Structure of the recombinant RNA polymerase from African Swine Fever Virus

Abstract African Swine Fever Virus (ASFV) is a Nucleo-Cytoplasmic Large DNA Virus which causes an incurable haemorrhagic fever in wild and domesticated pigs with a high impact on global food security. ASFV replicates in the cytoplasm of the infected cell and encodes its own transcription machinery that is independent of cellular factors. Little is known about how this system works at a molecular level and there are currently no approved vaccines nor antiviral drugs to treat the disease. Here, we present methods to produce recombinant ASFV RNA polymerase (RNAP), functional assays suitable to screen for inhibitors, and high-resolution cryo-Electron Microscopy structures of the viral RNAP in different conformational states. The viral RNAP bears a striking resemblance to RNAPII with bona fide homologs of nine of its twelve subunits. Key differences include the fusion of assembly platform subunits RPB3 and RPB11, and an unusual C-terminal domain of the stalk subunit vRPB7 that is related to the eukaryotic mRNA cap 2´-O-methyl transferase 1. Despite the structural conservation with eukaryotic and bacterial polymerases, the ASFV RNAP is resistant to the inhibitors rifampicin and alpha-amanitin, offering the grounds for the development of highly selective inhibitors. The cryo-EM structures and fully recombinant RNAP system together provide an important tool for the design, development, and screening of antiviral drugs in a low biosafety containment environment.