Oral Presentation Lorne Infection and Immunity 2014

Interaction of lyssavirus P-protein with STATs is critical to lethal rabies disease  (#27)

Linda Wiltzer 1 , Kazuma Okada 2 , Satoko Yamoaka 2 , Florence Larrous 3 , Henna Kuusisto 1 , Danielle Blondel 4 , Herve Bourhy 3 , David A Jans 1 , Naoto Ito 2 , Gregory W Moseley 1 5
  1. Department of Biochemistry and Molecular Biology, Monash University, Clayton, VIC, Australia
  2. Laboratory of Zoonotic Diseases, Gifu University, Gifu, Japan
  3. Unit Lyssavirus Dynamics and Host Adaptation, Pasteur Institute, Paris, France
  4. Laboratoire de Virologie Moléculaire et Structurale, CNRS, Paris, France
  5. Dept. of Biochemistry and Molecular Biology, Bio21 Institute, University of Melbourne, Parkville, VIC, Australia

Lyssaviruses, including Australian bat lyssavirus (ABLV), rabies virus (RABV), and Mokola virus, are a globally distributed genus of zoonotic pathogens that cause rabies disease with a remarkable case-fatality rate of 100% in humans. Human infection results in >55,000 deaths/year worldwide, and the spread of ABLV to humans and horses in Qld., coupled with recent outbreaks of RABV in the Indonesian archipelago, have highlighted to the risk of emergence and invasion by these viruses in Australasia. 

We found previously that the phospho(P-)proteins of lyssaviruses interact with signal transducers and activators of transcription (STATs) 1-3. This is thought to effect viral evasion of interferon (IFN)-mediated innate immune responses as a major pathogenicity factor. However, the significance of these interactions to in vivo infection and disease progression is unknown. Through an approach combining next-generation sequencing/bioinformatics, site-directed mutagenesis, single live-cell imaging, immune signalling assays, protein-protein interaction analysis, viral reverse genetics, and an in vivo disease model, we have identified the P-protein-STAT interaction site for the first time. We further developed mutations able to specifically inhibit P-protein’s function in antagonising IFN/STAT responses without impairing other critical functions of P-protein in genome replication and inhibition of viral induction of IFN.
Using these mutations, we generated the first fully viable “STAT-blind” lyssavirus. Growth of the mutant virus in IFN-deficient cells was indistinguishable from that of parental wild-type virus, but the mutant virus lacked the capacity to inhibit STAT responses and was severely impaired in its resistance to IFN. Intracerebral inoculation of mice indicated that the mutant virus was profoundly attenuated, showing strongly impaired growth in the CNS and causing no neurological symptoms or deaths. By contrast, the wild-type strain was invariably lethal.
These findings provide the first direct evidence that STAT interactions of P-protein are vital to lyssavirus infection of the CNS and the development of lethal rabies, with significance to efforts to generate new attenuated vaccine strains and/or novel therapeutics against this incurable disease.

  1. Conservation of a unique mechanism of immune evasion across the Lyssavirus genus. Wiltzer L, Larrous F, Oksayan S, Ito N, Marsh GA, Wang LF, Blondel D, Bourhy H, Jans DA, Moseley GW. J Virol. 2012 Sep;86(18):10194-9.
  2. The rabies virus interferon antagonist P protein interacts with activated STAT3 and inhibits Gp130 receptor signaling. Lieu KG, Brice A, Wiltzer L, Hirst B, Jans DA, Blondel D, Moseley GW. J Virol. 2013 Jul;87(14):8261-5.
  3. Role of interferon antagonist activity of rabies virus phosphoprotein in viral pathogenicity. Ito N, Moseley GW, Blondel D, Shimizu K, Rowe CL, Ito Y, Masatani T, Nakagawa K, Jans DA, Sugiyama M. J Virol. 2010 Jul;84(13):6699-710.