Transduction of the Scorpion Toxin Maurocalcine into Cells: EVIDENCE THAT THE TOXIN CROSSES THE PLASMA MEMBRANE

Eric Estève,Kamel Mabrouk,Alain Dupuis,Sophia Smida-Rezgui,Xavier Altafaj,Didier Grunwald,Jean-Claude Platel,Nicolas Andreotti,Isabelle Marty,Jean-Marc Sabatier,Michel Ronjat,Michel De Waard

doi:10.1074/jbc.m412521200

Abstract

Maurocalcine (MCa) is a 33-amino-acid residue peptide toxin isolated from the scorpion Scorpio maurus palmatus. External application of MCa to cultured myotubes is known to produce Ca2+ release from intracellular stores. MCa binds directly to the skeletal muscle isoform of the ryanodine receptor, an intracellular channel target of the endoplasmic reticulum, and induces long lasting channel openings in a mode of smaller conductance. Here we investigated the way MCa proceeds to cross biological membranes to reach its target. A biotinylated derivative of MCa was produced (MCa(b)) and complexed with a fluorescent indicator (streptavidine-cyanine 3) to follow the cell penetration of the toxin. The toxin complex efficiently penetrated into various cell types without requiring metabolic energy (low temperature) or implicating an endocytosis mechanism. MCa appeared to share the same features as the so-called cell-penetrating peptides. Our results provide evidence that MCa has the ability to act as a molecular carrier and to cross cell membranes in a rapid manner (1-2 min), making this toxin the first demonstrated example of a scorpion toxin that translocates into cells.

Highlights

Maurocalcine (MCa) is a 33-amino-acid residue peptide toxin isolated from the scorpion Scorpio maurus palmatus
With the recent identification of a ryanodine receptor type 1 (RyR1) binding site for MCa [6] and earlier evidence that MCa alters the gating of purified RyR1 channels reconstituted in lipid bilayers, we favor the hypothesis that MCa can reach its intracellular target directly by crossing the plasma membrane
We demonstrated that MCa has the ability to translocate into cells by crossing the plasma membrane

Summary

Introduction

Maurocalcine (MCa) is a 33-amino-acid residue peptide toxin isolated from the scorpion Scorpio maurus palmatus. Synthetic peptides corresponding to these domains are called cell-penetrating peptides (CPPs) and have in common the inherent trait of containing many basic residues (arginine and lysine), often oriented toward the same face of the molecule (Fig. 1B) Because of this structural feature, CPPs possess the interesting ability to cross biological membranes in a receptor- or transporter-independent manner [11,12,13]. These peptides seem to target the lipid bilayer directly, using complementary charges, and penetrate the cell through a mechanism called translocation, which remains unclear. Regardless of the precise mode of entry, CPPs possess unique features worth mentioning: (i) delivery in 100% of the cells [17], (ii) delivery in all cell types both in vitro and in vivo [18, 19], (iii) the ability to cross the blood-brain barrier [18], and (iv) the potential to act as carriers and to translocate large compounds, such as proteins and oligonucleotides [20]

Methods

Results

Conclusion