Ultrastructural Localization of the Herpes Simplex Virus Type 1 U _L 31, U _L 34, and U _S 3 Proteins Suggests Specific Roles in Primary Envelopment and Egress of Nucleocapsids

Abstract

The wild-type U L 31, U L 34, and U S 3 proteins localized on nuclear membranes and perinuclear virions; the U S 3 protein was also on cytoplasmic membranes and extranuclear virions. The U L 31 and U L 34 proteins were not detected in extracellular virions. U S 3 deletion caused (i) virion accumulation in nuclear membrane invaginations, (ii) delayed virus production onset, and (iii) reduced peak virus titers. These data support the herpes simplex virus type 1 deenvelopment-reenvelopment model of virion egress and suggest that the U S 3 protein plays an important, but nonessential, role in the egress pathway.Herpes simplex virus type 1 (HSV-1) virions contain a linear double-stranded DNA genome of approximately 152 kb that is packaged into an icosahedral capsid shell. An amorphous tegument layer surrounds the capsid and is, in turn, surrounded by an envelope composed of a host-derived lipid bilayer studded with viral integral membrane proteins. After the viral genome is replicated and packaged into capsids within the nucleus, assembled nucleocapsids acquire a primary lipid envelope by budding through the inner nuclear membrane (INM) into the space located between the inner and outer leaflets of the nuclear envelope (25,33). Whereas the derivation of the primary envelope from the INM is widely accepted, the route of transit of the nascent virions from the perinuclear space to the extracellular space is more controversial. An overview of the key players in herpesvirus egress and a comparison of the salient features of the two proposed envelopment models have been recently published (8,25).A single-step model of herpesvirus envelopment was proposed for the prototypical alphaherpesvirus HSV-1 (6,18,35,44). This model proposes that enveloped virions move through the endoplasmic reticulum (ER) and the Golgi apparatus in transport vesicles with concomitant modification of primary virion glycoproteins. The single-step envelopment model is supported by the observations that (i) enveloped particles within vesicles can be readily detected by electron microscopy and in fracture label studies (35, 44) and (ii) virion egress and virion-associated glycoprotein processing are both inhibited in cells treated with the ionophore monensin (18). On the other hand, neither of these observations can exclude the alternative deenvelopment-reenvelopment model. Such a model is supported by mounting ultrastructural and biochemical evidence (3,10,13,14,30,37,41,46,50) and has been proposed for HSV-1, other alphaherpesviruses such as varicella-zoster virus (VZV) and pseudorabies virus (PrV), and betaherpesviruses such as human cytomegalovirus. In this model, primary envelopment occurs by budding through the INM but the primary envelope surrounding the perinuclear virion is lost, presumably by fusion with the outer lamellae of the nuclear envelope. In a second step, reenvelopment occurs by wrapping of the nucleocapsid and its associated tegument with a lipid bilayer originating from a membranous cytoplasmic organelle bearing viral glycoprotein...

Bovine herpesvirus 1 infection and infectious bovine rhinotracheitis

“…In order to gain access to the cytoplasm, the virions operate a fusion of their primary envelope with the outer nuclear membrane. A viral kinase activity encoded by US3 is required for this de-envelopment since US3 deletion mutants of HSV-1 [164], PrV [93], and MDV [177] accumulate in the perinuclear space. Once translocated into the cytoplasm, naked capsids definitively acquire their mature tegument and secondary envelope by budding into a presumably trans Golgi compartment [126].…”

Transport and egress of herpes simplex virus in neurons

“…Despite these findings, pUL34 and pUL9 are unlikely to play significant roles during retrograde transport of HSV-1. Neither is present in mature HSV-1 virions [55,56], and pUL34 is not found in mature virions of the related PrV [57][58][59]. Furthermore, deletion of the UL34 gene does not prevent infection of cells by HSV-1 [60].…”

Mapping of Key Functions of the Herpes Simplex Virus 1 U S 3 Protein Kinase: the U S 3 Protein Can Form Functional Heteromultimeric Structures Derived from Overlapping Truncated Polypeptides

“…In the absence of the U S 3 protein kinase, capsids are retained in nuclei. Envelopment appears to be limited and occurs at the inner nuclear membrane invaginated into the nucleus (23,24,27). The presence of similar structures in cells infected with a mutant expressing only the U S 3.5 protein kinase suggests that U S 3.5 is less efficient in enabling the restructuring of the nuclear envelope to enable the release of capsids from nuclei (16).…”