Non-toxic venoms?

This article is part of a series highlighting new research in snake biology presented by herpetologists at the World Congress of Herpetology VII in Vancouver, British Columbia. If you want to learn more about the WCH, check out the June 2012 issue of Herpetological Review, or follow the Twitter hashtag #wch2012, with which I will tag all posts in this series.

A while back we heard about reasons why rattlesnakes frequently miss strikes at their squirrel prey. When they do hit their target, however, the prey tend to run off a little ways before kicking the bucket. This is because, although snake venoms are quick-acting, they mostly do not incapacitate immediately (although there are some sea snake venoms that are very, very quick). As a result, snakes that hunt using venom must be able to track down their prey after it has been bitten. This necessity has led Anthony Saviola, Steve Mackessy, and their colleagues at the University of Northern Colorado to an answer for a question that snake biologists have been asking for a long time: why do snake venoms contain molecules that are non-toxic?

Venom is a complex mixture of over one hundred proteins, peptides, enzymes, and small organic and inorganic molecules, many of which are toxic. It is essentially very strong saliva, capable of both breaking down food and chemically incapacitating or killing it. The evolution of venom has allowed many species of advanced snakes to utilize chemical rather than mechanical means of dispatching prey, which helps them avoid retaliation from sharp teeth and claws. Yet a few venom components serve neither to digest nor to kill prey. What on earth is their function?

Timber Rattlesnake, Crotalus horridus

Observations back to the 1960s suggest that rattlesnakes prefer the scent of envenomated mice over non-envenomated ones. When a rattlesnake strikes a prey item, a stereotypic behavior known as strike-induced chemosensory searching (SICS) is induced. This behavior is a fixed-action pattern that involves tongue-flicking to collect chemical information and a stereotyped searching movement pattern that allows the snake to determine which of the many chemical trails in its vicinity should be followed to find the envenomated prey.

To what chemical cues are rattlesnakes responding when they perform SICS? Mackessy separated Western Diamondback Rattlesnake (Crotalus atrox) venom into distinct fractions, each containing molecules of different sizes. This is accomplished by exploiting differences in the weight of each molecule, using techniques that allow different molecules to travel different distances through a gel medium based on how heavy they are. In the same amount of time, smaller, lighter molecules travel farther than larger, heavier ones. When Mackessy injected mice with the fractions containing the larger exonucleases, metalloproteinases, and phospholipases (classes of enzymes that break down, respectively, DNA, proteins, and fatty acids - so, the venom components that are active in digestion and incapacitation), the snakes showed little interest. Together, these well-known active compounds comprise about 85% of the total venom by mass, and they are responsible for all of the venom's digestive and killing activity, so it's a little surprising that snakes should show no interest in mice injected with them.

Structure of disintegrin heterodimer from Echis carinatus

However, when Mackessy injected mice with a venom fraction containing smaller molecules, called crotatroxin disintegrins, not known to have any enzymatic or toxic activity, the snakes behaved as they normally would have when scent-trailing mice injected with whole venom. Although disintegrins make up less than 10% of the venom by mass, they are clearly critical for the snake to find its prey following envenomation. What's more, the disintegrins alone don't induce trail-following behavior in snakes; rather, it is the product of the interaction of the disintegrins with the dead prey tissue that causes snakes to follow their trail.

Crotalus oreganus lutosus

Disintegrins are abundant in the venoms of most Western Rattlesnakes (Crotalus viridis sensu lato)¹, and are also found in other rattlesnakes, in Copperheads (Agkistrodon contortrix) and other snakes of the genus Agkistrodon (where they may be undergoing rapid evolution), and in most adult vipers. However, in juvenile rattlesnakes, and in many elapid snakes (cobras, coralsnakes, and other proteroglyphous snakes) disintegrins are present only at low levels or are absent entirely. Why should this be? One possible explanation is that elapids and juvenile vipers are more likely to hold onto their prey following a strike, so they don't need a chemical tracer to follow it because it doesn't usually get that far away. Some elapids do have disintegrins, however, and there is evidence that disintegrins in vipers also function to inhibit blood coagulation. In fact, a disintegrin molecule from Copperhead venom has been shown to slow the spread of breast and ovarian cancer in mice, so there could be much more to this story. One thing is sure: the Mackessy lab is one to watch if you're interested in snake venoms, and this won't be my last post on their fascinating research.

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1 Many of the nine previously recognized subspecies of the Western Rattlesnake (Crotalus viridis sensu lato) occur in the southwestern United States, and the complex has been subject to several molecular studies to reevaluate the taxonomic status of these subspecies. The consensus opinion largely follows Ashton and de Queiroz (2001), which recognized two species: C. viridis (Prairie Rattlesnake, two subspecies) and C. oreganus (Western Rattlesnake, six subspecies). This is the only species of rattlesnake that occurs where I live, in northeastern Utah.^↩

ACKNOWLEDGMENTS

Thanks to Todd Pierson for his photograph and to Steve Mackessy and Anthony Saviola for their coordination on this article, which was delayed in its release to coincide with the publication of their paper in BMC Biology.

REFERENCES

Ashton KG, de Queiroz A (2001) Molecular systematics of the western rattlesnake, Crotalus viridis (Viperidae), with comments on the utility of the D-loop in phylogenetic studies of snakes. Molecular Phylogenetics and Evolution 21:176-189. <link>

Calvete J, Sanz L, Juárez P, Mackessy S (2009) Snake venomics and disintegrins: portrait and evolution of a family of snake venom integrin antagonists. In: Mackessy S (ed) Handbook of Venoms and Toxins of Reptiles. CRC Press, Boca Raton, Florida, pp 337-357

Finn R (2001). Snake Venom Protein Paralyzes Cancer Cells. Journal of the National Cancer Institute 93:261-262 <link>

Furry K, Swain T, Chiszar D (1991) Strike-induced chemosensory searching and trail following by prairie rattlesnakes (Crotalus viridis) preying upon deer mice (Peromyscus maniculatus): chemical discrimination among individual mice. Herpetologica 47:69-78. <link>

Mackessy SP, Tu AT (1993) Biology of the sea snakes and biochemistry of their venoms. In: Tu AT (ed) Toxin-related Diseases: Poisons Originating from Plants, Animals and Spoilage. Oxford & IBH Publishing Co., New Delhi, pp 305-351 
<link>

Saviola AJ, Chiszar D, Busch C, Mackessy SP. 2013. Molecular basis for prey relocation in viperid snakes. BMC Biology 11 <link>

Soto JG et al. (2006) Genetic variation of a disintegrin gene found in the American copperhead snake (Agkistrodon contortrix). Gene 373:1-7. <link>

Life is Short, but Snakes are Long by Andrew M. Durso is licensed under a Creative Commons Attribution-NonCommercial-NoDerivs 3.0 Unported License.