Crystal structure of the RNA-dependent RNA polymerase from influenza C virus.
Negative-sense RNA viruses, such as influenza, encode large, multidomain RNA-dependent RNA polymerases that can both transcribe and replicate the viral RNA genome. In influenza virus, the polymerase (FluPol) is composed of three polypeptides: PB1, PB2 and PA/P3. PB1 houses the polymerase active site, whereas PB2 and PA/P3 contain, respectively, cap-binding and endonuclease domains required for transcription initiation by cap-snatching. Replication occurs through de novo initiation and involves a complementary RNA intermediate. Currently available structures of the influenza A and B virus polymerases include promoter RNA (the 5' and 3' termini of viral genome segments), showing FluPol in transcription pre-initiation states. Here we report the structure of apo-FluPol from an influenza C virus, solved by X-ray crystallography to 3.9 Å, revealing a new 'closed' conformation. The apo-FluPol forms a compact particle with PB1 at its centre, capped on one face by PB2 and clamped between the two globular domains of P3. Notably, this structure is radically different from those of promoter-bound FluPols. The endonuclease domain of P3 and the domains within the carboxy-terminal two-thirds of PB2 are completely rearranged. The cap-binding site is occluded by PB2, resulting in a conformation that is incompatible with transcription initiation. Thus, our structure captures FluPol in a closed, transcription pre-activation state. This reveals the conformation of newly made apo-FluPol in an infected cell, but may also apply to FluPol in the context of a non-transcribing ribonucleoprotein complex. Comparison of the apo-FluPol structure with those of promoter-bound FluPols allows us to propose a mechanism for FluPol activation. Our study demonstrates the remarkable flexibility of influenza virus RNA polymerase, and aids our understanding of the mechanisms controlling transcription and genome replication.
Sir William Dunn School of Pathology, University of Oxford, South Parks Road, Oxford OX1 3RE, UK.|Division of Structural Biology, Henry Wellcome Building for Genomic Medicine, University of Oxford, Oxford OX3 7BN, UK.
Narin
N
Hengrung
Division of Structural Biology, Henry Wellcome Building for Genomic Medicine, University of Oxford, Oxford OX3 7BN, UK.
Kamel
K
El Omari
Sir William Dunn School of Pathology, University of Oxford, South Parks Road, Oxford OX1 3RE, UK.
Itziar
I
Serna Martin
Sir William Dunn School of Pathology, University of Oxford, South Parks Road, Oxford OX1 3RE, UK.
Frank T
FT
Vreede
European Molecular Biology Laboratory, Grenoble Outstation and University Grenoble Alpes-Centre National de la Recherche Scientifique-EMBL Unit of Virus Host-Cell Interactions, 71 Avenue des Martyrs, CS 90181, 38042 Grenoble Cedex 9, France.
Stephen
S
Cusack
Diamond Light Source Ltd, Harwell Science &Innovation Campus, Didcot OX11 0DE, UK.
Robert P
RP
Rambo
Global Phasing Ltd, Sheraton House, Castle Park, Cambridge CB3 0AX, UK.
Clemens
C
Vonrhein
Global Phasing Ltd, Sheraton House, Castle Park, Cambridge CB3 0AX, UK.
Gérard
G
Bricogne
Division of Structural Biology, Henry Wellcome Building for Genomic Medicine, University of Oxford, Oxford OX3 7BN, UK.|Diamond Light Source Ltd, Harwell Science &Innovation Campus, Didcot OX11 0DE, UK.
David I
DI
Stuart
Division of Structural Biology, Henry Wellcome Building for Genomic Medicine, University of Oxford, Oxford OX3 7BN, UK.|Diamond Light Source Ltd, Harwell Science &Innovation Campus, Didcot OX11 0DE, UK.
Jonathan M
JM
Grimes
Sir William Dunn School of Pathology, University of Oxford, South Parks Road, Oxford OX1 3RE, UK.
Ervin
E
Fodor
D001051:Apoenzymes / Q000737:chemistry / Q000378:metabolism; D001665:Binding Sites; D018360:Crystallography, X-Ray; D004720:Endonucleases / Q000737:chemistry / Q000378:metabolism; D004789:Enzyme Activation; D009982:Gammainfluenzavirus / Q000201:enzymology*; D008958:Models, Molecular; D010442:Peptide Chain Initiation, Translational; D011401:Promoter Regions, Genetic / Q000235:genetics; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D021122:Protein Subunits / Q000737:chemistry / Q000378:metabolism; D012315:RNA Caps / Q000378:metabolism; D012367:RNA, Viral / Q000096:biosynthesis / Q000378:metabolism; D012324:RNA-Dependent RNA Polymerase / Q000737:chemistry* / Q000378:metabolism; D012261:Ribonucleoproteins / Q000737:chemistry
D001051:Apoenzymes; D021122:Protein Subunits; D012315:RNA Caps; D012367:RNA, Viral; D012261:Ribonucleoproteins; D012324:RNA-Dependent RNA Polymerase; D004720:Endonucleases