M. LINIGER¹, T.K.H. SCHEEL², J.M. LUNA³, G. AURAY¹, M. GERBER¹, I. KELLER⁴, R. BRUGGMANN⁴, N. RUGGLI¹, C.M. RICE<sup>3

1</sup>Department of Virology, Institute of Virology and Immunology IVI, Mittelhäusern, Switzerland, ²Copenhagen Hepatitis C Program, Department of Infectious Disease and Clinical Research Centre, Copenhagen University Hospital, Hvidovre, Denmark, ³Laboratory of Virology and Infectious Disease, Center for the Study of Hepatitis C, The Rockefeller University, New York, NY, USA, ⁴Interfaculty Bioinformatics Unit and Swiss Institute of Bioinformatics, University of Bern, Bern, Switzerland

Crosslinking immunoprecipitation of Argonaute proteins combined with high throughput sequencing was used in the context of viral infections to search for microRNA (miRNA) interactions with viral genomic RNA. This revealed specific miR-17 binding at the 3’ non-translated region of bovine viral diarrhoea virus (BVDV). The miR-17 binding site of BVDV is conserved in the related pestivirus classical swine fever virus (CSFV). Therefore, we studied the functional relevance of miR-17 binding by CSFV. To this end, CSFV with a defective miR-17 seed site was generated (CSFV-p3p4). Replication of the parent and mutant CSFV were analysed in the presence of locked antisense oligonucleotides (tinyLNA17) for miR-17 inhibition and of artificial miR-17p3p4 for seed site complementation, respectively. TinyLNA17 repressed the propagation of CSFV, suggesting that CSFV depends on miR-17 for efficient replication. In addition, replication of the CSFV-p3p4 mutant was strongly impaired. Complementation with artificial miR-17p3p4 restored the growth characteristics of CSFV-p3p4 completely. Luciferase-based miR-17 reporter assays suggested a virus-mediated sequestration of miR-17. Accordingly, differential transcriptome analysis of CSFV versus mock-infected cells showed functional de-repression of cellular miR-17 target mRNAs, thereby altering the host transcriptome. Taken together, these data demonstrate that miR-17 is a critical host factor for replication of CSFV.