Identification of Two Functional Domains within the Arenavirus Nucleoprotein

Macleod, Jesica M. Levingston; D'antuono, Alejandra Lorena; Loureiro, María Eugenia; Casabona, Juan Cruz; Gómez, Guillermo A.; López, Nora

doi:10.1128/jvi.01875-10

Cited by 42 publications

(61 citation statements)

References 63 publications

Supporting

Mentioning

Contrasting

Unclassified

Order By: Relevance

“…Our studies demonstrated that TCRV NP is able to self-oligomerize with a yet undefined molecularity and identified the N-ter domain (residues 1 to 332) as being essential for NP self-interaction (11). Biochemical evidence supporting the relevance of the N-ter domain in self-association has also been reported for the NP of the OW arenaviruses lymphocytic choriomeningitis virus (LCMV) and LASV (24,25).…”

mentioning

confidence: 60%

“…Furthermore, a 28-residue region predicted to fold into an ␣-helix (helix 5) within the N-ter domain was identified as being essential to sustain TCRV NP self-association (11). However, the mechanism by which NP homo-oligomerization contributes to RNP assembly as well as to viral transcription and replication remains unclear.…”

Section: Effect Of N-terminal and C-terminal Mutations On Tcrv Np Selmentioning

confidence: 99%

“…Previous studies on TCRV and LCMV demon-strated that NP self-associates in mammalian cells into oligomers and that the NP N-ter domain plays a relevant role in homooligomerization (11,24). Furthermore, a 28-residue region predicted to fold into an ␣-helix (helix 5) within the N-ter domain was identified as being essential to sustain TCRV NP self-association (11).…”

Section: Effect Of N-terminal and C-terminal Mutations On Tcrv Np Selmentioning

confidence: 99%

“…Plasmids expressing the Flag-or c-Myc-tagged versions of TCRV NP (pTCRV N-Flag and pTCRV N-myc, respectively) and plasmids pN⌬1-359, pN⌬334 -476, and pN⌬477-570, which express TCRV NP deletion mutants, were generated previously (9,11). Amino acid substitutions were introduced in the TCRV NP full-length sequence using a QuikChange site-directed mutagenesis kit (Stratagene), with pTCRV NFlag as the template and primers containing the mutated sequences.…”

Section: Cells and Virus Bsr (A Clone Of Bhk-21mentioning

confidence: 99%

“…Indeed, multiple copies of NP associate with the viral genomic and antigenomic RNAs forming ribonucleoprotein (RNP) complexes called nucleocapsids, which along with the L polymerase are the biologically active units for transcription of nonencapsidated, subgenomic viral mRNAs and for viral genome replication (6)(7)(8). Besides, interaction between NP and the matrix Z protein has been shown to be an important element for packaging of RNPs into viral particles during virion morphogenesis (9)(10)(11)(12)(13). Arenavirus NP has also been linked to inhibition of type I interferon induction (14)(15)(16)(17)(18)(19) and in the case of JUNV, NP has been proposed to act as a decoy substrate for apoptosis inhibition in infected cells (20).…”

mentioning

confidence: 99%

See 4 more Smart Citations

Differential Contributions of Tacaribe Arenavirus Nucleoprotein N-Terminal and C-Terminal Residues to Nucleocapsid Functional Activity

D'antuono

Loureiro

Foscaldi

et al. 2014

J Virol

Self Cite

View full text Add to dashboard Cite

The arenavirus nucleoprotein (NP) is the main protein component of viral nucleocapsids and is strictly required for viral genome replication mediated by the L polymerase. Homo-oligomerization of NP is presumed to play an important role in nucleocapsid assembly, albeit the underlying mechanism and the relevance of NP-NP interaction in nucleocapsid activity are still poorly understood. Here, we evaluate the contribution of the New World Tacaribe virus (TCRV) NP self-interaction to nucleocapsid functional activity. We show that alanine substitution of N-terminal residues predicted to be available for NP-NP interaction strongly affected NP self-association, as determined by coimmunoprecipitation assays, produced a drastic inhibition of transcription and replication of a TCRV minigenome RNA, and impaired NP binding to RNA. Mutagenesis and functional analysis also revealed that, while dispensable for NP self-interaction, key amino acids at the C-terminal domain were essential for RNA synthesis. Furthermore, mutations at these C-terminal residues rendered NP unable to bind RNA both in vivo and in vitro but had no effect on the interaction with the L polymerase. In addition, while all oligomerization-defective variants tested exhibited unaltered capacities to sustain NP-L interaction, NP deletion mutants were fully incompetent to bind L, suggesting that, whereas NP self-association is dispensable, the integrity of both the N-terminal and C-terminal domains is required for binding the L polymerase. Overall, our results suggest that NP self-interaction mediated by the N-terminal domain may play a critical role in TCRV nucleocapsid assembly and activity and that the C-terminal domain of NP is implicated in RNA binding. IMPORTANCEThe mechanism of arenavirus functional nucleocapsid assembly is still poorly understood. No detailed information is available on the nucleocapsid structure, and the regions of full-length NP involved in binding to viral RNA remain to be determined. In this report, novel findings are provided on critical interactions between the viral ribonucleoprotein components. We identify several amino acid residues in both the N-terminal and C-terminal domains of TCRV NP that differentially contribute to NP-NP and NP-RNA interactions and analyze their relevance for binding of NP to the L polymerase and for nucleocapsid activity. Our results provide insight into the contribution of NP self-interaction to RNP assembly and activity and reveal the involvement of the NP C-terminal domain in RNA binding. V iruses belonging to the Arenaviridae family are classified into two main groups, Old World (OW) and New World (NW), according to phylogeny, serological properties, and geographical distribution (1). Several members of this family, such as the NW Junin virus (JUNV) and the OW Lassa virus (LASV), produce serious hemorrhagic fever in humans. Tacaribe virus (TCRV), the prototype of NW arenaviruses, is nonpathogenic, closely related to JUNV, and has been used as a model in numerous studies (1-4). Arenaviruses ar...

