Sequestration of G3BP coupled with efficient translation inhibits stress granules in Semliki Forest virus infection

Panas, Marc D.; Varjak, Margus; Lulla, Aleksei; Eng, Kai Er; Merits, Andres; Hedestam, Gunilla B. Karlsson; McInerney, Gerald M.

doi:10.1091/mbc.e12-08-0619

Cited by 160 publications

(270 citation statements)

References 43 publications

Supporting

Mentioning

254

Contrasting

Order By: Relevance

“…1A). We did not observe the persistence of TIA-1-positive SGs in SFV-⌬P1ϩ2-infected cells (data not shown) as we had done previously in cells infected with SFV-⌬789, which does not recruit G3BP to viral replication complexes or similar structures (8). These data show that the prolinerich region of SFV was not important for the interaction between nsP3 and G3BP-1 in MEFs.…”

supporting

confidence: 76%

“…A surprising difference in the findings of the two groups was that the work of Fros and colleagues suggested that the CHIKV nsP3 sequence binding the SH3 domain of amphiphysins (398-PVAPPRRRR-406 [10]) was essential for colocalization of nsP3 and G3BP (9). In contrast to that work, our study showed that the SFV nsP3 C-terminal L/ITFGDFD repeat motifs at positions 449 to 455 and 466 to 472, both well conserved in the Old World alphaviruses (11), were necessary and sufficient for formation of the nsP3/G3BP complex in infected cells (8). Both these regions, although situated within the hypervariable domain (HVD) of nsP3 (12)(13)(14)(15), are highly conserved between both CHIKV and SFV, and it was therefore surprising that the viruses would differ in the region used to bind and recruit G3BP.…”

contrasting

confidence: 61%

“…Many diverse virus infections employ mechanisms to restrict the formation of SGs (4,5), which are induced by early events in the replication of many viruses, including the alphaviruses (6,7). We recently showed that in cells infected with the alphavirus Semliki Forest virus (SFV), nonstructural protein 3 (nsP3) binds RasGAP SH3-domain binding proteins 1 and 2 (G3BP-1 and G3BP-2, respectively), recruits them to foci containing other viral proteins and often double-stranded RNA (dsRNA), and, in doing so, inhibits SG assembly (8). Fros and colleagues (9) also showed that the expression of the closely related Chikungunya virus (CHIKV) nsP3 blocks SG assembly by recruitment of G3BP-1 into cytoplasmic foci.…”

mentioning

confidence: 99%

“…Transfections were performed as described previously (8). The boundaries of each of the deletions are presented in Fig.…”

mentioning

confidence: 99%

See 3 more Smart Citations

The C-Terminal Repeat Domains of nsP3 from the Old World Alphaviruses Bind Directly to G3BP

