The Homeodomain Derived Peptide Penetratin Induces Curvature of Fluid Membrane Domains

Antonin Lamazière,Gérard Chassaing,Olivier Lambert,Jesus Ayala-Sanmartin,Germain Trugnan,Claude Wolf

doi:10.1371/journal.pone.0001938

Abstract

BackgroundProtein membrane transduction domains that are able to cross the plasma membrane are present in several transcription factors, such as the homeodomain proteins and the viral proteins such as Tat of HIV-1. Their discovery resulted in both new concepts on the cell communication during development, and the conception of cell penetrating peptide vectors for internalisation of active molecules into cells. A promising cell penetrating peptide is Penetratin, which crosses the cell membranes by a receptor and metabolic energy-independent mechanism. Recent works have claimed that Penetratin and similar peptides are internalized by endocytosis, but other endocytosis-independent mechanisms have been proposed. Endosomes or plasma membranes crossing mechanisms are not well understood. Previously, we have shown that basic peptides induce membrane invaginations suggesting a new mechanism for uptake, “physical endocytosis”.Methodology/Principal FindingsHerein, we investigate the role of membrane lipid phases on Penetratin induced membrane deformations (liquid ordered such as in “raft” microdomains versus disordered fluid “non-raft” domains) in membrane models. Experimental data show that zwitterionic lipid headgroups take part in the interaction with Penetratin suggesting that the external leaflet lipids of cells plasma membrane are competent for peptide interaction in the absence of net negative charges. NMR and X-ray diffraction data show that the membrane perturbations (tubulation and vesiculation) are associated with an increase in membrane negative curvature. These effects on curvature were observed in the liquid disordered but not in the liquid ordered (raft-like) membrane domains.Conclusions/SignificanceThe better understanding of the internalisation mechanisms of protein transduction domains will help both the understanding of the mechanisms of cell communication and the development of potential therapeutic molecular vectors. Here we showed that the membrane targets for these molecules are preferentially the fluid membrane domains and that the mechanism involves the induction of membrane negative curvature. Consequences on cellular uptake are discussed.

Highlights

The delivery of active molecules into cells requires the step of efficiently cross the plasma membrane barrier
We observed that different basic peptides were able to induce membrane invaginations in giant vesicles, a process suggesting ‘‘physical endocytosis’’ mechanism for the basic peptide uptake [14]
Several studies suggest that increase in negatively charged phospholipid concentrations favours Penetratin and other basic peptide sequences binding to membranes by electrostatic adsorption

Summary

Introduction

The delivery of active molecules into cells requires the step of efficiently cross the plasma membrane barrier. These domains are usually present in transcription factors, are rich in basic residues and are responsible for the internalisation of the proteins into the cell cytoplasm [1,2] This last property resulted in new concepts about cell communication, and in the development of various molecular vectors such as Penetratin, Tat peptide and transportam which comprise the so called cell penetrating peptides (CPP). Protein membrane transduction domains that are able to cross the plasma membrane are present in several transcription factors, such as the homeodomain proteins and the viral proteins such as Tat of HIV-1 Their discovery resulted in both new concepts on the cell communication during development, and the conception of cell penetrating peptide vectors for internalisation of active molecules into cells. We have shown that basic peptides induce membrane invaginations suggesting a new mechanism for uptake, ‘‘physical endocytosis’’

Methods

Results

Conclusion