show abstract

mentioning

confidence: 60%

Section: Effect Of N-terminal and C-terminal Mutations On Tcrv Np Selmentioning

confidence: 99%

Section: Effect Of N-terminal and C-terminal Mutations On Tcrv Np Selmentioning

confidence: 99%

Section: Cells and Virus Bsr (A Clone Of Bhk-21mentioning

confidence: 99%

mentioning

confidence: 99%

See 3 more Smart Citations

Differential Contributions of Tacaribe Arenavirus Nucleoprotein N-Terminal and C-Terminal Residues to Nucleocapsid Functional Activity

D'antuono

Loureiro

Foscaldi

et al. 2014

J Virol

Self Cite

View full text Add to dashboard Cite

show abstract

The role of oligomerization for the biological functions of the arenavirus nucleoprotein

et al. 2013

View full text Add to dashboard Cite

The Lassa virus nucleoprotein (NP) is a multifunctional protein that plays an essential role in many aspects of the viral life cycle, including RNA encapsidation, viral transcription and replication, recruitment of ribonucleoprotein complexes to viral budding sites, and inhibition of the host cell interferon response. While it is known that NP is capable of forming oligomers, both the oligomeric state of NP in mammalian cells and the significance of NP oligomerization for its various functions remain unclear. Here, we demonstrate that Lassa virus NP solely forms trimers upon expression in mammalian cells. Using a minigenome assay we show that mutants that are not able to form stable trimers are no longer functional during transcription and/or replication of the minigenome, indicating that NP trimerization is essential for transcription and/or replication of the viral genome. However, mutations leading to destabilization of the NP trimer did not impact the incorporation of NP into virus-like particles or its ability to suppress interferon-induced gene expression, two important functions of arenavirus NP.

show abstract

Targeting virulence mechanisms for the prevention and therapy of arenaviral hemorrhagic fever

et al. 2013

View full text Add to dashboard Cite

A number of arenaviruses are pathogenic for humans, but they differ significantly in virulence. Lassa virus, found in West Afri ca, causes severe hemorrhagic fever (HF), while the other principal Old World arenavirus, lymphocytic choriomeningitis virus, causes mild illness in persons with normal immune function, and poses a threat only to immunocompromised individuals. The New World agents, including Junin, Machupo and Sabia virus, are highly pathogenic for humans. Arenaviral HF is characterized by high viremia and general immune suppression, the mechanism of which is unknown. Studies using viral reverse genetics, cell-based assays, animal models and human genome-wide association analysis have revealed potential mechanisms by which arenaviruses cause severe disease in humans. Each of the four viral gene products (GPC, L polymerase, NP, and Z matrix protein) and several host-cell factors (e.g., α-dystroglycan) are responsible for mediating viral entry, genome replication, and the inhibition of apoptosis, translation and interferon-beta (IFNβ) production. This review summarizes current knowledge of the role of each viral protein and host factor in the pathogenesis of arenaviral HF. Insights from recent studies are being exploited for the development of novel therapies.

show abstract

Identification of Two Functional Domains within the Arenavirus Nucleoprotein

Cited by 42 publications

References 63 publications

Differential Contributions of Tacaribe Arenavirus Nucleoprotein N-Terminal and C-Terminal Residues to Nucleocapsid Functional Activity

Differential Contributions of Tacaribe Arenavirus Nucleoprotein N-Terminal and C-Terminal Residues to Nucleocapsid Functional Activity

The role of oligomerization for the biological functions of the arenavirus nucleoprotein

Targeting virulence mechanisms for the prevention and therapy of arenaviral hemorrhagic fever

Contact Info

Product

Resources

About