Red-on-Yellow Queen: Bio-Layer Interferometry Reveals Functional Diversity Within Micrurus Venoms and Toxin Resistance in Prey Species

Abstract

Snakes in the family Elapidae largely produce venoms rich in three-finger toxins (3FTx) that bind to the ${\alpha }_1$ subunit of nicotinic acetylcholine receptors (nAChRs), impeding ion channel activity. These neurotoxins immobilize the prey by disrupting muscle contraction. Coral snakes of the genus Micrurus are specialist predators who produce many 3FTx, making them an interesting system for examining the coevolution of these toxins and their targets in prey animals. We used a bio-layer interferometry technique to measure the binding interaction between 15 Micrurus venoms and 12 taxon-specific mimotopes designed to resemble the orthosteric binding region of the muscular nAChR subunit. We found that Micrurus venoms vary greatly in their potency on this assay and that this variation follows phylogenetic patterns rather than previously reported patterns of venom composition. The long-tailed Micrurus tend to have greater binding to nAChR orthosteric sites than their short-tailed relatives and we conclude this is the likely ancestral state. The repeated loss of this activity may be due to the evolution of 3FTx that bind to other regions of the nAChR. We also observed variations in the potency of the venoms depending on the taxon of the target mimotope. Rather than a pattern of prey-specificity, we found that mimotopes modeled after snake nAChRs are less susceptible to Micrurus venoms and that this resistance is partly due to a characteristic tryptophan $\to$ serine mutation within the orthosteric site in all snake mimotopes. This resistance may be part of a Red Queen arms race between coral snakes and their prey.

Keywords: Arms race; Coral snake; Elapid; Mimotope; Three-finger toxin.

Fig. 1

The binding of Micrurus toxins to muscular nAChR mimotopes varies greatly depending on the individual venom as well as the target: A Example results of Micrurus corallinus A venom binding to Dipsadine mimotope. Shaded regions indicate the area under the curve for each replicate that is then averaged. B Cells show the area under the curve (Mean ± Standard Deviation, $N=3$) for the association step of the binding of each venom to each mimotope. Phylogeny on the left displays relationships between the venoms tested (short-tailed and long-tailed clades indicated by arrows, 3FTx-heavy venoms in purple and PLA₂-heavy venoms in teal), while the one on top displays the relationship between organisms on which mimotopes were modeled (clade of mimotopes based on snake sequences indicated with arrow). Crotalus horridus venom was used as a negative control as it contains many related protein families, but none which target the nAChR (Rokyta et al. 2013, 2015). Phylogenetic topology for Micrurus species was primarily adapted from the results of two recent phylogenies (Jowers et al. ; Reyes-Velasco et al. 2020) and checked for consistency with previous findings (Slowinski ; Gutberlet Jr and Harvey ; Pyron et al. ; Figueroa et al. ; Lee et al. ; Lomonte et al. ; Zheng and Wiens ; Zaher et al. ; Jowers et al. ; Gómez et al. 2021)

Fig. 2

Snake nAChR are consistently less vulnerable to $\mathrm{\alpha }$-neurotoxins, partially due to the W187S mutation: response of non-snake mimotopes to each Micrurus venom are on average higher than the response of snake mimotopes to that same venom (indicated by the ratios greater than 1), similarly the pseudo-ancestral blind snake mimotope without the W187S mutation are more vulnerable than the normal blind snake mimotope