German researchers have identified hundreds of host cell genes that affect influenza A virus replication. The work by Alexander Karlas and colleagues and published online in the journal Nature could help identify new drug targets which could be useful against a broad range of influenza viruses.
Influenza A viruses are a global public health threat that cause seasonal flu epidemics and periodic pandemics, the most recent major outbreak being the infamous swine flu pandemic that originated in Mexico in 2009. The influenza virus has a high mutation rate, which means that drugs and vaccines can become ineffective rapidly during a flu outbreak. The influenza virus can only replicate inside living cells and so novel drugs that target host cell functions required for virus replication are an attractive research area.
Karlas et al. conducted a genome-wide RNA interference screen in conjunction with a luciferase reporter assay to identify host cells factor that are important for influenza virus replication in human cells. First, they transfected human epithelial lung cells with 62,000 siRNAs against ~17,000 annotated and ~6,000 predicted human genes and 48 h later they infected the same cells with influenza A H1N1 virus and used immunofluorescence microscopy to check the virus infection rates. Secondly, they transferred the virus supernatants from the lung cells onto human embryonic kidney cells which contained an influenza-virus-specific luciferase reporter that bioluminesces in the presence of the virus to further quantify the virus infection and replication rates.
The scientists identified 287 human genes which appeared to be important in influenza virus replication, including the nuclear export factor genes NXF1 and XPO1, which have already been shown to be important for flu virus replication, and several genes which are connected with pre-mRNA splicing. They confirmed that siRNAs against ~59% (168 out of 287) of these genes decreased the number of endemic H1N1 and the 2009 pandemic H1N1 influenza viruses in human cells by at least five times. Interestingly, a subset of the same siRNAs also decreased replication of the highly pathogenic avian H5N1 influenza A strain. Furthermore, they found that a small molecule inhibitor of CDC-like kinase 1 reduced influenza virus replication by two orders of magnitude, because of impaired splicing of viral M2 messenger RNA. Finally, in vivo mouse studies confirmed the importance of the cell cycle regulator p27 in virus replication, p27-/- knockout mice were infected with H1N1 virus and 2 days later the researchers observed that the viral load within the lungs of the mice was significantly reduced.
This study “provided new and comprehensive information on host cell determinants of replication, and uncovered potential targets for novel antiviral strategies…against a broad spectrum of influenza viruses” write the authors and presents more information on the interaction between viruses and the human host.
Karlas, A., Machuy, N., Shin, Y., Pleissner, K., Artarini, A., Heuer, D., Becker, D., Khalil, H., Ogilvie, L., Hess, S., Mäurer, A., Müller, E., Wolff, T., Rudel, T., & Meyer, T. (2010). Genome-wide RNAi screen identifies human host factors crucial for influenza virus replication Nature DOI: 10.1038/nature08760