Assembly of complete, functionally active herpes simplex virus DNA replication compartments and recruitment of associated viral and cellular proteins in transient cotransfection assays.
AUTOR(ES)
Zhong, L
RESUMO
Early during the herpes simplex virus (HSV) lytic cycle or in the presence of DNA synthesis inhibitors, core viral replication machinery proteins accumulate in intranuclear speckled punctate prereplicative foci, some of which colocalize with numerous sites of host cellular DNA synthesis initiation known as replisomes. At later times, in the absence of inhibitors, several globular or large irregularly shaped replication compartments are formed; these compartments also contain progeny viral DNA and incorporate the IE175(ICP4) transcription factor together with several cellular proteins involved in DNA replication and repair. In this study, we demonstrate that several forms of both prereplication foci and active viral replication compartments that display an appearance similar to that of the compartments in HSV-infected cells can be successfully assembled in transient assays in DNA-transfected cells receiving genes encoding all seven essential HSV replication fork proteins together with oriS target plasmid DNA. Furthermore, bromodeoxyuridine (BrdU)-pulse-labeled DNA synthesis initiation sites colocalized with the HSV single-stranded DNA-binding protein (SSB) in these replication compartments, implying that active viral DNA replication may be occurring. The assembly of complete HSV replication compartments and incorporation of BrdU were both abolished by treatment with phosphonoacetic acid (PAA) and by omission of any one of the seven viral replication proteins, UL5, UL8, UL9, UL42, UL52, SSB, and Pol, that are essential for viral DNA replication. Consistent with the fact that both HSV IE175 and IE63(ICP27) localize within replication compartments in HSV-infected cells, the assembled HSV replication compartments were also able to recruit both of these essential regulatory proteins. Blocking viral DNA synthesis with PAA, but not omission of oriS, prevented the association of IE175 with prereplication structures. The assembled HSV replication compartments also redistributed cotransfected cellular p53 into the viral replication compartments. However, the other two HSV immediate-early nuclear proteins IE110(ICP0) and IE68(ICP22) did not enter the replication compartments in either infected or transfected cells.
ACESSO AO ARTIGO
http://www.pubmedcentral.nih.gov/articlerender.fcgi?artid=191447Documentos Relacionados
- Origin-Independent Assembly of Kaposi's Sarcoma-Associated Herpesvirus DNA Replication Compartments in Transient Cotransfection Assays and Association with the ORF-K8 Protein and Cellular PML
- Antibody-dependent cellular protection against herpes simplex virus dissemination as revealed by viral plauqe and infectivity assays.
- Proteomics of Herpes Simplex Virus Replication Compartments: Association of Cellular DNA Replication, Repair, Recombination, and Chromatin Remodeling Proteins with ICP8
- Evidence that the UL84 gene product of human cytomegalovirus is essential for promoting oriLyt-dependent DNA replication and formation of replication compartments in cotransfection assays.
- Proteins Specified by Herpes Simplex Virus: II. Viral Glycoproteins Associated with Cellular Membranes