Panas

Ahola

McInerney

2014

J Virol

Self Cite

129

View full text Add to dashboard Cite

bThe Old World alphaviruses block stress granule assembly by sequestration of RasGAP SH3-domain binding protein (G3BP). Here, we show that the proline-rich sequences in the hypervariable domain of nonstructural protein 3 (nsP3) of both Semliki Forest virus and Chikungunya virus were dispensable for binding to G3BP. nsP3 variants with or without this domain colocalized with G3BP. Furthermore, we show that the C-terminal repeat motifs of nsP3 were sufficient for G3BP binding. Stress granules (SGs) are foci of accumulation of translationally silent mRNP complexes, which are induced during cellular stress (1). Their assembly is dependent on the proteins TIA-1/R (2) and G3BP (3). Many diverse virus infections employ mechanisms to restrict the formation of SGs (4, 5), which are induced by early events in the replication of many viruses, including the alphaviruses (6, 7). We recently showed that in cells infected with the alphavirus Semliki Forest virus (SFV), nonstructural protein 3 (nsP3) binds RasGAP SH3-domain binding proteins 1 and 2 (G3BP-1 and G3BP-2, respectively), recruits them to foci containing other viral proteins and often double-stranded RNA (dsRNA), and, in doing so, inhibits SG assembly (8). Fros and colleagues (9) also showed that the expression of the closely related Chikungunya virus (CHIKV) nsP3 blocks SG assembly by recruitment of G3BP-1 into cytoplasmic foci. These reports therefore describe a function for Old World alphavirus nsP3 in inhibition of the stress response. A surprising difference in the findings of the two groups was that the work of Fros and colleagues suggested that the CHIKV nsP3 sequence binding the SH3 domain of amphiphysins ) was essential for colocalization of nsP3 and G3BP (9). In contrast to that work, our study showed that the SFV nsP3 C-terminal L/ITFGDFD repeat motifs at positions 449 to 455 and 466 to 472, both well conserved in the Old World alphaviruses (11), were necessary and sufficient for formation of the nsP3/G3BP complex in infected cells (8). Both these regions, although situated within the hypervariable domain (HVD) of nsP3 (12-15), are highly conserved between both CHIKV and SFV, and it was therefore surprising that the viruses would differ in the region used to bind and recruit G3BP.Recently, a variant of SFV (SFV-⌬P1ϩ2) was described, lacking the proline-rich SH3-domain binding sequences of nsP3 (10). It was shown that the deletion impairs the recruitment of amphiphysin proteins to foci of nsP3 accumulation and delays replication complex formation (10). To determine if this domain of SFV nsP3 was important for the recruitment of G3BP-1, we infected mouse embryonic fibroblasts (MEFs) maintained as described previously (16) with SFV wild type (wt) or SFV-⌬P1ϩ2 at a multiplicity of infection (MOI) of 1, fixed the cells at 8 h postinfection (hpi), and stained them for nsP3 and G3BP-1. Single-plane images were captured using a supercontinuum confocal TCS SP5 X microscope (Leica, Wetzlar, Germany) with a pulsed white light laser. No deficiency in G3BP-1 recruitment...

show abstract

supporting

confidence: 76%

contrasting

confidence: 61%

mentioning

confidence: 99%

“…Transfections were performed as described previously (8). The boundaries of each of the deletions are presented in Fig.…”

mentioning

confidence: 99%

See 2 more Smart Citations

The C-Terminal Repeat Domains of nsP3 from the Old World Alphaviruses Bind Directly to G3BP

Panas

Ahola

McInerney

2014

J Virol

Self Cite

129

View full text Add to dashboard Cite

show abstract

“…nsP1 is a methyltransferase and is associated with cellular membranes (4), nsP3 is a phosphoprotein that recruits host factor G3BP and consequently inhibits the formation of cellular stress granules (5,6), and nsP4 is the viral RNA-dependent RNA polymerase (3). nsP2 contains the viral helicase, protease, and a putative C-terminal methyltransferase domain; associates with many host proteins; and can effectively shut down host cell protein synthesis (7)(8)(9)(10)(11).…”

mentioning

confidence: 99%

The C-Terminal Domain of Chikungunya Virus nsP2 Independently Governs Viral RNA Replication, Cytopathicity, and Inhibition of Interferon Signaling

