Comparison of the Intranuclear Distributions of Herpes Simplex Virus Proteins Involved in Various Viral Functions

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Herpes simplex virus 1 (HSV-1) forms replication compartments (RCs), domains in which viral DNA replication, late-gene transcription, and encapsidation take place, in the host cell nucleus. The formation of these domains leads to compression and marginalization of host cell chromatin, which forms a dense layer surrounding the viral RCs and constitutes a potential barrier to viral nuclear egress or primary envelopment at the inner nuclear membrane. Surrounding the chromatin layer is the nuclear lamina, a further host cell barrier to egress. In this study, we describe an additional phase in RC maturation that involves disruption of the host chromatin and nuclear lamina so that the RC can approach the nuclear envelope. During this phase, the structure of the chromatin layer is altered so that it no longer forms a continuous layer around the RCs but instead is fragmented, forming islands between which RCs extend to reach the nuclear periphery. Coincident with these changes, the nuclear lamina components lamin A/C and lamin-associated protein 2 appear to be redistributed via a mechanism involving the U L 31 and U L 34 gene products. Viruses in which the U L 31 or U L 34 gene has been deleted are unable to undergo this phase of chromatin reorganization and lamina alterations and instead form RCs which are bounded by an intact host cell chromatin layer and nuclear lamina. We postulate that these defects in chromatin restructuring and lamina reorganization explain the previously documented growth defects of these mutant viruses.Herpes simplex virus 1 (HSV-1) is a large double-stranded DNA virus which replicates in the nucleus of infected host cells. Viral DNA replication and late gene transcription occur in replication compartments (RCs), nuclear domains which are formed when viral replication successively annexes large portions of the nucleus during the early stages of infection. During the later stages of replication, assembly of viral capsids occurs and DNA packaging takes place in RCs. Following this, capsids move to the inner nuclear membrane (INM), where primary envelopment takes place as they bud into the perinuclear space, acquiring an envelope derived from the INM. Herpesvirus particles are then thought to fuse with the outer nuclear membrane, releasing viral nucleocapsids into the host cell cytosol (17, 28). Finally, the nucleocapsid undergoes secondary envelopment and egress from the cell (24).During RC formation and annexation of space in the host cell nucleus, cellular chromatin is marginalized and compressed (18,26). This results in a layer of host cell chromatin surrounding the RC, which potentially constitutes a barrier through which viral capsids must move to reach the INM. Thus, to bud through the INM, viral nucleocapsids must move through the compacted host chromatin and the nuclear lamina. The nuclear lamina, a proteinaceous layer underlying the INM, is composed of integral INM proteins such as lamin-associated protein 2 (LAP2) and lamin B receptor, which interact with lamin proteins A, B, and C (2, ...

Section: Discussionsupporting

confidence: 63%

Section: Discussionmentioning

confidence: 99%

Herpes Simplex Virus 1 U _L 31 and U _L 34 Gene Products Promote the Late Maturation of Viral Replication Compartments to the Nuclear Periphery

Simpson-Holley

Baines

Roller

et al. 2004

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“…While the seven HSV DNA replication proteins are known, it is currently unclear as to what host proteins are involved in viral DNA replication. In addition to its role in DNA synthesis, ICP8 has been shown to affect viral transcription in at least two ways: (i) by repressing transcription from input parental viral genomes (33-35) and (ii) by stimulating late gene transcription (32).ICP8 and a number of other viral proteins, including the aforementioned viral replication proteins, the major viral transactivator ICP4, the immediate-early protein ICP27, and the major capsid protein VP5 accumulate within intranuclear structures referred to as replication compartments (9,19,47,59,71,73). Many of the processes required for viral replication, including viral DNA synthesis (18,71,74), viral transcription (47,57,71,74,75), virion assembly, and DNA packaging (19,51,93,96), occur within replication compartments.…”

mentioning

confidence: 99%

“…ICP8 and a number of other viral proteins, including the aforementioned viral replication proteins, the major viral transactivator ICP4, the immediate-early protein ICP27, and the major capsid protein VP5 accumulate within intranuclear structures referred to as replication compartments (9,19,47,59,71,73). Many of the processes required for viral replication, including viral DNA synthesis (18,71,74), viral transcription (47,57,71,74,75), virion assembly, and DNA packaging (19,51,93,96), occur within replication compartments.…”

mentioning

confidence: 99%

Proteomics of Herpes Simplex Virus Replication Compartments: Association of Cellular DNA Replication, Repair, Recombination, and Chromatin Remodeling Proteinswith ICP8

