Influenza viral infection represents a serious public health problem that causes contagious respiratory disease, which is most effectively prevented through vaccination to reduce transmission and future
infection. The nonstructural (NS) gene of influenza A virus encodes an
mRNA transcript that is alternatively spliced to express two
viral proteins, the nonstructural
protein 1 (NS1) and the nuclear export
protein (NEP). The importance of the NS gene of influenza A virus for viral replication and virulence has been well described and represents an attractive target to generate live attenuated influenza viruses with
vaccine potential. Considering that most
amino acids can be synthesized from several synonymous
codons, this study employed the use of misrepresented mammalian
codons (
codon deoptimization) for the de novo synthesis of a viral NS
RNA segment based on
influenza A/Puerto Rico/8/1934 (H1N1) (PR8) virus. We generated three different recombinant
influenza PR8 viruses containing
codon-deoptimized synonymous mutations in coding regions comprising the entire NS gene or the
mRNA corresponding to the individual
viral protein NS1 or NEP, without modifying the respective splicing and packaging signals of the viral segment. The fitness of these synthetic viruses was attenuated in vivo, while they retained immunogenicity, conferring both homologous and heterologous protection against influenza A virus challenges. These results indicate that influenza viruses can be effectively attenuated by synonymous
codon deoptimization of the NS gene and open the possibility of their use as a safe
vaccine to prevent
infections with these important human pathogens.
IMPORTANCE: Vaccination serves as the best therapeutic option to protect humans against
influenza viral infections. However, the efficacy of current
influenza vaccines is suboptimal, and novel approaches are necessary for the prevention of disease cause by this important human respiratory pathogen. The nonstructural (NS) gene of influenza virus encodes both the multifunctional nonstructural
protein 1 (NS1), essential for innate immune evasion, and the nuclear export
protein (NEP), required for the nuclear export of viral
ribonucleoproteins and for timing of the virus life cycle. Here, we have generated a recombinant
influenza A/Puerto Rico/8/1934 (H1N1) (PR8) virus containing a
codon-deoptimized NS segment that is attenuated in vivo yet retains immunogenicity and protection efficacy against homologous and heterologous influenza virus challenges. These results open the exciting possibility of using this NS
codon deoptimization methodology alone or in combination with other approaches for the future development of
vaccine candidates to prevent
influenza viral infections.