The human T-cell leukemia virus (HTLV-1) Rex protein is essential for the cytoplasmic accumulation of incompletely spliced transcripts that code for the viral structural proteins. In this study effects of Rex on the amounts of total, spliced and unspliced RNA from HTLV-1 were determined. In transfected fibroblasts Rex production resulted in reduced amounts of spliced RNA and increased quantities of unspliced RNA in the nucleus. However, the total amount of viral RNA was not affected and the stability of spliced transcripts was not changed, thus indicating that only the rate of splicing was reduced. Rex action also reduced splicing in immortalized human cord blood T-cells. However, the total amount of viral transcripts and the stability of unspliced RNA in these cells were also increased in the presence of Rex. This indicates that Rex also prevents the degradation of unspliced transcripts in T-cells. The changes in the relative amounts of spliced and unspliced RNA induced by Rex were observed not only in the cytoplasm but also in the nucleus. Thus Rex affects the nucleocytoplasmic transport, splicing and stability of HTLV-1 RNA in the nucleus. These observations may suggest that Rex directs the unspliced viral RNA to the cytoplasm via a nuclear compartment that is not accessible to splicing and degradation factors.

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The virulence of influenza A viruses depends on the activity of the viral RNA polymerase complex and viral regulatory phosphoproteins. We identified that the protein kinase C (PKC) inhibitor Gö 6976 had a post-entry anti-influenza viral effect, by using a polymerase activity-based reporter assay. This inhibitory effect was observed for influenza virus-infected cells as well as for cells transiently transfected with constructs for the RNA polymerase complex. Importantly, the in vitro analysis of viral protein phosphorylation identified PKCa as a kinase phosphorylating PB1 and NS1, but not PB2, PA or NP. Gö 6976 was able to block PKC-specific phosphorylation in vitro. Thus, our data suggest that PKC contributes to the phosphorylation of influenza PB1 and NS1 proteins which appears to be functionally relevant for both viral RNA polymerase activity and efficient viral replication.Influenza A viruses are pathogens that cause significant morbidity and mortality in humans and a variety of animal species (Palese & Shaw, 2007;Wright et al., 2007). Influenza viruses possess a segmented negative-sense RNA genome which is replicated in the nucleus of infected cells. The virulence of individual virus strains is based on a complex co-action of determinants, including the viral haemagglutinin (HA), the RNA-dependent RNA polymerase (subunits PB1, PB2 and PA) and the nonstructural protein (NS1) (Geiss et al., 2002;Tumpey et al., 2005Tumpey et al., , 2007. The RNA-dependent RNA polymerase complex is responsible for both transcription and genomic replication (Torreira et al., 2007;Palese & Shaw, 2007). PB1 represents the central catalytic subunit of the polymerase complex (Honda & Ishihama, 1997;Poch et al., 1989;Torreira et al., 2007), PB2 is responsible for recognition and binding of the cap structures of host mRNAs which are used for priming viral transcription (Fechter & Brownlee, 2005;Guilligay et al., 2008), while PA combines essential activities required for genome replication and protein processing (Fodor & Smith, 2004;Maier et al., 2008;Palese & Shaw, 2007). The viral nucleoprotein (NP) strictly associates with the RNA polymerase complex to form viral ribonucleoproteins (RNPs) (Noda et al., 2006;Portela & Digard, 2002), it stimulates RNA polymerase activity and mediates nuclear RNP translocation (Neumann et al., 1997;Bui et al., 2002;Portela & Digard, 2002). The viral regulatory protein NS1 provides a number of functions required for efficient replication and gene expression (Hale et al., 2008a). An interaction between NS1 and the viral transcription-replication complex was reported (Marion et al., 1997) but awaits a confirmation in functional aspects. NS1 and the RNP components including NP are phosphoproteins in various types and strains of influenza viruses (Sanz-Ezquerro et al., 1998;Bui et al., 2002;Arrese & Portela, 1996;Skorko et al., 1991;Marschall et al., 1999;Neumann et al., 1997;Privalsky & Penhoet, 1978, 1981. Several cellular protein kinases, including protein kinase C (PKC), casein kinase II, cyclin-dependen...

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