This Article
Right arrow Full Text
Right arrow Full Text (PDF)
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Services
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrowReprints and Permissions
Right arrow Copyright Information
Right arrow Books from ASM Press
Right arrow MicrobeWorld
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Fraefel, C.
Right arrow Articles by Ackermann, M.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Fraefel, C.
Right arrow Articles by Ackermann, M.

 Previous Article  |  Next Article 

Journal of Virology, January 2004, p. 389-398, Vol. 78, No. 1
0022-538X/04/$08.00+0     DOI: 10.1128/JVI.78.1.389-398.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.

Spatial and Temporal Organization of Adeno-Associated Virus DNA Replication in Live Cells

Cornel Fraefel,1* Anne Greet Bittermann,2 Hansruedi Büeler,3 Irma Heid,1 Thomas Bächi,2 and Mathias Ackermann1

Institute of Virology,1 Center for Microscopy,2 Institute of Molecular Biology, University of Zurich, Zurich, Switzerland3

Received 30 July 2003/ Accepted 16 September 2003

Upon cell entry, the genomes of herpes simplex virus type 1 (HSV-1) and adenovirus (Ad) associate with distinct nuclear structures termed ND10 or promyelocytic leukemia (PML) nuclear bodies (NBs). PML NB morphology is altered or disrupted by specific viral proteins as replication proceeds. We examined whether adeno-associated virus (AAV) replication compartments also associate with PML NBs, and whether modification or disruption of these by HSV-1 or Ad, both of which are helper viruses for AAV, is necessary at all. Furthermore, to add a fourth dimension to our present view of AAV replication, we established an assay that allows visualization of AAV replication in live cells. A recombinant AAV containing 40 lac repressor binding sites between the AAV inverted terminal repeats was constructed. AAV Rep protein and helper virus-mediated replication of this recombinant AAV genome was visualized by binding of enhanced yellow fluorescent protein-lac repressor fusion protein to double-stranded AAV replication intermediates. We demonstrate in live cells that AAV DNA replication occurs in compartments which colocalize with AAV Rep. Early after infection, the replication compartments were small and varied in numbers from 2 to more than 40 per cell nucleus. Within 4 to 8 h, individual small replication compartments expanded and fused to larger structures which filled out much of the cell nucleus. We also show that AAV replication compartments can associate with modified PML NBs in Ad-infected cells. In wild-type HSV-1-infected cells, AAV replication compartments and PML NBs did not coexist, presumably because PML was completely disrupted by the HSV-1 ICP0 protein. However, alteration or disruption of PML appears not to be a prerequisite for AAV replication, as the formation of replication compartments was normal when the ICP0 mutants HSV-1 dl1403 and HSV-1 FXE, which do not affect PML NBs, were used as the helper viruses; under these conditions, AAV replication compartments did not associate with PML NBs.

* Corresponding author. Mailing address: Institute of Virology, University of Zurich, Winterthurerstrasse 266a, CH-8057 Zurich, Switzerland. Phone: 41 1 635 8713. Fax: 41 1 635 8911. E-mail: cornelf{at}vetvir.unizh.ch.

Journal of Virology, January 2004, p. 389-398, Vol. 78, No. 1
0022-538X/04/$08.00+0     DOI: 10.1128/JVI.78.1.389-398.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.



This article has been cited by other articles:

  • de Oliveira, A. P., Glauser, D. L., Laimbacher, A. S., Strasser, R., Schraner, E. M., Wild, P., Ziegler, U., Breakefield, X. O., Ackermann, M., Fraefel, C. (2008). Live Visualization of Herpes Simplex Virus Type 1 Compartment Dynamics. J. Virol. 82: 4974-4990 [Abstract] [Full Text]  
  • Cervelli, T., Palacios, J. A., Zentilin, L., Mano, M., Schwartz, R. A., Weitzman, M. D., Giacca, M. (2008). Processing of recombinant AAV genomes occurs in specific nuclear structures that overlap with foci of DNA-damage-response proteins. J. Cell Sci. 121: 349-357 [Abstract] [Full Text]  
  • Nash, K., Chen, W., Muzyczka, N. (2008). Complete In Vitro Reconstitution of Adeno-Associated Virus DNA Replication Requires the Minichromosome Maintenance Complex Proteins. J. Virol. 82: 1458-1464 [Abstract] [Full Text]  
  • Cotmore, S. F., Gottlieb, R. L., Tattersall, P. (2007). Replication Initiator Protein NS1 of the Parvovirus Minute Virus of Mice Binds to Modular Divergent Sites Distributed throughout Duplex Viral DNA. J. Virol. 81: 13015-13027 [Abstract] [Full Text]  
  • Nash, K., Chen, W., McDonald, W. F., Zhou, X., Muzyczka, N. (2007). Purification of Host Cell Enzymes Involved in Adeno-Associated Virus DNA Replication. J. Virol. 81: 5777-5787 [Abstract] [Full Text]  
  • Glauser, D. L., Strasser, R., Laimbacher, A. S., Saydam, O., Clement, N., Linden, R. M., Ackermann, M., Fraefel, C. (2007). Live Covisualization of Competing Adeno-Associated Virus and Herpes Simplex Virus Type 1 DNA Replication: Molecular Mechanisms of Interaction. J. Virol. 81: 4732-4743 [Abstract] [Full Text]  
  • Glauser, D. L., Saydam, O., Balsiger, N. A., Heid, I., Linden, R. M., Ackermann, M., Fraefel, C. (2005). Four-Dimensional Visualization of the Simultaneous Activity of Alternative Adeno-Associated Virus Replication Origins. J. Virol. 79: 12218-12230 [Abstract] [Full Text]  
  • Geoffroy, M.-C., Epstein, A. L., Toublanc, E., Moullier, P., Salvetti, A. (2004). Herpes Simplex Virus Type 1 ICP0 Protein Mediates Activation of Adeno-Associated Virus Type 2 rep Gene Expression from a Latent Integrated Form. J. Virol. 78: 10977-10986 [Abstract] [Full Text]  


Copyright © 2009 by the American Society for Microbiology.   For an alternate route to Journals.ASM.org, visit: http://intl-journals.asm.org | More Info»