Hepatitis C Virus E1 Protein Induces Modification of Membrane Permeability inE. coliCells

Abstract

The E1 gene of hepatitis C virus (HCV) has been cloned and expressed in BL21(DE3)pLys Escherichia coli strain by pET3a vector to analyze changes in membrane permeability produced by this protein. We showed that the expression of E1 (aa 192-383), as well as of two C-terminal fragments (aa 331-383 and aa 341-383) corresponding to the transmembrane (TM) region of this protein, induced a rapid lysis of cells. On the contrary, the expression of a mutant of E1 (aa 192-340), lacking the last 40 amino acids, did not cause cell lysis. The analysis of permeability changes revealed that modification of membrane permeability to several compounds were observed only in clones expressing E1 and C-terminal fragments, while the synthesis of the C-terminal-deleted mutant had little or no effect on permeability. These findings demonstrate that the TM domain of E1 protein has membrane-active properties that may be involved in some aspects of virus-cell interaction.

A New Determinant of Endoplasmic Reticulum Localization Is Contained in the Juxtamembrane Region of the Ectodomain of Hepatitis C Virus Glycoprotein E1

“…A possible explanation for this finding is that the TMD of E1 protein has a toxic effect on the cells. Interestingly, rapid cell lysis, probably due to permeability changes of the inner membrane, has been recently documented in prokaryotic cells expressing this region (34). These results were entirely consistent with the previously described identification of an ER retention signal in the TMD domain of E1 (9).…”

Suggested role of the Golgi apparatus and endoplasmic reticulum for crucial sites of hepatitis C virus replication in human lymphoblastoid cells infected in vitro

“…Furthermore, the observed subcellular localisation of the E2 envelope protein in large cytoplasmic vacuoles, on ER membranes and, in some cells, on plasma membrane, strongly suggest that this viral protein might be directly involved in both CPE and cell-cell fusion phenomena. As previously mentioned, it has been reported that the E1 and E2 viral envelope proteins display cell fusion activity and could modify the cell membrane permeability [Ciccaglione et al, 1998;Takikawa et al, 2000]. As demonstrated by TEM and confocal microscopic observations on reinfected cultures and by PCR results, the de novo released viral particles are able to reinfect other coresident new cells, thus maintaining long-term infection in the culture system.…”

“…The E1 and E2 envelope glycoproteins are transmembrane proteins and their extracellular domains are thought to play an important role in the interactions between the virus and its receptorial complex [Miyamura and Matsuura, 1993;Ishii et al, 1998;Pileri et al, 1998]. It has also been reported that these viral envelope proteins exhibit cell fusion activity and could modify cell membrane permeability [Ciccaglione et al, 1998;Takikawa et al, 2000]. The NS2 protein is known to be a metalloprotease [Hijikata et al, 1993;Reed et al, 1995].…”

Expression of HCV E1 Protein in Baculovirus-Infected Cells: Effects on Cell Viability and Apoptosis Induction

“…Although we cannot establish whether fulllength E1 and/or one of these products are responsible for inducing the cell death program, it is conceivable that Green fluorescence activation of apoptosis required the presence of the Cterminal region. This hypothesis is sustained (1) by the demonstration that this region of E1 induces cell lysis also in prokaryotic cells [2][3][4] and (2) by our preliminary data obtained in mammalian cells which indicated that transfection of the HCV sequence corresponding to the E1 transmembrane domain induced a significant decrease in cell viability in transfected cells caused by apoptosis activation. So, it seems that this portion of E1 is sufficient for the induction of a toxic effect both in prokaryotic and in eukaryotic cells.…”

“…In previous work, we demonstrated that the expression of HCV E1 protein in Escherichia coli is toxic and induces rapid cell lysis due to the alteration of membrane permeability [2,4]. This effect was due to an intrinsic ability of the TM region of E1 to modify the permeability of biological membranes with possibly important consequences on cell survival.…”