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Journal of Virology, November 2007, p. 12572-12581, Vol. 81, No. 22
0022-538X/07/$08.00+0     doi:10.1128/JVI.00351-07
Copyright © 2007, American Society for Microbiology. All Rights Reserved.

Promoter Complex in the Central Part of the Mouse Mammary Tumor Virus Long Terminal Repeat{triangledown}

Francoise Rouault,1,2 Shiva Badihi Nejad Asl,2,3 Stefanie Rungaldier,2,3 Eva Fuchs,2,3 Brian Salmons,1 and Walter H. Günzburg2,3*

Austrianova Biomanufacturing AG,1 Christian Doppler Laboratory for Gene Therapeutic Vector Development,2 Research Institute for Virology and Biomedicine, Veterinärplatz 1, A-1210 Vienna, Austria3

Received 17 February 2007/ Accepted 26 July 2007

Unique among the retroviruses, mouse mammary tumor virus (MMTV) carries, in addition to the usual long terminal repeat (LTR) promoter, another promoter, P2, which is located in the central part of the proviral U3 sequence, within the LTR open reading frame (ORF). Using an in vitro reporter system based on a sensitive luciferase expression assay, we investigated the regulation of the P2 promoter in the context of the Mtv-2 and Mtv-8 genomes. Irrespective of the genomic source, the activity of the P2 promoter is regulated by a downstream-located enhancer and an upstream-located negative regulatory element (NRE), the activity of which overrides the activator. During this study, we unexpectedly detected another independent neighboring promoter that we called P3. The novel P3 promoter does not seem to be controlled by any NRE but is influenced by the same enhancer that modulates the P2 promoter. The respective transcription starts of the two promoters located in this tight cluster are only 61 bases apart. The transcripts originating from this promoter complex carry the same first intron, which is bound by canonical splice donor and splice acceptor sites located in the LTR. One novel doubly spliced transcript carrying a 459-nucleotide-long ORF was detected in several MMTV-carrying murine cells and could be successfully expressed in murine cells as a His-tagged fusion product. The novel viral protein, the function of which remains to be elucidated, has an apparent molecular mass of 20 kDa.

* Corresponding author. Mailing address: Research Institute for Virology and Biomedicine, Veterinärplatz 1, A-1210 Vienna, Austria. Phone: (43) 25077-2301. Fax: (43) 25077-2390. E-mail: walter.guenzburg{at}vu-wien.ac.at

{triangledown} Published ahead of print on 8 August 2007.

Journal of Virology, November 2007, p. 12572-12581, Vol. 81, No. 22
0022-538X/07/$08.00+0     doi:10.1128/JVI.00351-07
Copyright © 2007, American Society for Microbiology. All Rights Reserved.





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