Transgenic Tarantula Toxin: A novel tool to study mechanosensitive ion channels in Drosophila

Denis Beqja,Sunna Haidar,Mihail Nikolaev,Yilan Shen,Barry Denholm

doi:10.1016/j.jinsphys.2020.104116

Denis Beqja, Sunna Haidar + Show 3 more

Open Access

https://doi.org/10.1016/j.jinsphys.2020.104116

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Journal: Journal of insect physiology	Publication Date: Sep 28, 2020
Citations: 6	License type: cc-by

Affiliation: University of Edinburgh

Abstract

The tarantula venom toxin GsMTx4 is the only known specific inhibitor of cation-selective mechanosensitive ion channels (MSCs). Its specificity, potency, and ease of use on isolated tissues and cells have made it a powerful pharmacological tool to identify and probe the physiological function of MSCs. In some contexts, however, it would be desirable to deliver the toxin in a controlled way in vivo. Here we describe a novel tool to allow spatial and temporal control of GsMTx4 delivery in vivo in Drosophila. To test the tool, we targeted MSCs required for mechanical nociception in a specific subset of sensory neurons in intact larvae. Expression of GsMTx4 in these neurons results in robust inhibition of mechanical nociception, demonstrating the toxin is active when expressed in vivo. The tool will be particularly useful to manipulate MSC activity in a spatially and temporally-controlled manner to study their role in development, physiology and behaviour in intact, free moving animals.

Highlights

Venom from the tarantula Grammostola spatulata contains a potent and specific inhibitor of mechanosensitive ion channels (MSCs)
Two versions were generated: (i) a full-length variant containing the full Grammostola spatulata mechanotoxin #4 (GsMTx4) cDNA (GsMTx4FL) encoding a pre-pro-peptide including a N-terminal signal peptide and a pro-sequence, both of which are subsequently cleaved to make the active peptide; (ii) a variant consisting of the active peptide alone (GsMTx4-AP)
We tested the ability of GsMTx4 to block mechanical nociception by driving expression in ppk-GAL4 neurons

Summary

Introduction

Venom from the tarantula Grammostola spatulata contains a potent and specific inhibitor of mechanosensitive ion channels (MSCs). For these reasons we generated transgenic fly lines with a genetically encoded GsMTx4 gene under the control of the UAS-GAL4 system to allow spatial and temporal delivery of the toxin in the intact animal. We tested the function of the transgenic GsMTx4 lines by assessing their ability to block mechanical nociception in larval ppk-GAL4 sensory neurons – a subset of multidendritic neurons that tile the body wall and respond to a variety of external stimuli including mechanical force (Fig. 2A)[7].

Results

Conclusion