Conformational changes in Lassa virus L protein associated with promoter binding and RNA synthesis activity

Abstract

Lassa virus is endemic in West Africa and can cause severe hemorrhagic fever. The viral L protein transcribes and replicates the RNA genome via its RNA-dependent RNA polymerase activity. Here, we present nine cryo-EM structures of the L protein in the apo-, promoter-bound pre-initiation and active RNA synthesis states. We characterize distinct binding pockets for the conserved 3’ and 5’ promoter RNAs and show how full-promoter binding induces a distinct pre-initiation conformation. In the apo- and early elongation states, the endonuclease is inhibited by two distinct L protein peptides, whereas in the pre-initiation state it is uninhibited. In the early elongation state, a template-product duplex is bound in the active site cavity together with an incoming non-hydrolysable nucleotide and the full C-terminal region of the L protein, including the putative cap-binding domain, is well-ordered. These data advance our mechanistic understanding of how this flexible and multifunctional molecular machine is activated.

The mechanism of genome replication and transcription in bunyaviruses

“…However, arguably the major breakthrough in recent years has been the visualization of these proteins in action by determining structures of L proteins stalled in key active states. Following the pioneering work on influenza virus polymerase [ 46 , 47 ], snapshots have now been obtained for LACV, LASV, and the SFTSV L proteins [ 48 – 50 ]. LACV L structures revealed the key conformational changes necessary for genome replication and transcription [ 49 ].…”

“…LACV L structures revealed the key conformational changes necessary for genome replication and transcription [ 49 ]. SFTSV and LASV L snapshots depicted the conformational changes associated with promoter binding and genome replication activities [ 48 , 50 ] ( Fig 1 ). In addition, LASV, Junin virus (JUNV), and MACV L have been determined in complex with the viral matrix protein Z, providing structural insights into the regulation of Arenaviridae polymerase activity [ 51 – 53 ], which had been detected previously in in vitro and cell-based assays [ 54 – 59 ].…”

Lassa fever — the road ahead

“…The glycoprotein complex (GPC) mediates viral attachment and cell entry 28 . The Large (L) protein is an RNA polymerase involved in transcription and replication 29 , 30 . L protein has additional functions such as cap-snatching.…”

Revisiting Viral RNA-Dependent RNA Polymerases: Insights from Recent Structural Studies

“…The structures of the arenaviral L–Z complexes aided in the understanding of the molecular mechanism of the on/off switching of vRNA production. The complexes of the Old World arenavirus , Lassa virus (PDB 7OCH, 7OE3, 7OE7, 7OEA, 7OEB, 7OJJ, 7OJK, 7OJL, 7OJN), and the New World arenavirus, Machupo virus (PDB 7CKM, 7ELC), revealed a single copy of the Z bound to the L polymerase at the interface of the PA subunit carboxy (C)-terminal-like (PA-C-like) regions and the RdRP ( Figure 5 ) [ 50 , 51 ]. The central L-binding domain of the Z was shown to regulate the polymerase activity by associating with the motifs D and E, whereas the flexible extremities were proposed to interact with diverse proteins during the assembly for genome encapsidation.…”