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Journal of Virology, June 2009, p. 5708-5717, Vol. 83, No. 11
0022-538X/09/$08.00+0     doi:10.1128/JVI.00300-09
Copyright © 2009, American Society for Microbiology. All Rights Reserved.

Restriction of Human Polyomavirus BK Virus DNA Replication in Murine Cells and Extracts{triangledown}

Cathal Mahon,1,{dagger} Bo Liang,2,{dagger} Irina Tikhanovich,1,{ddagger} Johanna R. Abend,3,{ddagger} Michael J. Imperiale,3 Heinz P. Nasheuer,1* and William R. Folk2*

National University of Ireland, Galway, Dept. of Biochemistry, Galway, Ireland,1 University of Missouri—Columbia, Dept. of Biochemistry, Columbia, Missouri 65212,2 University of Michigan, Dept. of Microbiology and Immunology, Ann Arbor, Michigan 481093

Received 11 February 2009/ Accepted 19 March 2009

BK virus (BKV) causes persistent and asymptomatic infections in most humans and is the etiologic agent of polyomavirus-associated nephropathy (PVAN) and other pathologies. Unfortunately, there are no animal models with which to study activation of BKV replication in the human kidney and the accompanying PVAN. Here we report studies of the restriction of BKV replication in murine cells and extracts and the cause(s) of this restriction. Upon infection of murine cells, BKV expressed large T antigen (TAg), but viral DNA replication and progeny were not detected. Transfection of murine cells with BKV TAg expression vectors also caused TAg expression without accompanying DNA replication. Analysis of the replication of DNAs containing chimeric BKV and murine polyomavirus origins revealed the importance of BKV core origin sequences and TAg for DNA replication. A sensitive assay was developed with purified BKV TAg that supported TAg-dependent BKV DNA replication with human but not with murine cell extracts. Addition of human replication proteins, DNA polymerase {alpha}-primase, replication protein A, or topoisomerase I to the murine extracts with BKV TAg did not rescue viral DNA replication. Notably, addition of murine extracts to human extracts inhibited BKV TAg-dependent DNA replication at a step prior to or during unwinding of the viral origin. These findings and differences in replication specificity between BKV TAg and the TAgs of simian virus 40 (SV40) and JC virus (JCV) and their respective origins implicate features of the BKV TAg and origin distinct from SV40 and JCV in restriction of BKV replication in murine cells.

* Corresponding author. Mailing address for Heinz P. Nasheuer: National University of Ireland, Galway, Dept. of Biochemistry, Galway, Ireland. Phone: 35391492430. Fax: 35391495504. E-mail: h.nasheuer{at}nuigalway.ie. Mailing address for William R. Folk: University of Missouri—Columbia, Dept. of Biochemistry, Columbia, MO 65212. Phone: (573) 884-2921. Fax: (573) 884-4597. E-mail: folkw{at}missouri.edu

{triangledown} Published ahead of print on 18 March 2009.

{dagger} These authors contributed equally.

{ddagger} These authors contributed equally.

Journal of Virology, June 2009, p. 5708-5717, Vol. 83, No. 11
0022-538X/09/$08.00+0     doi:10.1128/JVI.00300-09
Copyright © 2009, American Society for Microbiology. All Rights Reserved.





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