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Journal of Virology, August 2009, p. 7440-7448, Vol. 83, No. 15
0022-538X/09/$08.00+0     doi:10.1128/JVI.02390-08
Copyright © 2009, American Society for Microbiology. All Rights Reserved.

Autographa californica Multiple Nucleopolyhedrovirus me53 (ac140) Is a Nonessential Gene Required for Efficient Budded-Virus Production{triangledown}

Jondavid de Jong,1 Basil M. Arif,2 David A. Theilmann,3 and Peter J. Krell1*

Department of Molecular and Cellular Biology, University of Guelph, Guelph, Ontario N1G 2W1, Canada,1 Great Lakes Forestry Centre, Sault Ste. Marie, Ontario P6A 2E5, Canada,2 Pacific Agri-Food Research Centre, Agriculture and Agri-Food Canada, Summerland, British Columbia V0H 1Z0, Canada3

Received 18 November 2008/ Accepted 13 May 2009

me53 is a highly conserved baculovirus gene found in all lepidopteran baculoviruses that have been fully sequenced to date. The putative ME53 protein contains a zinc finger domain and has been previously described as a major early transcript. We generated an me53-null bacmid (Ac{Delta}me53GFP), as well as a repair virus (AcRepME53:HA-GFP) carrying me53 with a C-terminal hemagglutinin (HA) tag, under the control of its native early and late promoter elements. Sf9 and BTI-Tn-5b1 cells transfected with Ac{Delta}me53GFP resulted in a 3-log reduction in budded-virus (BV) production compared to both the parental Autographa californica multiple nucleopolyhedrosis virus and the repair bacmids, demonstrating that although me53 is not essential for replication, replication is compromised in its absence. Our data also suggest that me53 does not affect DNA replication. Cell fractionation showed that ME53 is found in both the nucleus and the cytoplasm as early as 6 h postinfection. Deletion of the early transcriptional start site resulted in a 10- to 360-fold reduction of BV yield; however, deletion of the late promoter (ATAAG) resulted in a 160- to 1,000-fold reduction, suggesting that, in the context of BV production, ME53 is required both early and late in the infection cycle. Additional Western blot analysis of purified virions from the repair virus revealed that ME53:HA is associated with both BV and occlusion-derived virions. Together, these results indicate that me53, although not essential for viral replication, is required for efficient BV production.

* Corresponding author. Mailing address: Department of Molecular and Cellular Biology, University of Guelph, Guelph, Ontario N1G 2W1, Canada. Phone: (519) 824-4120, ext. 53368. Fax: (519) 837-1802. E-mail: pkrell{at}uoguelph.ca

{triangledown} Published ahead of print on 20 May 2009.

Journal of Virology, August 2009, p. 7440-7448, Vol. 83, No. 15
0022-538X/09/$08.00+0     doi:10.1128/JVI.02390-08
Copyright © 2009, American Society for Microbiology. All Rights Reserved.





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