Title : Restriction and recruitment – gene duplication and the origin and evolution of snake venom toxins

CC-BY-NC-ND 4.0 International license not peer-reviewed) is the author/funder. It is made available under a The copyright holder for this preprint (which was. Abstract The genetic and genomic mechanisms underlying evolutionary innovations are of fundamental importance to our understanding of animal evolution. Snake venom represents one such innovation and has been hypothesised to have originated and diversified via a process that involves duplication of genes encoding body proteins and subsequent recruitment of the copy to the venom gland where natural selection can act 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 (neofunctionalisation) is an even rarer process that requires the evolution of novel combinations of transcription factor binding sites in upstream regulatory regions. This hypothesis concerning the evolution of snake venom is therefore very unlikely. Nonetheless, it is often assumed to be established fact and this has hampered research into the true origins of snake venom toxins. We have generated transcriptomic data for a diversity of body tissues and salivary and venom glands from venomous and non-venomous reptiles, which has allowed us to critically evaluate this hypothesis. Our comparative transcriptomic analysis of venom and salivary glands and body tissues in five species of reptile reveals that snake venom does not evolve via the hypothesised process of duplication and recruitment of 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 non-venomous reptiles and have therefore been restricted to the venom gland following duplication, not recruited. Thus snake venom evolves via the duplication and subfunctionalisation of genes encoding existing salivary proteins. These results highlight the danger of the " just-so story " in evolutionary biology, where an elegant and intuitive idea is repeated so often that it assumes the mantle of established fact, to the detriment of the field as a whole.. CC-BY-NC-ND 4.0 International license not peer-reviewed) is the author/funder. It is made available under a The copyright holder for this preprint (which was .