Analysis of Assembly and Budding of Lujo Virus

Abstract

cThe recently identified arenavirus Lujo virus (LUJV) causes fatal hemorrhagic fever in humans. We analyzed its mechanism of viral release driven by matrix protein Z and the cell surface glycoprotein precursor GPC. The L domains in Z are required for efficient virus-like particle release, but Tsg101, ALIX/AIP1, and Vps4A/B are unnecessary for budding. LUJV GPC is cleaved by site 1 protease (S1P) at the RKLM motif, and treatment with the S1P inhibitor PF-429242 reduced LUJV production.T he family Arenaviridae is divided into Old World (OW) and New World (NW) arenaviruses on the basis of the results of geographical, serological, and phylogenetic analyses. The OW arenaviruses include Lassa virus (LASV), the causative agent of Lassa fever, which is endemic to West Africa and has been estimated to infect several hundred thousand individuals annually. The OW arenavirus Lujo virus (LUJV) was associated with a fatal outbreak of hemorrhagic fever in South Africa in 2008. The first patient was transferred from Zambia to South Africa, and four of five patients died (1). LUJV represents a new member of the family Arenaviridae and is the second OW arenavirus reported to cause hemorrhagic fever (1-3). Arenaviruses are enveloped viruses with bisegmented negative-strand RNA genomes consisting of L and S segments. The L segment encodes viral matrix protein Z and RNA-dependent RNA polymerase L, while the S segment encodes nucleoprotein NP and the cell surface glycoprotein precursor GPC in the opposite direction (2). Whole-genome sequence data revealed a unique placement of LUJV among currently known arenaviruses, predicting altered pathogenesis and replication strategies (3). The alignment of LUJV Z is especially unique, as it possesses PT/SAP and YREL sequences, which are known as L-domain motifs critical for virus budding in other viruses and are not conserved among other arenaviruses (4). In addition, although LUJV belongs to the OW arenaviruses, phylogenetic analysis showed that LUJV GPC shows sequence similarity to NW arenaviruses. Interestingly, a recent study showed that the GP-mediated cell entry of LUJV utilizes mechanisms different from those of both OW and NW arenaviruses (5). These observations prompted us to analyze the late stage of LUJV replication, driven by Z and GPC. As the late step of the LUJV life cycle in the cell has not been characterized, we examined the molecular mechanism of LUJV assembly and budding.First, we examined whether the putative L domains within LUJV Z have roles in virus-like particle (VLP) production. The L domain is a short amino acid motif that interacts with cellular factors to facilitate the budding process (4, 6, 7). There are three well-characterized L-domain motifs, PT/SAP, PPxY, and YPXnL (or YxxL), that interact with Tsg101 (8), Nedd4-like E3 ligases (9-14), and ALIX/AIP1 (15-19), respectively. The roles of L domains in arenavirus budding have been well characterized (20)(21)(22). LUJV Z possesses the YREL motif, which is not conserved in other arenaviruses, and a PSAP motif at the...

A Proteomics Survey of Junín Virus Interactions with Human Proteins Reveals Host Factors Required for Arenavirus Replication

“…Knockdown of the ESCRT-I protein TSG101 results in decreased VLP production for LCMV and LASV (17,42). Perturbation of the ESCRT accessory protein VPS4 reduces VLP production for Tacaribe virus and LASV but not Lujo virus (41,47,94). Recently, our laboratory has shown that LCMV utilizes the ESCRT complex specifically for defective interfering particle production but not the release of infectious virus (44).…”

Molecular Mechanisms Underlying the Cellular Entry and Host Range Restriction of Lujo Virus

“…The S segment of the viral genome encodes the glycoprotein (GP) precursor and the nucleoprotein, and the L segment encodes the matrix protein (Z) and the RNA-dependent RNA polymerase (L) ( 2 , 7 ). GP is synthesized as a single polypeptide chain and cleaved posttranslationally into GP1 and GP2 by a cellular proprotein convertase, subtilisin kexin isozyme 1/site 1 protease ( 8 , 9 ). GP1 is located on the top of the viral surface GP spike and mediates the attachment of virions to the target cell.…”

Microtubule-dependent transport of arenavirus matrix protein demonstrated using live-cell imaging microscopy

“…In addition, Z protein plays a crucial role in virion assembly and release (2,13,17). Although the importance of viral late domain motifs in Z protein and the interaction of Z protein with the cellular endosomal sorting complexes required for transport (ESCRT) for virion formation and budding have been well studied (2,13,(17)(18)(19)(20)(21)(22), the transport of Z proteins is been yet fully understood. In the present study, we performed live-cell imaging analyses of Z protein transport in LCMV-infected cells and characterised the movement both over short time period (within 3 min, not certified by peer review) is the author/funder.…”