The surface glycoprotein E2 of bovine viral diarrhoea virus contains an intracellular localization signal

Abstract

The intracellular transport of the surface glycoprotein E2 of bovine viral diarrhoea virus was analysed by expressing the cloned gene in the absence of other viral proteins. Immunofluorescence analysis and surface biotinylation indicated that E2 is located in an early compartment of the secretory pathway and not transported to the cell surface. In agreement with this result, E2 was found to contain only high-mannose oligosaccharide side-chains but no N-glycans of the complex type. To define the intracellular localization signal of the E2 protein, chimeric proteins were generated. E2 chimeras containing the MT (membrane anchor plus carboxy-terminal domain) of the G protein of vesicular stomatitis virus (VSV) or of the F protein of bovine respiratory syncytial virus (BRSV) were transported to the cell surface. On the other hand, VSV G protein containing the MT domain of E2 was detected only in the ER, indicating that this domain contains an ER localization signal. A chimeric E2 protein, in which not the membrane anchor but only the carboxy-terminal end was replaced by the corresponding domain of the BRSV F protein, was also localized in the ER. Therefore, it was concluded that the membrane anchor contains the ER localization signal of E2. Interestingly, the ER export signal within the VSV G protein cytoplasmic tail was found to overrule the ER localization signal in the E2 protein membrane anchor.

The Molecular Biology of Pestiviruses

“…Furthermore, it seems that misfolded E2 cannot associate with E1 (Branza-Nichita et al, 2001). E2 is retained in the cell through its transmembrane anchor (Grummer et al, 2001;K€ ohl et al, 2004). More specifically, R1047 in the transmembrane region plays a role in the intracellular retention of E2, as mutation of this amino acid led to cell surface localization of E2 (K€ ohl et al, 2004).…”

“…Intracellular budding has long been proposed for pestiviruses. All three envelope glycoproteins are retained in the cells (Burrack et al, 2012;Grummer et al, 2001;K€ ohl et al, 2004;C. Radtke & B.A.…”

Sequence-optimised E2 constructs from BVDV-1b and BVDV-2 for DNA immunisation in cattle

“…This was surprising because glycoprotein E2 is normally not transported to the cell surface but retained in the endoplasmic reticulum [34]. This is due to the presence of an arginine residue within the membrane anchor of E2 [34]. The arginine residue is also observed in the amino-acid sequence of membrane anchors of our recombinant E2.…”

“…However, surface localisation of E2 has also been observed in cells infected with recombinant baculovirus [39] or recombinant VSV [21] that express E2. As suggested by Köhl et al [34,35] it is conceivable that, in some transfected cells, the cell retention machinery becomes saturated by overexpressed E2 glycoproteins which are then transported to the cell surface.…”

“…A proportion of COS cells transfected with pCMV-LPsynE2.1 and pCMVLPsynE2 expressed E2 at the cell surface. This was surprising because glycoprotein E2 is normally not transported to the cell surface but retained in the endoplasmic reticulum [34]. This is due to the presence of an arginine residue within the membrane anchor of E2 [34].…”