Virology 410:385C394 [PubMed] [Google Scholar] 10

Virology 410:385C394 [PubMed] [Google Scholar] 10. PCBP1 and PCBP2 were found to bind the viral 5 untranslated region (5UTR). Small interfering RNA (siRNA)-mediated silencing of PCBP1 and PCBP2 in cells resulted in significantly reduced PRRSV genome replication and transcription without adverse effect on initial polyprotein synthesis. Overall, the results presented here point toward an important role for PCBP1 and PCBP2 in regulating PRRSV RNA synthesis. INTRODUCTION Porcine reproductive and respiratory syndrome virus (PRRSV) is considered the most significant swine pathogen worldwide. Viral infection leads to late-term reproductive failure in pregnant sows and respiratory distress in young piglets. The most recent study (in 2011) of annual economic losses due to PRRSV infections to the U.S. swine industry alone has suggested the loss to be over $660 million (29). PRRSV is a member of the family and families, form the order (4). PRRSV contains a plus-sense [(+)-sense] single-stranded RNA genome of 15 kb in length. The RNA genome is polyadenylated at the 3 end and possesses a 5 cap structure (28). The nonstructural proteins (nsp’s) are encoded in the 5-proximal two-thirds of the genomic RNA and are synthesized as polyproteins from two open reading frames (ORFs): ORF1a and ORF1b. Upon translation, the two polyproteins are cotranslationally and posttranslationally processed by four virus-encoded proteases, i.e., nsp1, nsp1, nsp2, and nsp4, to produce 14 different nsp’s (39, 48). Some of these nsp’s are assembled to form the viral replicase complex and, along with host cell components, carry out Morin hydrate and regulate viral RNA replication, subgenomic (sg) mRNA transcription, and translation. The PRRSV structural proteins encoded by the 3-proximal one-third of the genomic RNA are translated from a set of six nested sg CD14 mRNAs generated through a process of discontinuous RNA transcription (33, Morin hydrate 37). The viral genomic RNA acts as a template for protein synthesis, genome replication (synthesis of genome-length RNAs), and transcription (synthesis of sg-length RNAs) as well as for packaging into virions. These processes must be regulated by viral and host cell factors for efficient replication of the virus. Identification of such factors is challenging, as it requires uncoupling of the tightly linked processes of genome replication and transcription. Studies have identified EAV nsp1 and nsp10 as the two proteins essential for sg mRNA synthesis (45, 51). While EAV nsp1 is synthesized as a single protein due to the lack of a PCP active-site residue, PRRSV nsp1 is autoproteolytically processed to produce nsp1 and nsp1 proteins (10). Mutations of the protease active site of PRRSV nsp1 led to a replication-competent but sg mRNA transcription-incompetent virus (22). The same study also described that mutation in the nsp1 protease active site results in a total loss of RNA replication, presumably because of the absence of cleavage of the nsp1-nsp2 junction. More recently, it was shown that all the three subdomains of EAV nsp1 (zinc finger, PCP, and Morin hydrate PCP) are important for transcription (31). Importantly, it was shown that the PCP domain’s involvement in sg mRNA transcription is separate from its protease function. Furthermore, nsp1 was found to regulate the relative amounts of viral mRNA by controlling the accumulation of full-length and sg-length minus (?)-strand templates (31). Besides its role in viral sg mRNA transcription, EAV nsp1 is also involved in polyprotein processing and virion biogenesisunderscoring its multifunctional role in the life cycle of the virus (45). Similar roles can also be predicted for the PRRSV orthologs, nsp1 and nsp1. Besides their involvement in viral gene expression, both subunits of nsp1 are known inhibitors of cellular innate immune response (3, 7, 19, 34, 42). While the.