The Triatoma Virus Structural Protein VP4 Induces Membrane Permeability through Dynamic Pores

Abstract

In naked viruses, membrane breaching is a key step that must be performed for genome transfer into the target cells. Despite its importance, the mechanisms behind this process remain poorly understood. The small protein VP4, codified in the genome of most Picornavirales order viruses, has been shown to be involved in membrane alterations. Here we have analyzed the permeabilization activity of the natively non-myristolated VP4 protein from Triatoma virus (TrV), a virus belonging to the Dicistroviridae family within the Picornavirales order. The VP4 protein was produced as a Maltose Binding Protein (MBP) fusion to achieve its successful expression. This recombinant VP4 protein is able to produce membrane permeabilization in model membranes in a lipid-dependent manner. The membrane-induced permeability was also influenced by the pH, being greater at higher pH values. We demonstrate that the permeabilization activity elicited by the protein occurs through discrete pores that are reversibly inserted on the membrane. Sizing experiments using fluorescent dextrans, cryo-electron microscopy imaging and other additional techniques showed that recombinant VP4 forms heterogeneous proteo-lipidic pores rather than common proteinaceous channels. These results show that VP4 protein is involved in the membrane alterations required for genome transfer or cell entry steps during dicistrovirus infection.