This Article Full Text Full Text (PDF) Other Versions of this Article: JVI.01284-06v1 80/23/11743    most recent Alert me when this article is cited Alert me if a correction is posted Services Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Reprints and Permissions Copyright Information Books from ASM Press MicrobeWorld Citing Articles Citing Articles via Google Scholar Google Scholar Articles by Bjarnadottir, H. Articles by Jonsson, J. J. Search for Related Content PubMed PubMed Citation Articles by Bjarnadottir, H. Articles by Jonsson, J. J.

 Previous Article  |  Next Article 

Journal of Virology, December 2006, p. 11743-11755, Vol. 80, No. 23
0022-538X/06/$08.00+0     doi:10.1128/JVI.01284-06
Copyright © 2006, American Society for Microbiology. All Rights Reserved.

Encapsidation Determinants Located Downstream of the Major Splice Donor in the Maedi-Visna Virus Leader Region

Department of Biochemistry and Molecular Biology, University of Iceland, Faculty of Medicine, Reykjavik, Iceland,¹ Department of Genetics and Molecular Medicine, Landspitali-University Hospital, Reykjavik, Iceland²

Received 19 June 2006/ Accepted 1 September 2006

We investigated the role of the 5'-untranslated region between the primer binding site and the gag initiation codon in ovine lentivirus maedi-visna virus (MVV) genomic RNA encapsidation. We identified five computer-predicted stem-loops, three of which were highly conserved in primary sequence and structure. One stable 83-nucleotide (nt) stem-loop (SL4) was not conserved in the primary sequence, but phylogenetic analysis revealed several base pair covariations. The deletion of individual stem-loops did not markedly affect the relative encapsidation efficiency (REE). Only one mutant, carrying a disruption of a 31-nt stem-loop (SL5), had 58% REE in fetal ovine synovial (FOS) cells. A 168-nt deletion (3MSD) downstream of the major splice donor (MSD) which removed three stem-loops, including SL5, resulted in 24% and 20% REE in FOS and 293T cells, respectively. A 100-nt deletion (5MSD) upstream of the MSD resulted in 15-fold lower cellular genomic RNA levels than the wild-type levels in 293T cells. The 5MSD mutant and a double mutant (DM) (5MSD and 3MSD) did not express detectable levels of virion proteins in 293T cells. In contrast, the region deleted in 5MSD was dispensable in FOS cells, and the DM had the same REE as the 3MSD virus. Thus, the region upstream of the MSD contains sequences critical for RNA and protein expression in a cell type-specific fashion. Our results indicate that MVV encapsidation determinants are located downstream of the MSD. These results provide comparative insight into lentiviral encapsidation and can be utilized in the design of MVV-based gene transfer vectors.

Published ahead of print on 13 September 2006.