Rotavirus Infection Induces the Phosphorylation of eIF2α but Prevents the Formation of Stress Granules

ABSTRACT Early during the infection process, rotavirus causes the shutoff of cell protein synthesis, with the nonstructural viral protein NSP3 playing a vital role in the phenomenon. In this work, we have found that the translation initiation factor 2α (eIF2α) in infected cells becomes phosphorylated early after virus infection and remains in this state throughout the virus replication cycle, leading to a further inhibition of cell protein synthesis. Under these restrictive conditions, however, the viral proteins and some cellular proteins are efficiently translated. The phosphorylation of eIF2α was shown to depend on the synthesis of three viral proteins, VP2, NSP2, and NSP5, since in cells in which the expression of any of these three proteins was knocked down by RNA interference, the translation factor was not phosphorylated. The modification of this factor is, however, not needed for the replication of the virus, since mutant cells that produce a nonphosphorylatable eIF2α sustained virus replication as efficiently as wild-type cells. In uninfected cells, the phosphorylation of eIF2α induces the formation of stress granules, aggregates of stalled translation complexes that prevent the translation of mRNAs. In rotavirus-infected cells, even though eIF2α is phosphorylated these granules are not formed, suggesting that the virus prevents the assembly of these structures to allow the translation of its mRNAs. Under these conditions, some of the cellular proteins that form part of these structures were found to change their intracellular localization, with some of them having dramatic changes, like the poly(A) binding protein, which relocates from the cytoplasm to the nucleus in infected cells, a relocation that depends on the viral protein NSP3.

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Stress granule-inducing eukaryotic translation initiation factor 4A inhibitors block influenza A virus replication

10.1101/194589 ◽

2017 ◽

Cited By ~ 1

Author(s):

Patrick D. Slaine ◽

Mariel Kleer ◽

Nathan Smith ◽

Denys A. Khaperskyy ◽

Craig McCormick

Keyword(s):

Protein Synthesis ◽

Influenza A Virus ◽

Viral Protein ◽

Influenza A ◽

Translation Initiation Factor ◽

Host Protein ◽

Initiation Factor ◽

Viral Protein Synthesis ◽

Infected Cells ◽

Pateamine A

ABSTRACTEukaryotic translation initiation factor 4A (eIF4A) is a helicase that facilitates assembly of the translation preinitiation complex by unwinding structured mRNA 5’ untranslated regions. Pateamine A (PatA) and silvestrol are natural products that disrupt eIF4A function and arrest translation, thereby triggering the formation of cytoplasmic aggregates of stalled preinitiation complexes known as stress granules (SGs). Here we examined the effects of eIF4A inhibition by PatA and silvestrol on influenza A virus (IAV) protein synthesis and replication in cell culture. Treatment of infected cells with either PatA or silvestrol at early times post-infection results in SG formation, arrest of viral protein synthesis and failure to replicate the viral genome. PatA, which irreversibly binds to eIF4A, sustained long-term blockade of IAV replication following drug withdrawal, and inhibited IAV replication at concentrations that had minimal cytotoxicity. By contrast, the antiviral effects of silvestrol were fully reversible; drug withdrawal caused rapid SG dissolution and resumption of viral protein synthesis. IAV inhibition by silvestrol was invariably associated with cytotoxicity. PatA blocked replication of genetically divergent IAV strains, suggesting common dependence on host eIF4A activity. This study demonstrates the feasibility of targeting core host protein synthesis machinery to prevent viral replication.IMPORTANCEInfluenza A virus (IAV) relies on cellular protein synthesis to decode viral messenger RNAs. Pateamine A and silvestrol are natural products that inactivate an essential protein synthesis protein known as eIF4A. Here we show that IAV is sensitive to these eIF4A inhibitor drugs. Treatment of infected cells with pateamine A or silvestrol prevented synthesis of viral proteins, viral genome replication and release of infectious virions. The irreversible eIF4A inhibitor pateamine A sustained long-term blockade of viral replication, whereas viral protein synthesis quickly resumed after silvestrol was removed from infected cells. Prolonged incubation of either infected or uninfected cells with these drugs induced the programmed cell death cascade called apoptosis. Our findings suggest that core components of the host protein synthesis machinery are viable targets for antiviral drug discovery. The most promising drug candidates should selectively block protein synthesis in infected cells without perturbing bystander uninfected cells.

