A human cytomegalovirus-encoded microRNA regulates expression of multiple viral genes involved in replication.
Although multiple studies have documented the expression of over 70 novel virus-encoded microRNAs (miRNAs), the targets and functions of most of these regulatory RNA species are unknown. In this study a comparative bioinformatics approach was employed to identify potential human cytomegalovirus (HCMV) mRNA targets of the virus-encoded miRNA miR-UL112-1. Bioinformatics analysis of the known HCMV mRNA 3' untranslated regions (UTRs) revealed 14 potential viral transcripts that were predicted to contain functional target sites for miR-UL112-1. The potential target sites were screened using luciferase reporters that contain the HCMV 3'UTRs in co-transfection assays with miR-UL112-1. Three of the 14 HCMV miRNA targets were validated, including the major immediate early gene encoding IE72 (UL123, IE1), UL112/113, and UL120/121. Further analysis of IE72 regulation by miR-UL112-1 with clones encoding the complete major immediate early region revealed that the IE72 3'UTR target site is necessary and sufficient to direct miR-UL112-1-specific inhibition of expression in transfected cells. In addition, miR-UL112-1 regulation is mediated through translational inhibition rather than RNA degradation. Premature expression of miR-UL112-1 during HCMV infection resulted in a significant decrease in genomic viral DNA levels, suggesting a functional role for miR-UL112-1 in regulating the expression of genes involved in viral replication. This study demonstrates the ability of a viral miRNA to regulate multiple viral genes.
Grey
Finn
F
Vaccine and Gene Therapy Institute, Oregon Health Sciences University, Portland, Oregon, United States of America. greyf@ohsu.edu
Meyers
Heather
H
White
Elizabeth A
EA
Spector
Deborah H
DH
Nelson
Jay
J
D001483:Base Sequence; D015152:Blotting, Northern; D015153:Blotting, Western; D019295:Computational Biology; D003587:Cytomegalovirus / Q000235:genetics*; D003586:Cytomegalovirus Infections / Q000235:genetics*; D004261:DNA Replication; D015870:Gene Expression; D005786:Gene Expression Regulation; D005814:Genes, Viral; D006801:Humans; D035683:MicroRNAs / Q000235:genetics*; D008969:Molecular Sequence Data; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014162:Transfection; D020506:Untranslated Regions; D014779:Virus Replication / Q000235:genetics*
D035683:MicroRNAs; D020506:Untranslated Regions