The Price of Success: Biophysical Costs of Tetrodotoxin Resistance in Voltage-Gated Sodium Channels

Utah State University, Logan UT

Investigators

Abstract

The overall goal of this research is to gain a deeper understanding of the mechanisms by which evolution takes place at the molecular level. By studying different populations of a species of garter snake, some of which live in areas also inhabited by toxic newts, we have discovered structural changes in a protein known as the voltage-gated sodium channel that result in an greater level of resistance to the neurotoxin, tetrodotoxin (TTX. This protein is found in the membranes of all electrically excitable cells, including neurons, heart muscle, and skeletal muscle. Previous research revealed that unique changes in the amino acid sequence in skeletal muscle sodium channels resulted in a profound increase in TTX resistance in snakes that live with, and prey upon, toxic newts. Snakes that live in areas where there are no newts, however, have what would be considered a normal sequence of amino acids in their skeletal muscle sodium channels. Selective pressure has, therefore, resulted in molecular changes that may form the basis of speciation. These molecular changes, however, may have come at a cost. Coincidental studies found that highly TTX-resistant snakes have a lower maximum crawl speed than toxin-sensitive snakes. This project is therefore based on the hypothesis that the molecular changes that impart TTX-resistance may also result in decreased performance in sodium channels resulting in slower skeletal muscle contractions. This hypothesis will be tested by measuring a variety of biophysical properties in sodium channels from the populations of snakes previously studied, and comparing these properties to the level of toxin resistance. An alternative hypothesis is that sodium channel expression levels are altered in resistant snakes. This hypothesis will be tested by quantifying the number of sodium channels in snake muscle from TTX-resistant and TTX-sensitive populations. Toxin resistance in neuronal sodium channels will also be measured, based on the hypothesis that these channels must also be TTX-resistant in snakes that prey upon newts. The structure of neuronal sodium channels will be compared across different snake populations, as was previously done with skeletal muscle sodium channels. This project will further elucidate how this critically important protein works by determining whether there are molecular interactions between the channel pore, where TTX binds, and the voltage-sensitive structures that control channel gating, and how changes in protein structure in response to selective pressure form the basis of evolution at the level of a single, highly conserved molecule. This high profile research comes at a time when the principles of evolution are increasingly scrutinized by the public and in the media. The results of this project will provide a scientific perspective for the ongoing discussion about evolutionary principles. These results will be disseminated at local public schools, as well as at national scientific conferences, and in the popular and scientific press. This project will be performed at Utah State University, an institution with strong research programs for undergraduates and underrepresented minorities, and with an NSF ADVANCE grant to encourage women in science. The laboratory in which the research will be performed consistently employs members from these groups to engage them in the excitement of basic research.