivary proteins. These results highlight the danger of the elegant and intuitive "just-so story" in evolutionary biology.

Keywords: evolution; gene duplication; neofunctionalization; snake venom; subfunctionalization.

Fig. 1.—

Restriction and recruitment. Duplicated genes may be either restricted or recruited to the venom gland, with recruitment dependent on the evolution of new combinations of transcription factor binding sites in upstream regulatory regions. Mutation/loss of regulatory regions is indicated with an X.

Fig. 2.—

Tissue distribution of putative toxin gene families. Many proposed toxin gene families are expressed in a wide range of tissues, including the salivary or venom gland and have therefore been restricted to the venom gland following duplication, not recruited. Tissue abbreviations: Sal, salivary gland; VG, venom gland; Bra, brain; Liv, liver; K, kidney; O, ovary; P, pooled tissue (see text for details). Species abbreviations: Ema, leopard gecko (Eublepharis macularius); Pre, royal python (Python regius); Oae, rough green snake (Opheodrys aestivus); Pgu, corn snake (Pantherophis guttatus); Eco, painted saw-scaled viper (Echis coloratus); Oha, king cobra (Ophiophagus hannah); Tel, garter snake (Thamnophis elegans).

Fig. 3.—

Maximum-likelihood tree of complement C3 genes. complement C3 genes are expressed in a diversity of tissues, including venom and salivary glands. Following a gene duplication event (marked with *, shaded dark gray) one paralog has been restricted to the venom gland in the king cobra (Ophiophagus hannah) and the monocled cobra (Naja kaouthia). The two distinct king cobra sequences most likely represent geographic variation between Indonesian and Chinese populations. An additional gene duplication event appears to have occurred in the Austrelaps superbus lineage (marked with +, shaded light gray). Lineages for which body (nonvenom gland) sequences are available are colored blue and bootstrap values for 500 replicates are shown above branches.