Fig
Fig.5(a) shows NMuMG cell entry by non-neutralized gL+and gLvirions, with gN co-staining for comparison. this by holding gH in an antigenically unique heterodimer until after endocytosis. Second, gLvirions were more vulnerable to gB-directed neutralization. This covered multiple epitopes and thus seemed to reflect a general opening up of the gHgB access complex, which gL again normally restricts to late endosomes. gL therefore limits MuHV-4 neutralization by providing redundancy in cell binding and by keeping key elements of the virion fusion machinery hidden until after endocytosis. == Bosentan Hydrate INTRODUCTION == Most vaccines depend on eliciting neutralizing antibodies (Zinkernagel & Hengartner, 2006). Herpesvirus service providers remain infectious despite making antibody responses. Preventing herpesvirus infections by vaccination is usually therefore a difficult challenge. We are using murid herpesvirus 4 (MuHV-4) to understand gammaherpesvirus neutralization. MuHV-4 binds to cells via heparan sulfate, using either gp70, a product of ORF4 (Gilletet al., 2007a), or gHgL (Gilletet al., 2008a). Immune sera can block cell binding (Gillet al., 2006), but they block membrane fusion poorly, allowing opsonized virions to infect macrophages and dendritic cells via IgG Fc receptors (Rosaet al., 2007). Bypassing cell-binding blocks in this way is not unique to MuHV-4 (Inadaet al., 1985;Maidjiet al., Bosentan Hydrate 2006). How might herpesvirus membrane fusion be blocked better? Answering this means understanding how fusion works. Virus-specific glycoproteins, such as herpes simplex virus gD, can modulate fusion (Avitabileet al., 2007;Atanasiuet al., 2007) and some herpesviruses can express option fusion complexes by using different accessory glycoproteins (Borza & Hutt-Fletcher, 2002;Wang & Shenk, 2005), Bosentan Hydrate but the core machinery, comprising the gHgL heterodimer and gB (Browneet al., 2001), is usually conserved. MuHV-4 membrane fusion is usually pH-dependent and occurs in late endosomes (Gilletet al., 2008b). Fusion is usually associated with conformation changes in both gH and gB (Gilletet al., 2008b,c). gB probably switches between pre- and post-fusion says, like the structurally homologous vesicular stomatitis computer virus glycoprotein G (Rocheet al., 2007), but gH is different. It switches from a gL-dependent to a gL-independent conformation in late endosomes (Gilletet al., 2008c), implying that gH and gL dissociate. Yet gLvirions, which constitutively express the downstream form of gH (gH-only), remain infectious; indeed, they show premature rather than impaired membrane fusion (Gilletet al., 2008c). It therefore appears that gH changes from gHgL to gH-only before engaging in fusion. Not only is usually gH different in gLvirions: gB also shows conformational instability. This is consistent with a knock-on effect of the switch in gH, as gHgL and gB are associated in the virion membrane (Gillet & Stevenson, 2007a). Rabbit polyclonal to NF-kappaB p65.NFKB1 (MIM 164011) or NFKB2 (MIM 164012) is bound to REL (MIM 164910), RELA, or RELB (MIM 604758) to form the NFKB complex. A link probably intramembrane is usually managed between gH and gB, even without gL (Gillet & Stevenson, 2007a), but any extracellular conversation must switch, as the gHgL and gH-only conformations are antigenically very different (Gillet al., 2006). The gB N terminus covers a part of gHgL, and deleting it also seems to destabilize gB (Gillet & Stevenson, 2007b). This region may therefore bridge the gB and gHgL extracellular domains. The gB and gH conformation changes present problems for antibodies that would block membrane fusion (Gillet al., 2006;Gilletet al., 2006). First, antibodies must act indirectly, either by blocking conformation changes (probably the major mechanism for gHgL) or by causing steric hindrance (probably the major mechanism for gB) (Gilletet al., 2008b). Second, they must remain attached to their targets in late endosomes and compete with conformation changes that are energetically favourable at low pH. With glycan shielding (Gillet & Stevenson, 2007b) and poor immunogenicity (Gilletet al., 2007b) also factored in, it is perhaps unsurprising that total MuHV-4 neutralization is so hard. The central functions of gL in MuHV-4 cell binding and membrane fusion suggest an additional role for it in virion neutralization. Whether gL itself is usually a neutralization target is unknown, but gHgL is the major mAb-defined target on wild-type virions (Gillet Bosentan Hydrate al., 2006). This neutralization operates downstream of cell binding, presumably by inhibiting the post-endocytic dissociation of gL from gH. Disrupting gL would remove gHgL as a target, but could instead reveal other gH epitopes. In order to understand how gL affects neutralization, we compared the infectivity of gL+and gLvirions after exposure to immune sera or mAbs. Our results explain some of the resistance of wild-type MuHV-4 virions to neutralization and shed new light on herpesvirus access. == METHODS == == Mice, sera and mAbs. == Female C57BL/6 or BALB/c mice (Harlan UK) were infected intranasally with MuHV-4 when 68 weeks aged, in accordance with local ethics and Home Office.