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 acids and a proximal 5′ splice site producing M2 mRNA that encodes the essential M2 ion-channel protein. In this study, we demonstrated 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 acids. We nominated this novel transcript as M4 mRNA. M4 mRNA was detected in A/WSN/33-infected cells derived from different species. Sequence comparison of M1 mRNA in both A/WSN/33 and Udorn/72 at position 143 to 151 reveals that this novel 5′ splice site generated in WSN was due to one nucleotide difference at position 147. Several strains of influenza A viruses other than WSN also possess the potential M4 5′ splice site by sequence analysis from the files of GenBank.
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