The Herpes Simplex Virus 1 Virion Host Shutoff Protein Enhances Translation of Viral Late mRNAs by Preventing mRNA Overload

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In a previous study, it was observed that cells infected with herpes simplex virus 2 (HSV-2) failed to accumulate stress granules (SGs) in response to oxidative stress induced by arsenite treatment. As a follow-up to this observation, we demonstrate here that disruption of arsenite-induced SG formation by HSV-2 is mediated by a virion component. Through studies on SG formation in cells infected with HSV-2 strains carrying defective forms of UL41, the gene that encodes vhs, we identify vhs as a virion component required for this disruption. Cells infected with HSV-2 strains producing defective forms of vhs form SGs spontaneously late in infection. In addition to core SG components, these spontaneous SGs contain the viral immediate early protein ICP27 as well as the viral serine/threonine kinase Us3. As part of these studies, we reexamined the frameshift mutation known to reside within the UL41 gene of HSV-2 strain HG52. We demonstrate that this mutation is unstable and can rapidly revert to restore wild-type UL41 following low-multiplicity passaging. Identification of the involvement of virion-associated vhs in the disruption of SG formation will enable mechanistic studies on how HSV-2 is able to counteract antiviral stress responses early in infection. In addition, the ability of Us3 to localize to stress granules may indicate novel roles for this viral kinase in the regulation of translation. IMPORTANCEEukaryotic cells respond to stress by rapidly shutting down protein synthesis and storing mRNAs in cytoplasmic stress granules (SGs). Stoppages in protein synthesis are problematic for all viruses as they rely on host cell machinery to synthesize viral proteins. Thus, many viruses target SGs for disruption or modification. Infection by herpes simplex virus 2 (HSV-2) was previously observed to disrupt SG formation induced by oxidative stress. In this follow-up study, we identify virion host shutoff protein (vhs) as a viral protein involved in this disruption. The identification of a specific viral protein involved in disrupting SG formation is a key step toward understanding how HSV-2 interacts with these antiviral structures. Additionally, this understanding may provide insights into the biology of SGs that may find application in studies on human motor neuron degenerative diseases, like amyotrophic lateral sclerosis (ALS), which may arise as a result of dysregulation of SG formation.

Section: Discussionmentioning

confidence: 99%

The Herpes Simplex Virus 2 Virion-Associated Ribonuclease vhs Interferes with Stress Granule Formation

Finnen

Hay

Dauber

et al. 2014

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“…SCG2 knockdown showed an increase in the amount of secreted HSV1, the opposite effect of what was observed in reovirus infections. These results suggest that SGG2 plays different roles in the secretory pathways of nonenveloped reoviruses and the enveloped HSV (25)(26)(27).…”

Section: Scg2 Is Differentially Regulated By Reovirus and Hsv Infectimentioning

confidence: 93%

“…SCG2 is able to induce the formation of secretory granule-like structures in nonneuroendocrine cells, indicating a crucial role of SCG2 in generating secretory vesicles, and functions in the packaging and sorting of peptide hormones and neuropeptides into secretory vesicles (5). Two domains on the SCG2 full-length protein (domain spanning human SCG2 residues 25 to 41 [hSCG2 [25][26][27][28][29][30][31][32][33][34][35][36][37][38][39][40][41] ] and hSCG2 334 -348 ) are target- …”

mentioning

confidence: 99%

Differential Reovirus-Specific and Herpesvirus-Specific Activator Protein 1 Activation of Secretogranin II Leads to Altered Virus Secretion

