Nuclear Herpesvirus Capsid Motility Is Not Dependent on F-Actin

Abstract

A considerable part of the herpesvirus life cycle takes place in the host nucleus. While much progress has been made to understand the molecular processes required for virus replication in the nucleus, much less is known about the temporal and spatial dynamics of these events. Previous studies have suggested that nuclear capsid motility is directed and dependent on actin filaments (F-actin), possibly using a myosin-based, ATP-dependent mechanism. However, the conclusions from these studies were indirect. They either relied on the effects of F-actin depolymerizing drugs to deduce an F-actin dependency or they visualized nuclear F-actin but failed to show a direct link to capsid motility. Moreover, no direct link between nuclear capsid motility and a molecular motor has been established. In this report, we reinvestigate the involvement of F-actin in nuclear herpesvirus capsid transport. We show for representative members of all three herpesvirus subfamilies that nuclear capsid motility is not dependent on nuclear F-actin and that herpesvirus infection does not induce nuclear F-actin in primary fibroblasts. Moreover, in these cells, three F-actin-inhibiting drugs failed to effect capsid motility. Only latrunculin A treatment stalled nuclear capsids but did so by an unexpected effect: the drug induced actin rods in the nucleus. Immobile capsids accumulated around actin rods, and immunoprecipitation experiments suggested that capsid motility stopped because latrunculin-induced actin rods nonspecifically bind nuclear capsids. Interestingly, capsid motility was unaffected in cells that do not induce actin rods. Based on these data, we conclude that herpesvirus nuclear capsid motility is not dependent on F-actin.

The diffusive way out: Herpesviruses remodel the host nucleus, enabling capsids to access the inner nuclear membrane

“…Phalloidin staining in fixed cells gave the same results. 12 We could only detect nuclear F-actin structures in infected superior cervical ganglion (SCG) neurons using Phalloidin as reported earlier, but not with Lifeact. We were therefore unable to directly visualize if nuclear F-actin structures support capsid motility because i) all cell types tested with the exception of primary cultures of SCG neurons did not induce detectable nuclear F-actin ii) Nuclear F-actin in SCG neurons could only be detected after fixation and staining with fluorescent phalloidin.…”

Remodeling nuclear architecture allows efficient transport of herpesvirus capsids by diffusion

“…However, further study of this hypothesis has been missing for more than a decade. We recently reinvestigated this idea, and did not find any evidence that herpesviruses induce nuclear F-actin (10).…”

“…Earlier, we and others provided data that suggested that herpesvirus capsids use an active, directed mechanism based on F-actin to transport through the nucleoplasm (8,9) and/or that nuclear F-actin is involved in capsid assembly. However, we recently showed that herpesviruses do not induce nuclear F-actin in most cells, and, more importantly, that nuclear capsid motility is not dependent on nuclear F-actin (10). This led us to reinvestigate nuclear capsid motility by single particle tracking to address whether molecular motors power it.…”

Phosphorylation of Bovine Herpesvirus 1 VP8 Plays a Role in Viral DNA Encapsidation and Is Essential for Its Cytoplasmic Localization and Optimal Virion Incorporation

“…The herpesvirus particle is composed of a capsid containing the double-stranded DNA genome, which is surrounded by a tegument layer and an envelope containing viral glycoproteins (1). During the herpesvirus life cycle, genome synthesis, capsid assembly, DNA incorporation (2), and primary tegumentation (3) occur in the nuclei of infected cells. Subsequently, nucleocapsids leave the nucleus by budding through the nuclear membrane via an envelopment-de-envelopment pathway (4,5).…”