| Summary: | 碩士 === 長庚大學 === 基礎醫學研究所 === 87 === It has been shown that influenza A virus M1 mRNA has two alternative 5' splice sites: a distal 5' splice site producing M3 mRNA that has the coding potential for 9 amino acid and a proximal 5' splice site producing M2 mRNA that encodes the essential M2 ion-channel protein. In this study, we demonstrate that the laboratory widely used strain A/WSN/33, but not A/Udorn/72 possessed another novel 5' splice site producing a transcript with the coding potential for 54 amino acid. We nominated this novel transcript as M4 mRNA. M4 mRNA was detected in A/WSN/33-infected cells derived from different species, and the uninfected cells transfected with M1 cDNA, only the M3 mRNA 5' splice site was used. M2 mRNA was detected after blocking M3 mRNA 5' splice site by mutation. If M4 protein can be translated, it needs to be study further. Sequence comparison of M1 mRNA in both A/WSN/33 and A/Udorn/72 at position 143 to 157 reveals that the novel 5' splice site generated in WSN was due to one nucleotide difference at position 147. Several strains of influenza A virus other than WSN also possess the potential M4 5' splice site by sequence analysis from the files of GenBank and recently isolates in Taiwan. M4 5' splice site seem to be a genetic maker from phylogenetic analysis. To study the biological function of the novel 5' splice site, we construct transfectant viruses by reverse genetics. In addition, we used RNase protection assay to quantitate the efficiency of alternative splicing in M1 mRNA of A/WSN/33.
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