Glycoproteins gB and gH Are Required for Syncytium Formation but Not for Herpesvirus-Induced Nuclear Envelope Breakdown

Abstract

Herpesvirus nucleocapsids are assembled in the nucleus, whereas maturation into infectious virions takes place in the cytosol. Since, due to their size, nucleocapsids cannot pass the nuclear pores, they traverse the nuclear envelope by vesicle-mediated transport. Nucleocapsids bud at the inner nuclear membrane into the perinuclear space, forming primary enveloped particles and are released into the cytosol after fusion of the primary envelope with the outer nuclear membrane. The nuclear egress complex (NEC), consisting of the conserved herpesvirus proteins (p)UL31 and pUL34, is required for this process, whereas the viral glycoproteins gB and gH, which are essential for fusion during penetration, are not. We recently described herpesvirus-induced nuclear envelope breakdown (NEBD) as an alternative egress pathway used in the absence of the NEC. However, the molecular details of this pathway are still unknown. It has been speculated that glycoproteins involved in fusion during entry might play a role in NEBD. By deleting genes encoding glycoproteins gB and gH from the genome of NEBD-inducing pseudorabies viruses, we demonstrate that these glycoproteins are not required for NEBD but are still necessary for syncytium formation, again emphasizing fundamental differences in herpesvirus-induced alterations at the nuclear envelopes and plasma membranes of infected cells. Herpesvirus maturation takes place in the nucleus and in the cytoplasm of infected cells. After capsid assembly and DNA encapsidation in the nucleus, the newly formed nucleocapsids leave the nucleus by budding at the inner nuclear membrane and fission, resulting in a primary enveloped particle located in the perinuclear cleft. Subsequent fusion with the outer nuclear membrane releases the nucleocapsid into the cytosol, where final tegumentation and envelopment take place (reviewed in references 1-4).The nuclear egress complex (NEC) consisting of conserved herpesvirus proteins homologous to pUL31 and pUL34 of herpes simplex virus 1 (HSV-1) is necessary for efficient nuclear egress. The type II tail-anchored membrane protein pUL34 localizes to the nuclear envelope and interacts with the soluble nucleoplasmic pUL31, thereby linking it to the inner nuclear membrane (5, 6). The NEC recruits cellular (protein kinase C) and viral kinases (pUS3 and pUL13), which phosphorylate lamins, resulting in local dissolution of the nuclear lamina (7-10).In the absence of either pUL31 or pUL34, viral replication is severely impaired but not totally abolished (5,6,11,12). To uncover potential other pathways for leaving the nucleus, the residual infectivity of pseudorabies viruses (PrV) lacking pUL34 or pUL31 was used for serial passaging in cell culture. After repeated passaging infectious virus progeny designated as PrV-⌬UL34Pass and PrV⌬UL31Pass could be isolated which efficiently replicated in the absence of either component of the NEC. In contrast to wild-type PrV, the passaged mutants leave the nucleus via a fragmented nuclear envelope designated as nuclear escape, the...

Nuclear envelope breakdown induced by herpes simplex virus type 1 involves the activity of viral fusion proteins

“…The dependence on gB and gH function for NEBD seen here contrasts with a recent report showing that single deletions of gB or gH do not inhibit the NEBD induced by PrV variants ΔUL31Pass and ΔUL34Pass (Schulz et al, 2013). This parallels a difference in gB and gH function in de-envelopment fusion during capsid nuclear egress – double deletions of gB and gH do not inhibit de-envelopment fusion in PrV, but do inhibit in HSV infections (Farnsworth et al, 2007; Klupp et al, 2008).…”

The Knowns and Unknowns of Herpesvirus Nuclear Egress

“…However, we have not yet been able to identify the decisive factor. Viral glycoproteins involved in fusion, gB and gH, play no role in NE breakdown during infection (149). In contrast, although NE breakdown was also observed in HSV-1-infected cells, this was dependent on viral glycoproteins gB and gH but could not compensate for NEC function (12).…”

Human Cytomegalovirus Egress: Overcoming Barriers and Co-Opting Cellular Functions

“…These findings strongly argued that HSV gB, which has been shown to require expression of gH/gL for the fusion of incoming virions in permissive cells, was required for fusion at the ONM of primary enveloped capsids during nuclear egress. However, similar phenotypes have not been observed in PRV mutant viruses with deletions of homologous envelope glycoproteins [117]. To date, the differences between the nuclear egress phenotypes of these two closely related α-herpesviruses following deletion of essential glycoproteins have not been resolved.…”