pH-Controlled two-step uncoating of influenza virus.
Upon endocytosis in its cellular host, influenza A virus transits via early to late endosomes. To efficiently release its genome, the composite viral shell must undergo significant structural rearrangement, but the exact sequence of events leading to viral uncoating remains largely speculative. In addition, no change in viral structure has ever been identified at the level of early endosomes, raising a question about their role. We performed AFM indentation on single viruses in conjunction with cellular assays under conditions that mimicked gradual acidification from early to late endosomes. We found that the release of the influenza genome requires sequential exposure to the pH of both early and late endosomes, with each step corresponding to changes in the virus mechanical response. Step 1 (pH 7.5-6) involves a modification of both hemagglutinin and the viral lumen and is reversible, whereas Step 2 (pH <6.0) involves M1 dissociation and major hemagglutinin conformational changes and is irreversible. Bypassing the early-endosomal pH step or blocking the envelope proton channel M2 precludes proper genome release and efficient infection, illustrating the importance of viral lumen acidification during the early endosomal residence for influenza virus infection.
Drittes Physikalisches Institut, Georg-August-Universität, Göttingen, Germany.
Sai
S
Li
Institut für Biologie, AG Molekulare Biophysik, Humboldt-Universität zu Berlin, Berlin, Germany.
Christian
C
Sieben
Institut für Chemie und Biochemie, Forschungszentrum für Elektronenmikroskopie, Freie Universität Berlin, Berlin, Germany.
Kai
K
Ludwig
Institut für Biologie, AG Molekulare Biophysik, Humboldt-Universität zu Berlin, Berlin, Germany.
Chris T
CT
Höfer
Institut für Biologie, AG Molekulare Biophysik, Humboldt-Universität zu Berlin, Berlin, Germany.
Salvatore
S
Chiantia
Institut für Biologie, AG Molekulare Biophysik, Humboldt-Universität zu Berlin, Berlin, Germany. Electronic address: andreas.herrmann@rz.hu-berlin.de.
Andreas
A
Herrmann
Drittes Physikalisches Institut, Georg-August-Universität, Göttingen, Germany; Center for Nanoscale Microscopy and Molecular Physiology of the Brain (CNMPB), Göttingen, Germany. Electronic address: frederic.eghiaian@inserm.fr.
Frederic
F
Eghiaian
Drittes Physikalisches Institut, Georg-August-Universität, Göttingen, Germany; Center for Nanoscale Microscopy and Molecular Physiology of the Brain (CNMPB), Göttingen, Germany.
Iwan A T
IA
Schaap
D000818:Animals; D020285:Cryoelectron Microscopy; D004285:Dogs; D011992:Endosomes / Q000737:chemistry / Q000821:virology*; D016679:Genome, Viral; D019267:Hemagglutinin Glycoproteins, Influenza Virus / Q000737:chemistry; D006863:Hydrogen-Ion Concentration; D053122:Influenza A Virus, H3N2 Subtype / Q000737:chemistry / Q000235:genetics / Q000502:physiology* / Q000648:ultrastructure; D008081:Liposomes / Q000737:chemistry; D061985:Madin Darby Canine Kidney Cells; D055595:Mechanical Phenomena; D018625:Microscopy, Atomic Force; D011487:Protein Conformation; D012261:Ribonucleoproteins / Q000737:chemistry; D014763:Viral Matrix Proteins / Q000737:chemistry; D014764:Viral Proteins / Q000737:chemistry; D014771:Virion / Q000737:chemistry; D057646:Virus Uncoating; D000086082:Viroporin Proteins
D019267:Hemagglutinin Glycoproteins, Influenza Virus; D008081:Liposomes; D012261:Ribonucleoproteins; D014763:Viral Matrix Proteins; D014764:Viral Proteins; C488937:M2 protein, Influenza A virus; D000086082:Viroporin Proteins