Elevation of CpG frequencies in influenza A genome attenuates pathogenicity but enhances host response to infection

Previously, we demonstrated that frequencies of CpG and UpA dinucleotides profoundly influence the replication ability of echovirus 7 (Tulloch et al., 2014). Here, we show that that influenza A virus (IAV) with maximised frequencies of these dinucleotides in segment 5 showed comparable attenuation in cell culture compared to unmodified virus and a permuted control (CDLR). Attenuation was also manifested in vivo, with 10-100 fold reduced viral loads in lungs of mice infected with 200PFU of CpG-high and UpA-high mutants. However, both induced powerful inflammatory cytokine and adaptive (T cell and neutralising antibody) responses disproportionate to their replication. CpG-high infected mice also showed markedly reduced clinical severity, minimal weight loss and reduced immmunopathology in lung, yet sterilising immunity to lethal dose WT challenge was achieved after low dose (20PFU) pre-immunisation with this mutant. Increasing CpG dinucleotide frequencies represents a generic and potentially highly effective method for generating safe, highly immunoreactive vaccines. DOI: http://dx.doi.org/10.7554/eLife.12735.001

[1]  J. Peiris,et al.  Generation of Live Attenuated Influenza Virus by Using Codon Usage Bias , 2015, Journal of Virology.

[2]  P. Simmonds,et al.  Attenuation of dengue (and other RNA viruses) with codon pair recoding can be explained by increased CpG/UpA dinucleotide frequencies , 2015, Proceedings of the National Academy of Sciences.

[3]  Charles B. Ward,et al.  Large-scale recoding of an arbovirus genome to rebalance its insect versus mammalian preference , 2015, Proceedings of the National Academy of Sciences.

[4]  P. Simmonds,et al.  RNA virus attenuation by codon pair deoptimisation is an artefact of increases in CpG/UpA dinucleotide frequencies , 2014, eLife.

[5]  Steven F. Baker,et al.  Influenza A Virus Attenuation by Codon Deoptimization of the NS Gene for Vaccine Development , 2014, Journal of Virology.

[6]  Judy Qiu,et al.  Total Synthesis of a Functional Designer Eukaryotic Chromosome , 2014, Science.

[7]  T. Opriessnig,et al.  Computer-aided codon-pairs deoptimization of the major envelope GP5 gene attenuates porcine reproductive and respiratory syndrome virus. , 2014, Virology.

[8]  P. Simmonds,et al.  The influence of CpG and UpA dinucleotide frequencies on RNA virus replication and characterization of the innate cellular pathways underlying virus attenuation and enhanced replication , 2014, Nucleic acids research.

[9]  Jonathan J Deeks,et al.  Cellular immune correlates of protection against symptomatic pandemic influenza , 2013, Nature Medicine.

[10]  B. Clotet,et al.  Changes in codon-pair bias of human immunodeficiency virus type 1 have profound effects on virus replication in cell culture , 2013, Retrovirology.

[11]  B. Futcher,et al.  Deliberate reduction of hemagglutinin and neuraminidase expression of influenza virus leads to an ultraprotective live vaccine in mice , 2013, Proceedings of the National Academy of Sciences.

[12]  Walter N. Moss,et al.  Influenza A Virus Coding Regions Exhibit Host-Specific Global Ordered RNA Structure , 2012, PloS one.

[13]  P. Simmonds.  SSE: a nucleotide and amino acid sequence analysis platform , 2012, BMC Research Notes.

[14]  Walter N. Moss,et al.  Identification of potential conserved RNA secondary structure throughout influenza A coding regions. , 2011, RNA.

[15]  Y. Pilpel,et al.  Determinants of translation efficiency and accuracy , 2011, Molecular systems biology.

[16]  Vishvanath Nene,et al.  Faculty Opinions recommendation of Live attenuated influenza virus vaccines by computer-aided rational design. , 2010 .

[17]  Thomas H Segall-Shapiro,et al.  Creation of a Bacterial Cell Controlled by a Chemically Synthesized Genome , 2010, Science.

[18]  Julia R Gog,et al.  Genome packaging in influenza A virus. , 2010, The Journal of general virology.

[19]  Jie Ma,et al.  Computational identification of rare codons of Escherichia coli based on codon pairs preference , 2010, BMC Bioinformatics.

[20]  E. Wimmer,et al.  Synthetic viruses: a new opportunity to understand and prevent viral disease , 2009, Nature Biotechnology.

[21]  A. Vincent,et al.  Genetic Inactivation of Poliovirus Infectivity by Increasing the Frequencies of CpG and UpA Dinucleotides within and across Synonymous Capsid Region Codons , 2009, Journal of Virology.

[22]  Jean A. Roayaei,et al.  Live attenuated or inactivated influenza vaccines and medical encounters for respiratory illnesses among US military personnel. , 2009, JAMA.

[23]  Maido Remm,et al.  Preferred and avoided codon pairs in three domains of life , 2008, BMC Genomics.

[24]  J. Gog,et al.  Mutational Analysis of cis-Acting RNA Signals in Segment 7 of Influenza A Virus , 2008, Journal of Virology.

[25]  S. Garcia-Vallvé,et al.  CAIcal: A combined set of tools to assess codon usage adaptation , 2008, Biology Direct.

[26]  J. R. Coleman,et al.  Virus Attenuation by Genome-Scale Changes in Codon Pair Bias , 2008, Science.

[27]  A. Osterhaus,et al.  Influenza virus-specific cytotoxic T lymphocytes: a correlate of protection and a basis for vaccine development. , 2007, Current opinion in biotechnology.

[28]  J. Gog,et al.  Codon conservation in the influenza A virus genome defines RNA packaging signals , 2007, Nucleic acids research.

[29]  K. Fujii,et al.  Contributions of Two Nuclear Localization Signals of Influenza A Virus Nucleoprotein to Viral Replication , 2006, Journal of Virology.

[30]  A. Vincent,et al.  Modulation of Poliovirus Replicative Fitness in HeLa Cells by Deoptimization of Synonymous Codon Usage in the Capsid Region , 2006, Journal of Virology.

[31]  Ian Stansfield,et al.  tRNA properties help shape codon pair preferences in open reading frames , 2006, Nucleic acids research.

[32]  P. Sharp,et al.  Variation in the strength of selected codon usage bias among bacteria , 2005, Nucleic acids research.

[33]  Cecile Viboud,et al.  Impact of influenza vaccination on seasonal mortality in the US elderly population. , 2005, Archives of internal medicine.

[34]  Manuel A. S. Santos,et al.  Comparative context analysis of codon pairs on an ORFeome scale , 2005, Genome Biology.

[35]  A. Osterhaus,et al.  Efficient generation and growth of influenza virus A/PR/8/34 from eight cDNA fragments. , 2004, Virus research.

[36]  Chin-fen Yang,et al.  Multiple amino acid residues confer temperature sensitivity to human influenza virus vaccine strains (FluMist) derived from cold-adapted A/Ann Arbor/6/60. , 2003, Virology.

[37]  Ivan Ivanov,et al.  Missing Codon Pairs in the Genome of Escherichia Coli , 2002, Bioinform..

[38]  G. W. Hatfield,et al.  Nonrandom utilization of codon pairs in Escherichia coli. , 1989, Proceedings of the National Academy of Sciences of the United States of America.

[39]  M Yarus,et al.  Sense codons are found in specific contexts. , 1985, Journal of molecular biology.

[40]  J. Bennetzen,et al.  Codon selection in yeast. , 1982, The Journal of biological chemistry.