Taylor

Knipe

2004

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210

262

In this study, we have used immunoprecipitation and mass spectrometry to identify over 50 cellular and viral proteins that are associated with the herpes simplex virus 1 (HSV-1) ICP8 single-stranded DNA-binding protein. Many of the coprecipitating cellular proteins are known members of large cellular complexes involved in (i) DNA replication or damage repair, including RPA and MSH6; (ii) nonhomologous and homologous recombination, including the catalytic subunit of the DNA-dependent protein kinase, Ku86, and Rad50; and (iii) chromatin remodeling, including BRG1, BRM, hSNF2H, BAF155, mSin3a, and histone deacetylase 2. It appears that DNA mediates the association of certain proteins with ICP8, while more direct protein-protein interactions mediate the association with other proteins. A number of these proteins accumulate in viral replication compartments in the infected cell nucleus, indicating that these proteins may have a role in viral replication. WRN, which functions in cellular recombination pathways via its helicase and exonuclease activities, is not absolutely required for viral replication, as viral yields are only very slightly, if at all, decreased in WRN-deficient human primary fibroblasts compared to control cells. In Ku70-deficient murine embryonic fibroblasts, viral yields are increased by almost 50-fold, suggesting that the cellular nonhomologous endjoining pathway inhibits HSV replication. We hypothesize that some of the proteins coprecipitating with ICP8 are involved in HSV replication and may give new insight into viral replication mechanisms.Herpes simplex virus 1 (HSV-1) is a large, double-stranded DNA virus that replicates in the host cell nucleus. HSV encodes over 80 gene products that contribute to viral replication in either cultured cells or animal hosts (76). Due to the limited size of the HSV-1 genome, the virus cannot code for every function required for its propagation; thus, HSV-1 must rely upon factors supplied by the host cell for replication. For example, HSV exclusively uses the host cell RNA polymerase II for the transcription of viral genes (4, 16). The exact number and identity of the cellular factors required for HSV replication is unknown, but the identification of such factors is an active area of research as it may shed light on mechanisms of viral replication, the cellular process, or the factor itself. It is this concept that induced us to identify cellular proteins that associate with HSV-1 ICP8.The HSV-1 single-stranded DNA-binding protein, ICP8, is a 128-kDa multifunctional zinc metalloprotein (31, 37) encoded by the U L 29 gene (61). ICP8, in concert with the other HSV DNA replication proteins, including the helicase-primase complex (U L 5, U L 8, U L 52), the origin-binding protein (U L 9), and the polymerase holoenzyme (U L 30/U L 42), is required for viral DNA synthesis (11,12). While the seven HSV DNA replication proteins are known, it is currently unclear as to what host proteins are involved in viral DNA replication. In addition to its role in DNA synthesis, IC...

“…Evidence for posttranscriptional effects of ICP27 include the observations that ICP27 binds RNA via its RGG motif (7,38,53,76), regulates pre-mRNA 3Ј processing (33,51,52), stabilizes labile 3Ј ends of mRNA (7), interacts with cellular protein p32 (8) and spliceosome-associated protein 145 (9), inhibits the splicing of both viral and cellular transcripts (34), induces retention of intron-containing transcripts in the nucleus (61) but shuttles viral intronless transcripts from the nucleus to the cytoplasm (60,76,83), and regulates distribution of host small nuclear ribonucleoproteins (snRNPs) (59,78). In addition, ICP27 has been found to interact with ICP0 and ICP4 (54,57), to colocalize with ICP4 within HSV replication compartments in infected cells (23) and transfected cells (102), and to affect the posttranslational modification and DNA-binding ability of ICP4 (57,97,98). An additional protein needed for late gene transcription is the single-stranded-DNA-binding protein, ICP8, one of the seven viral ␤ proteins necessary and sufficient for viral DNA replication in transfected cells (13,96).…”

mentioning

confidence: 99%

Association of Herpes Simplex Virus Type 1 ICP8 and ICP27 Proteins with Cellular RNA Polymerase II Holoenzyme

Zhou

Knipe

2002

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Herpes simplex virus 1 (HSV-1) infection causes the shutoff of host gene transcription and the induction of a transcriptional program of viral gene expression. Cellular RNA polymerase II is responsible for transcription of all the viral genes, but several viral proteins stimulate viral gene transcription. ICP4 is required for all delayed-early and late gene transcription, ICP0 stimulates transcription of viral genes, and ICP27 stimulates expression of some early genes and transcription of at least some late viral genes. The early DNA-binding protein, ICP8, also stimulates late gene transcription. We therefore investigated which HSV proteins interact with RNA polymerase II. Using immunoprecipitation and Western blotting methods, we observed the coprecipitation of ICP27 and ICP8 with RNA polymerase II holoenzyme. The association of ICP27 with RNA polymerase II was detectable as early as 3 h postinfection, while ICP8 association became evident by 5 h postinfection, and the association of both was independent of viral DNA synthesis. Infections with ICP27 gene mutant viruses revealed that ICP27 is required for the association of ICP8 with RNA polymerase II, while studies with ICP8 gene deletion mutants showed no apparent role for ICP8 in the association of ICP27 with RNA polymerase II. The association of ICP27 and ICP8 with RNA polymerase II holoenzyme appeared to be independent of nucleic acids. We hypothesize that the interaction of ICP27 with RNA polymerase II holoenzyme reflects its role in stimulating early and late gene expression and/or its role in inhibiting host transcription and that the interaction of ICP8 with RNA polymerase II holoenzyme reflects its role in stimulating late gene transcription