264 : Production and extreme RNA editing of defective interfering particles is associated with interferon induction by human metapneumovirus
Type I interferon (IFN) production is the hallmark for host innate immune responses upon virus infection. While most respiratory viruses carry antagonists of this response, reports on HMPV have been contradictive. Here we demonstrate that HMPV accumulates defective interfering particles (DIs) rapidly upon in vitro passage and that the presence of DIs is associated with IFN production. While low passage recombinant HMPV did not induce IFN, the same virus passaged at high multiplicity of infection (moi) caused high IFN production. Using 454-sequencing, we show that upon high moi passage, each virus particle contained >100 DIs, a number unprecedented for any virus, to our knowledge. This was confirmed by Northern Blot analyses. We identified >40 different DIs in a single virus stock that varied in length and sequence, but all appeared to be generated via the “snap-back” mechanism, and all were formed at the trailer end. Importantly, each of the DIs was edited extensively – presumably by host RNA-editing enzymes such as ADAR – as evidenced by the fact that up to 45% of original Adenosine and Cytodine residues had mutated to Guanidine or Thymidine respectively. Real time PCR and Western blot assays demonstrated the upregulation of ADAR in HMPV-DI infected cells.These data indicate that HMPV has an unusual propensity to generate DIs, and that these DIs are edited at an unprecedented high frequency. These data further indicate that the contradiction in published data on HMPV IFN induction and antagonism may be due to the presence or absence of DIs in virus stocks. The interaction of HMPV DIs with the RNA editing machinery and IFN responses warrants further investigations.