The Portal Vertex of KSHV Promotes Docking of Capsids at the Nuclear Pores

Abstract

Kaposi’s sarcoma-associated herpesvirus (KSHV) is a cancer-related herpesvirus. Like other herpesviruses, the KSHV icosahedral capsid includes a portal vertex, composed of 12 protein subunits encoded by open reading frame (ORF) 43, which enables packaging and release of the viral genome into the nucleus through the nuclear pore complex (NPC). Capsid vertex-specific component (CVSC) tegument proteins, which directly mediate docking at the NPCs, are organized on the capsid vertices and are enriched on the portal vertex. Whether and how the portal vertex is selected for docking at the NPC is unknown. Here, we investigated the docking of incoming ORF43-null KSHV capsids at the NPCs, and describe a significantly lower fraction of capsids attached to the nuclear envelope compared to wild-type (WT) capsids. Like WT capsids, nuclear envelope-associated ORF43-null capsids co-localized with different nucleoporins (Nups) and did not detach upon salt treatment. Inhibition of nuclear export did not alter WT capsid docking. As ORF43-null capsids exhibit lower extent of association with the NPCs, we conclude that although not essential, the portal has a role in mediating the interaction of the CVSC proteins with Nups, and suggest a model whereby WT capsids can dock at the nuclear envelope through a non-portal penton vertex, resulting in an infection ‘dead end’.

Role of HSV-1 Capsid Vertex-Specific Component (CVSC) and Viral Terminal DNA in Capsid Docking at the Nuclear Pore

“…Combined, our observations suggest that reduced levels of CVSC proteins on A-and B-capsids compared to that of C-capsids, result in lower capsid-NPC binding. Our findings are supported by recently published data for Kaposi's sarcoma-associated herpesvirus (KSHV) showing that KSHV capsids (with capsid structure homologous to HSV-1 capsids) dock at the NPCs even when portal-ring-forming protein ORF43 is deleted (ORF43-null KSHV capsids) [15]. Based on this result, authors suggest that capsid-NPC binding is mediated by CVSC complexes at non-portal penton vertices (this type of non-portal binding results in infection "dead-end" particles since DNA cannot be ejected) [15].…”

“…It has been observed that docking of HSV-1 at the NPC is mediated by the minor capsid protein UL25, which binds Nup214 situated on the cytoplasmic NPC surface [10,11], and by tether tegument protein UL36, which binds Nup358 localized on the cytoplasmic NPC filaments [9,14] (gene names are used to refer to their respective proteins). It was suggested that these interactions ensure alignment of the capsid portal with the NPC opening pore, facilitating DNA ejection into the nucleoplasm [15]. The HSV-1 capsid is comprised mainly of the major capsid protein VP5 (UL19), organized in 11 pentameric (penton forming vertices) and 150 hexameric (hexon) capsomer subunits [16].…”

Intranuclear HSV-1 DNA ejection induces major mechanical transformations suggesting mechanoprotection of nucleus integrity

“…Using a pull-down assay combined with qPCR, we showed earlier that HSV-1 capsid–NPC binding in isolated reconstituted nuclei triggers the ejection of viral DNA and its internalization in the nucleus, driven by capsid DNA pressure of ∼18 atm ( 10 ). A benefit of the reconstituted virus–nucleus system is that it can be used to isolate the effect of the central step of viral infection—capsid docking at the nucleus and viral genome uncoating—on nucleus mechanics while avoiding interference from other processes occurring within the cell cytoplasm during viral replication ( 12 ). There are, on average, more than a thousand NPCs covering a significant area of the nucleus surface ( 41 ).…”

“… 8 , 9 ) of cell-free reconstituted nuclei incubated with HSV-1 capsids, which dock at the nuclear pore complexes (NPCs) and eject their DNA into the nucleus ( 10 , 11 ). This isolated nucleus system recapitulates capsid–nucleus binding and viral DNA ejection (without DNA replication) in vivo without interference from trafficking kinetics of capsids moving toward the nucleus ( 12 ) while allowing direct probing of nucleus mechanics with the AFM cantilever. Previously, nucleus mechanics was investigated with AFM studies in intact cells ( 13 ).…”

The Ins and Outs of Herpesviral Capsids: Divergent Structures and Assembly Mechanisms across the Three Subfamilies

“…One conserved function most likely involves capsid docking at the nucleus and genome delivery. KSHV capsids lacking a portal complex do not efficiently dock at the nucleus [102], likely because the portal vertex also lacks the CATC. CATC proteins may also have subfamilyspecific functions.…”