Supplementary Materials

Supplementary Material for:

Antibodies to influenza nucleoprotein cross-react with human hypocretin receptor 2

Syed Sohail Ahmed,* Wayne Volkmuth, José Duca, Lorenzo Corti, Michele Pallaoro, Alfredo Pezzicoli, Anette Karle, Fabio Rigat, Rino Rappuoli, Vas Narasimhan, Ilkka Julkunen, Arja Vuorela, Outi Vaarala, Hanna Nohynek, Franco Laghi Pasini, Emanuele Montomoli, Claudia Trombetta, Christopher M. Adams, Jonathan Rothbard, Lawrence Steinman*

*Corresponding author. E-mail: sohail.q.ahmed{at}gsk.com (S.S.A.); steinman{at}stanford.edu (L.S.)

Published 1 July 2015, Sci. Transl. Med. 7, 294ra105 (2015)
DOI: 10.1126/scitranslmed.aab2354

This PDF file includes:

  • Materials and Methods
  • Fig. S1. Visualization of three-dimensional proteins and surface-exposed domains from published crystal structures of human HCRT receptor 2 and trimer NP structure from the 1/Wilson-Smith/1933 influenza strain.
  • Fig. S2. Trimer NP structure from the 1/Wilson-Smith/1933 influenza strain interrogated for structural chains, known T cell epitopes, sequence conservation, and functional determinants.
  • Fig. S3. The conservation of influenza NP epitopes identified in this study with other influenza strains from 1902 to 2013.
  • Fig. S4. Clinical serum samples with lower signal on ELISA confirmed to be background staining (1:400 dilution) based on microscopic patterns of reactivity.
  • Fig. S5. Microscopy of cell lines engineered to express HCRT receptor 1.
  • Fig. S6. Inhibition of IgG binding to HCRT receptor 2 by isoleucine variant of influenza NP peptide.
  • Fig. S7. Source data for blocking experiments done in triplicate on six subjects with narcolepsy and history of Pandemrix vaccination.
  • Fig. S8. Modeling of influenza NP peptide (amino acids 111 to 122) fit within the HLA-DQB1*0602 (allele strongly associated with narcolepsy).
  • Fig. S9. Western blots for influenza NP using monoclonal antibody lnA245 or monoclonal antibody lnA108.
  • Fig. S10. Extracted ion chromatogram (EIC) generated for the influenza NP peptides ELILYDKEEIR (isoleucine variant), ELILYDKEEMR (methionine variant), and IVVDYMMQKPGK (control sequence from influenza HA contained in all vaccines).
  • Fig. S11. Lineage of influenza vaccine reassortants and laboratory strains generated through crosses with high-yielding donor strains.
  • Fig. S12. Crosses of X-157 high-yielding donor strain with other historical influenza strains.
  • Fig. S13. The identity of the “X” contained in the influenza NP YDKEEXR sequence from NYMC X-157 (CY095712).
  • Fig. S14. Influenza HA–associated immune response in subjects vaccinated with Focetria in 2009.
  • Table S1. Percent of individuals with HLA-DQB1*0602 allele associated with narcolepsy.
  • Table S2. In-cell ELISA IgG binding value (absorbance 450 nm/615 nm) for sera with non–HCRT receptor 2–specific background staining.
  • Table S3. HLA-DQA1*0102:DQB1*0602 (allele tightly associated with narcolepsy-cataplexy) binding to 21- and 15-mer peptides.
  • Table S4. T cell studies published in the literature on the influenza NP motif “YDKEEIRRIWRQ” providing functional evidence for HLA class II processing of this epitope.
  • Table S5. Influenza A virus NP antibody inhibition test with sera from individuals with A(H1N1) pdm09 infection.
  • Table S6. Influenza A virus NP antibody inhibition test with sera from Finnish children not known to have narcolepsy in 2004/2005.
  • Table S7. Source data for influenza NP-associated immune response in subjects vaccinated with Focetria in 2009.
  • References (7679)

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Other Supplementary Material for this manuscript includes the following:

  • Movie S1 (Microsoft Word format). Influenza A virus NP indicating exposed surface location of NP mimic of HCRT receptors 2 and 1.

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