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Journal of Virology, July 2007, p. 7164-7170, Vol. 81, No. 13
0022-538X/07/$08.00+0     doi:10.1128/JVI.00112-07
Copyright © 2007, American Society for Microbiology. All Rights Reserved.

Marek's Disease Virus Type 2 (MDV-2)-Encoded MicroRNAs Show No Sequence Conservation with Those Encoded by MDV-1

Avian Oncogenic Virus Group, Institute for Animal Health, Compton, Berkshire RG20 7NN, United Kingdom,¹ LRF Molecular Haematology Unit, Nuffield Department of Clinical Laboratory Sciences, University of Oxford, Oxford OX3 9DU, United Kingdom,² Division of Bioinformatics, Biozentrum, University of Basel, Klingelbergstr 50-70, CH-4056 Basel, Switzerland³

Received 17 January 2007/ Accepted 18 April 2007

MicroRNAs (miRNAs) are increasingly being recognized as major regulators of gene expression in many organisms, including viruses. Among viruses, members of the family Herpesviridae account for the majority of the currently known virus-encoded miRNAs. The highly oncogenic Marek's disease virus type 1 (MDV-1), an avian herpesvirus, has recently been shown to encode eight miRNAs clustered in the MEQ and LAT regions of the viral genome. The genus Mardivirus, to which MDV-1 belongs, also includes the nononcogenic but antigenically related MDV-2. As MDV-1 and MDV-2 are evolutionarily very close, we sought to determine if MDV-2 also encodes miRNAs. For this, we cloned, sequenced, and analyzed a library of small RNAs from the lymphoblastoid cell line MSB-1, previously shown to be coinfected with both MDV-1 and MDV-2. Among the 5,099 small RNA sequences determined from the library, we identified 17 novel MDV-2-specific miRNAs. Out of these, 16 were clustered in a 4.2-kb long repeat region that encodes R-LORF2 to R-LORF5. The single miRNA outside the cluster was located in the short repeat region, within the C-terminal region of the ICP4 homolog. The expression of these miRNAs in MSB-1 cells and infected chicken embryo fibroblasts was further confirmed by Northern blotting analysis. The identification of miRNA clusters within the repeat regions of MDV-2 demonstrates conservation of the relative genomic positions of miRNA clusters in MDV-1 and MDV-2, despite the lack of sequence homology among the miRNAs of the two viruses. The identification of these novel miRNAs adds to the growing list of virus-encoded miRNAs.

Published ahead of print on 25 April 2007.

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