Virus Research 190 (2014) 69–74 Contents lists available at ScienceDirect Virus Research j ourna l h o mepa ge: www.elsevier.com/locate/virusres Transcriptional regulation of gene expression of infectious salmon anaemia virus segment 7 Rimatulhana B. Ramly, Christel M. Olsen, Stine Braaen, Elisabeth F. Hansen, Espen Rimstad ∗ Department of Food Safety and Infection Biology, Faculty of Veterinary Medicine and Biosciences, Norwegian University of Life Sciences (NMBU), P.O. Box 8146 Dep, 0033 Oslo, Norway a r t i c l e i n f o Article history: Received 12 February 2014 Received in revised form 22 May 2014 Accepted 7 July 2014 Available online 16 July 2014 Keywords: Infectious salmon anaemia virus Gene expression of NS and NEP a b s t r a c t The nuclear replication and gene splicing of orthomyxoviruses are unique among RNA viruses. Segment 7 of infectious salmon anaemia virus (ISAV) is the only segment that undergoes splicing. Two proteins are encoded by this segment, the non-structural antagonist (ISAV-NS) of the innate immune response that is translated from the unspliced collinear transcript, and a nuclear exporting protein (ISAV-NEP) that is translated from the spliced mRNA. Here we report the transcription profiles for these ISAV proteins. The appearance of the spliced ISAV-NEP mRNA was delayed and the relative amount was less but slowly accumulated to 20–30% to that of the collinear NS mRNA. In cells transfected with segment 7 the ratio between spliced and collinear mRNA was approximately 10%. A highly conserved, possible structured RNA, in the region of the 3 ′ splicing site of the segment is speculated as being important for the regulation of the efficiency of the splicing. © 2014 Elsevier B.V. All rights reserved. 1. Introduction Infectious salmon anaemia virus (ISAV) has a genome of eight single-stranded RNA molecules of negative polarity and is the type virus of the genus Isavirus in the family Orthomyxoviridae. It is the etiologic agent of infectious salmon anaemia (ISA) a disease that cause high mortality in aquaculture of Atlantic salmon (Salmo salar). The number of ISA outbreaks has been highly reduced due to improvements in hygienic and management measures. The tran- scription and replication of ISAV take place in the nucleus of the infected cell (Brinson et al., 2011; Sandvik et al., 2000), where the viral genomic-, complementary- and messenger RNAs are syn- thesized by the viral transcriptase complex. Like other orthomyx- oviruses ISAV mRNAs have capped, heterogeneous 5 ′ -ends and their synthesis is inhibited by -amanitin, a specific inhibitor of the cellular RNA polymerase II due to the need of capped host nuclear RNAs as primers for mRNA synthesis (Sandvik et al., 2000). The nuclear replication requires nuclear export of viral mRNAs, and nuclear import of the viral proteins of the ribonucleoproteins (RNP), i.e. the nucleoprotein (NP) and the viral transcriptase com- plex, as well as of matrix protein (M), and finally nuclear export of the RNP–M complex. This transport must be temporally and ∗ Corresponding author. Tel.: +47 22964766. E-mail address: espen.rimstad@nmbu.no (E. Rimstad). quantitatively well regulated and the nuclear exporting protein (NEP) has been shown to play a vital role in this matter for influenza virus (Boulo et al., 2007). An ISAV protein with properties compar- ative to those of NEP of influenza viruses is encoded by genomic segment 7 (Ramly et al., 2013), and this segment also encodes the non-structural (NS) protein of ISAV (Garcia-Rosado et al., 2008). ISAV-NS is encoded by a collinear mRNA transcript, whereas ISAV- NEP is encoded by the only known spliced mRNA of ISAV (Biering et al., 2002; Kibenge et al., 2007; Mjaaland et al., 1997; Ritchie et al., 2002). The organization of the ISAV genomic segment 7 resembles that of the corresponding segment of the influenza viruses, where the interferon (IFN) antagonist NS1 and NEP are transcribed by the collinear and spliced transcripts, respectively. This indicates a remarkably conservation of gene organization between the piscine ISAV and the avian/mammalian influenza viruses. The 34 kDa ISAV-NS and the 17.5 kDa -NEP share the 22 N- terminal amino acids (Biering et al., 2002; Clouthier et al., 2002; Ritchie et al., 2002). The smaller ISAV-NEP contains leucine-rich nuclear export signals, got trapped in the nucleus when CRM-1 dependent nuclear export was inhibited with Leptomycin B, and found to interact with both NP and M (Rimatulhana et al., 2013). The ISAV-NS has antagonistic effect on transcription of IFN-type I and IFN-type I induced proteins, but does not have the RNA binding properties or the nuclear localization signals of the NS1 proteins of influenza viruses and it localizes mainly perinucle- arly in the cytoplasm (Garcia-Rosado et al., 2008; McBeath et al., http://dx.doi.org/10.1016/j.virusres.2014.07.008 0168-1702/© 2014 Elsevier B.V. All rights reserved.