SSRC Annual Report 2012

The SSRC was established in 1950 and enjoys a long and storied history of research accomplishments. The impact of our research has had on the state, the nation and the world is evident in the success of our organization, which has ultimately been driven by the desire and dedication of our employees. Although an organization\u27s annual report provides a venue to applaud the hard work and dedication of its employees for its success during the previous year, this particular report also celebrates our accomplished past and acknowledges significant historical milestones celebrated this year

siRNA against the G gene of human metapneumovirus

Background: Human metapneumovirus (hMPV) is a significant viral respiratory pathogen of infants and children, the elderly and immunocompromised individuals. Disease associated with hMPV infection resembles that of human respiratory syncytial virus (RSV) and includes bronchiolitis and pneumonia. The glycosylated G attachment protein of hMPV is required for viral entry in vivo and has also been identified as an inhibitor of innate immune responses. Findings: We designed and validated two siRNA molecules against the G gene using A549 cells and demonstrated consistent 88-92% knock-down for one siRNA molecule, which was used in subsequent experiments. Significant reduction of G mRNA in A549 cells infected with hMPV did not result in a reduction in viral growth, nor did it significantly increase the production of type I interferon (α/β) in response to infection. However, there was a moderate increase in IFN-β mRNA expression in response to infection in siG-transfected cells compared to untransfected and si-mismatch-transfected cells. Expression of G by recombinant adenovirus did not affect type I IFN expression. Conclusion: G has been previously described as a type I interferon antagonist, although our findings suggest it may not be a significant antagonist

SSRC Annual Report 2013

The SSRC was established in 1950 and enjoys a long and storied history of research accomplishments. The impact of our research has had on the state, the nation and the world is evident in the success of our organization, which has ultimately been driven by the desire and dedication of our employees. Although an organization\u27s annual report provides a venue to applaud the hard work and dedication of its employees for its success during the previous year, this particular report also celebrates our accomplished past and acknowledges significant historical milestones celebrated this year

Mutation of the elongin C binding domain of human respiratory syncytial virus non-structural protein 1 (NS1) results in degradation of NS1 and attenuation of the virus

Background: Human respiratory syncytial virus (RSV) is an important cause of lower respiratory tract disease in the paediatic population, immunocompromised individuals and the elderly worldwide. However, despite global efforts over the past several decades there are no commercially available vaccines. RSV encodes 2 non-structural proteins, NS1 and NS2, that are type I interferon antagonists. RSV restricts type I interferon signaling and the expression of antiviral genes by degrading STAT2. It has been proposed that NS1 binds to elongin C to form a ubiquitin ligase (E3) complex that targets STAT2 for ubiquitination and proteosomal degradation. Results: Here, we have engineered a live recombinant RSV in which the 3 consensus amino acids of the NS1 elongin C binding domain have been replaced with alanine (NS1F-ELCmut). Mutation of this region of NS1 resulted in attenuation of RSV replication in A549 cells to levels similar to that observed when the NS1 gene is completely deleted (NS1). This mutation also resulted in moderate attenuation in Vero cells. Attenuation was correlated to intracellular degradation of the mutated NS1 protein. Time course analysis showed that mutant NS1 protein accumulated in cytoplasmic bodies that contained the lysosomal marker LAMP1. However lack of cleavage of LC3 suggested that autophagy was not involved. Induction of IFN- mRNA expression also was observed in association with the degradation of NS1 protein and attenuation of viral growth. Conclusions: These results indicate that the elongin C binding region of NS1 is crucial for survival of the protein and that disruption of this region results in the degradation of NS1 and restriction of RSV replication