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Molecular and Cellular Biology, December 2006, p. 8814-8825, Vol. 26, No. 23
0270-7306/06/$08.00+0     doi:10.1128/MCB.00636-06
Copyright © 2006, American Society for Microbiology. All Rights Reserved.

Transcriptional Regulation Is Affected by Subnuclear Targeting of Reporter Plasmids to PML Nuclear Bodies{triangledown} ,{dagger}

Gregory J. Block,{ddagger} Christopher H. Eskiw,§ Graham Dellaire, and David P. Bazett-Jones*

Research Institute, The Hospital for Sick Children, Toronto, Canada

Received 12 April 2006/ Returned for modification 26 June 2006/ Accepted 30 August 2006

Whereas the PML protein has been reported to have both transcriptional coactivator and corepressor potential, the contribution of the PML nuclear body (PML NB) itself to transcriptional regulation is not well understood. Here we demonstrate that plasmid DNA artificially tethered to PML or the PML NB-targeting domain of Sp100 is preferentially localized to PML NBs. Using the tethering technique, we targeted a simian virus 40 promoter-driven luciferase reporter plasmid to PML NBs, resulting in the repression of the transgene transcriptional activity. Conversely, the tethering of a cytomegalovirus promoter-containing reporter plasmid resulted in activation. Targeting a minimal eukaryotic promoter did not affect its activity. The expression of targeted promoters could be modulated by altering the cellular concentration of PML NB components, including Sp100 and isoforms of the PML protein. Finally, we demonstrate that ICP0, the promiscuous herpes simplex virus transactivator, increases the level of transcriptional activation of plasmid DNA tethered to the PML NB. We conclude that when PML NB components are artificially tethered to reporter plasmids, the PML NB contributes to the regulation of the tethered DNA in a promoter-dependent manner. Our findings demonstrate that transient transcription assays are sensitive to the subnuclear localization of the transgene plasmid.

* Corresponding author. Mailing address: The Hospital for Sick Children, 555 University Avenue, Toronto, Ontario M5G 1X8, Canada. Phone: (416) 813-2181. Fax: (416) 813-2235. E-mail: dbjones{at}sickkids.ca.

{triangledown} Published ahead of print on 11 September 2006.

{dagger} Supplemental material for this article may be found at http://mcb.asm.org/.

§ Present address: Sir William Dunn School of Pathology, Oxford University, Oxford, United Kingdom.

{ddagger} Present address: Department of Gene Therapy, Tulane University, New Orleans, La.

Molecular and Cellular Biology, December 2006, p. 8814-8825, Vol. 26, No. 23
0270-7306/06/$08.00+0     doi:10.1128/MCB.00636-06
Copyright © 2006, American Society for Microbiology. All Rights Reserved.



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