Berard

Severini

Coombs

2015

Viruses utilize host cell machinery for propagation and manage to evade cellular host defense mechanisms in the process. Much remains unknown regarding how the host responds to viral infection. We recently performed global proteomic screens of mammalian reovirus TIL-and T3D-infected and herpesvirus (herpes simplex virus 1 [HSV-1])-infected HEK293 cells. The nonenveloped RNA reoviruses caused an upregulation, whereas the enveloped DNA HSV-1 caused a downregulation, of cellular secretogranin II (SCG2). SCG2, a member of the granin family that functions in hormonal peptide sorting into secretory vesicles, has not been linked to virus infections previously. We confirmed SCG2 upregulation and found SCG2 phosphorylation by 18 h postinfection (hpi) in reovirus-infected cells. We also found a decrease in the amount of reovirus secretion from SCG2 knockdown cells. Similar analyses of cells infected with HSV-1 showed an increase in the amount of secreted virus. Analysis of the stress-activated protein kinase (SAPK)/Jun N-terminal protein kinase (JNK) pathway indicated that each virus activates different pathways leading to activator protein 1 (AP-1) activation, which is the known SCG2 transcription activator. We conclude from these experiments that the negative correlation between SCG2 quantity and virus secretion for both viruses indicates a virusspecific role for SCG2 during infection. IMPORTANCEMammalian reoviruses affect the gastrointestinal system or cause respiratory infections in humans. Recent work has shown that all mammalian reovirus strains (most specifically T3D) may be useful oncolytic agents. The ubiquitous herpes simplex viruses cause common sores in mucosal areas of their host and have coevolved with hosts over many years. Both of these virus species are prototypical representatives of their viral families, and investigation of these viruses can lead to further knowledge of how they and the other more pathogenic members of their respective families interact with the host. Here we show that secretogranin II (SCG2), a protein not previously studied in the context of virus infections, alters virus output in a virus-specific manner and that the quantity of SCG2 is inversely related to amounts of infectious-virus secretion. Herpesviruses may target this protein to facilitate enhanced virus release from the host. Viruses that infect a host cell rely on the host machinery to replicate while managing to evade the immune system. Thus, there is an intricate relationship between the host and the virus that can be characterized by measuring changes in the host's protein effector molecules during the course of infection. In a previous study (1), we identified numerous HEK293 protein abundance changes after infection with the prototypic mammalian reovirus subtype 1 Lang (T1L), including the upregulation of secretogranin II (SCG2) at 24 h postinfection (hpi).SCG2 is a protein in the structurally and functionally related granin family that also includes chromogranins A and B, secretogranin III, secretogranin V (se...

“…The ability of vhs to bind to translation initiation factors may also be indicative of a direct role for vhs in translation. Cap-independent translation via cis-acting elements and enhanced translation of viral late mRNAs have both been ascribed to vhs (41)(42)(43)(44). As vhs may interact with mRNA in two distinct manners and has the ability to bind eIF4A, eIF4B, and eIF4H, all of which are known SG components (45), predicting how vhs is able to disrupt arsenite-induced SG formation is not straightforward.…”

mentioning

confidence: 99%

Herpes Simplex Virus 2 Virion Host Shutoff Endoribonuclease Activity Is Required To Disrupt Stress Granule Formation

Finnen

Zhu

et al. 2016

We previously established that cells infected with herpes simplex virus 2 (HSV-2) are disrupted in their ability to form stress granules (SGs) in response to oxidative stress and that this disruption is mediated by virion host shutoff protein (vhs), a virionassociated endoribonuclease. Here, we test the requirement for vhs endoribonuclease activity in disruption of SG formation. We analyzed the ability of HSV-2 vhs carrying the point mutation D215N, which ablates its endoribonuclease activity, to disrupt SG formation in both transfected and infected cells. We present evidence that ablation of vhs endoribonuclease activity results in defects in vhs-mediated disruption of SG formation. Furthermore, we demonstrate that preformed SGs can be disassembled by HSV-2 infection in a manner that requires vhs endoribonuclease activity and that, befitting this ability to promote SG disassembly, vhs is able to localize to SGs. Together these data indicate that endoribonuclease activity must be maintained in order for vhs to disrupt SG formation. We propose a model whereby vhs-mediated destruction of SG mRNA promotes SG disassembly and may also prevent SG assembly. IMPORTANCEStress granules (SGs) are transient cytoplasmic structures that form when a cell is exposed to stress. SGs are emerging as potential barriers to viral infection, necessitating a more thorough understanding of their basic biology. We identified virion host shutoff protein (vhs) as a herpes simplex virus 2 (HSV-2) protein capable of disrupting SG formation. As mRNA is a central component of SGs and the best-characterized activity of vhs is as an endoribonuclease specific for mRNA in vivo, we investigated the requirement for vhs endoribonuclease activity in disruption of SG formation. Our studies demonstrate that endoribonuclease activity is required for vhs to disrupt SG formation and, more specifically, that SG disassembly can be driven by vhs endoribonuclease activity. Notably, during the course of these studies we discovered that there is an ordered departure of SG components during their disassembly and, furthermore, that vhs itself has the capacity to localize to SGs. Stress granules (SGs) are dynamic, non-membrane-bound cytoplasmic compartments that contain translationally silent mRNAs that remain associated with a cadre of translation initiation proteins, poly(A) binding protein (PABP), and the small ribosomal subunit in the form of messenger ribonucleoprotein complexes (mRNPs) (1). SGs assemble rapidly in response to a variety of stressful conditions and likewise can disassemble rapidly when stress is alleviated. Natural stresses that eukaryotic cells encounter include starvation, elevated temperature (heat shock), oxidation, and virus infection, all of which can be sensed by four distinct eukaryotic initiation factor 2 (eIF2) kinases. Following the sensing of stress, these kinases become activated and phosphorylate the alpha subunit of eIF2 (eIF2␣), leading to a failure to initiate new rounds of translation (2). The ensuing stoppage in prote...