Fros

Maten

Vlak

et al. 2013

J Virol

118

View full text Add to dashboard Cite

Alphavirus nonstructural protein 2 (nsP2) has pivotal roles in viral RNA replication, host cell shutoff, and inhibition of antiviral responses. Mutations that individually rendered other alphaviruses noncytopathic were introduced into chikungunya virus nsP2. Results show that (i) nsP2 mutation P718S only in combination with KR649AA or adaptive mutation D711G allowed noncytopathic replicon RNA replication, (ii) prohibiting nsP2 nuclear localization abrogates inhibition of antiviral interferon-induced JAK-STAT signaling, and (iii) nsP2 independently affects RNA replication, cytopathicity, and JAK-STAT signaling. Chikungunya virus (CHIKV) is a member of the Alphavirus genus within the Togaviridae family. In humans, infection by this mosquito-borne virus can result in the development of a high fever, rash, and incapacitating, sometimes chronic, arthralgia. In the last decade, CHIKV outbreaks occurred throughout the Indian Ocean region, including La Reunion, infecting up to onethird of the human population, before infecting millions of people in India and Southern Asia (1, 2). CHIKV is a positive-strand RNA virus that replicates in the cytoplasm of infected cells. The genome contains four nonstructural proteins (nsP1 to nsP4) that are directly translated from the genomic RNA (gRNA). The viral structural proteins are translated later in infection from subgenomic mRNA (sgRNA) (3).nsP1 is a methyltransferase and is associated with cellular membranes (4), nsP3 is a phosphoprotein that recruits host factor G3BP and consequently inhibits the formation of cellular stress granules (5, 6), and nsP4 is the viral RNA-dependent RNA polymerase (3). nsP2 contains the viral helicase, protease, and a putative C-terminal methyltransferase domain; associates with many host proteins; and can effectively shut down host cell protein synthesis (7-11). Alphavirus nsP2 also contains a nuclear localization signal (NLS) in its C-terminal domain (CHIKV nsP2 KR649-650) (Fig. 1A, top). nsP2 from related Semliki Forest virus (SFV) and Sindbis virus (SINV) has been shown to translocate to the nucleus (12-14), as specific mutations within the NLS retained SFV nsP2 in the cytoplasm and reduced its cytopathicity (15). In the nucleus, nsP2 of Old World alphaviruses (SFV, SINV, and CHIKV) has been reported to inhibit host cell mRNA transcription via degradation of a subunit of DNA-directed RNA polymerase II (RPB1) (16). Mutation of a conserved proline residue in a site homologous to CHIKV nsP2 P718 (Fig. 1A, bottom) rendered SINV noncytopathic and alleviated the transcriptional inhibition via RPB1 (16-18).In addition, alphavirus nsP2 has been shown to antagonize the host's main antiviral response, the interferon (IFN) response, in two ways: (i) beta interferon (IFN-␤) transcription via global host shutoff and (ii) downstream type I/II IFN-induced Janus kinase/ signal transducers and activators of transcription (JAK-STAT) signaling (16,(19)(20)(21)(22). Upon activation, phosphorylated STAT1/2 proteins translocate as dimers into the nucleus to activate t...

show abstract

Regulation of stress granules and P‐bodies during RNA virus infection

2013

View full text Add to dashboard Cite

RNA granules are structures within cells that play major roles in gene expression and homeostasis. Two principle kinds of RNA granules are conserved from yeast to mammals: stress granules (SGs), which contain stalled translation initiation complexes, and processing bodies (P-bodies, PBs), which are enriched with factors involved in RNA turnover. Since RNA granules are associated with silenced transcripts, viruses subvert RNA granule function for replicative advantages. This review, focusing on RNA viruses, discusses mechanisms that manipulate stress granules and P-bodies to promote synthesis of viral proteins. Three main themes have emerged for how viruses manipulate RNA granules; i) cleavage of key host factors, ii) control of PKR activation and iii) redirecting RNA granule components for new or parallel roles in viral reproduction, at the same time disrupting RNA granules. Viruses utilize one or more of these routes to achieve robust and productive infection.

show abstract

Sequestration of G3BP coupled with efficient translation inhibits stress granules in Semliki Forest virus infection

Cited by 160 publications

References 43 publications

The C-Terminal Repeat Domains of nsP3 from the Old World Alphaviruses Bind Directly to G3BP

The C-Terminal Repeat Domains of nsP3 from the Old World Alphaviruses Bind Directly to G3BP

The C-Terminal Domain of Chikungunya Virus nsP2 Independently Governs Viral RNA Replication, Cytopathicity, and Inhibition of Interferon Signaling

Regulation of stress granules and P‐bodies during RNA virus infection

Contact Info

Product

Resources

About