Based on the concept of anti-sense RNA expression, we plan to establish and characterize systems which exhibit resistance to animal virus infection. 1.) Using bovine papilloma virus vectors, we will further develop permanent cell lines (C127) which express high levels of influenza virus genes in the anti-sense. These cells will be examined for their ability prevent replication of influenza viruses. In addition, we will establish cell lines which express anti-sense RNAs from tandem copies of full length and/or of domains of influenza virus genes. Such cell lines may show an enhanced resistance to influenza virus infection because of gene dosage effect of the anti-sense RNAs. Cell lines permanently expressing RNAs analogous to defective interfering (DI) RNAs of influenza virus will also be examined for their ability to interfere with viral replication. 2.) We will test alternative cell, vector and promoter systems such as 293 cells and EBV-based vectors containing CMV promoters to express virus-specific anti-sense RNAs in order to establish optimal resistance conditions and to allow infection with different influenza viruses. 3.) We will study the mechanism of action by which anti-sense RNA expression inhibits influenza virus replication. 4.) We will attempt to generate transgenic mice made resistant to influenza virus infection via anti-sense RNA expression. 5.) To determine the usefulness of anti-sense RNA expression as a general antiviral approach, we will try to establish a second viral anti-sense RNA expression system by introducing a vesicular stomatitis virus (VSV) gene into tissue culture cells. 6.) We will attempt to develop ribo- and/or deoxyribo- oligonucleotides which are complementary to influenza virus- specific sequences, for use as antiviral agents.
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