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Mammalian Orthoreovirus Particles Induce and Are Recruited into Stress Granules at Early Times Postinfection

Journal of Virology ◽

10.1128/jvi.01239-09 ◽

2009 ◽

Vol 83 (21) ◽

pp. 11090-11101 ◽

Cited By ~ 51

Author(s):

Qingsong Qin ◽

Craig Hastings ◽

Cathy L. Miller

Keyword(s):

Protein Synthesis ◽

Uv Light ◽

Stress Granules ◽

Translation Initiation Factor ◽

Cellular Stress Response ◽

Initiation Factor ◽

Viral Mrna ◽

Mammalian Orthoreovirus ◽

Infected Cells ◽

Core Particles

ABSTRACT Infection with many mammalian orthoreovirus (MRV) strains results in shutoff of host, but not viral, protein synthesis via protein kinase R (PKR) activation and phosphorylation of translation initiation factor eIF2α. Following inhibition of protein synthesis, cellular mRNAs localize to discrete structures in the cytoplasm called stress granules (SGs), where they are held in a translationally inactive state. We examined MRV-infected cells to characterize SG formation in response to MRV infection. We found that SGs formed at early times following infection (2 to 6 h postinfection) in a manner dependent on phosphorylation of eIF2α. MRV induced SG formation in all four eIF2α kinase knockout cell lines, suggesting that at least two kinases are involved in induction of SGs. Inhibitors of MRV disassembly prevented MRV-induced SG formation, indicating that viral uncoating is a required step for SG formation. Neither inactivation of MRV virions by UV light nor treatment of MRV-infected cells with the translational inhibitor puromycin prevented SG formation, suggesting that viral transcription and translation are not required for SG formation. Viral cores were found to colocalize with SGs; however, cores from UV-inactivated virions did not associate with SGs, suggesting that viral core particles are recruited into SGs in a process that requires the synthesis of viral mRNA. These results demonstrate that MRV particles induce SGs in a step following viral disassembly but preceding viral mRNA transcription and that core particles are themselves recruited to SGs, suggesting that the cellular stress response may play a role in the MRV replication cycle.

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Replication and Cytopathic Effect of Oncolytic Vesicular Stomatitis Virus in Hypoxic Tumor Cells In Vitro and In Vivo

Journal of Virology ◽

10.1128/jvi.78.17.8960-8970.2004 ◽

2004 ◽

Vol 78 (17) ◽

pp. 8960-8970 ◽

Cited By ~ 36

Author(s):

John H. Connor ◽

Christine Naczki ◽

Costas Koumenis ◽

Douglas S. Lyles

Keyword(s):

Protein Synthesis ◽

Vesicular Stomatitis Virus ◽

Viral Protein ◽

Translation Initiation Factor ◽

Initiation Factor ◽

Progeny Virus ◽

Hypoxic Stress ◽

Α Subunit ◽

Hypoxic Conditions ◽

Vesicular Stomatitis

ABSTRACT Tumor hypoxia presents an obstacle to the effectiveness of most antitumor therapies, including treatment with oncolytic viruses. In particular, an oncolytic virus must be resistant to the inhibition of DNA, RNA, and protein synthesis that occurs during hypoxic stress. Here we show that vesicular stomatitis virus (VSV), an oncolytic RNA virus, is capable of replication under hypoxic conditions. In cells undergoing hypoxic stress, VSV infection produced larger amounts of mRNA than under normoxic conditions. However, translation of these mRNAs was reduced at earlier times postinfection in hypoxia-adapted cells than in normoxic cells. At later times postinfection, VSV overcame a hypoxia-associated increase in α subunit of eukaryotic initiation factor 2 (eIF-2α) phosphorylation and initial suppression of viral protein synthesis in hypoxic cells to produce large amounts of viral protein. VSV infection caused the dephosphorylation of the translation initiation factor eIF-4E and inhibited host translation similarly under both normoxic and hypoxic conditions. VSV produced progeny virus to similar levels in hypoxic and normoxic cells and showed the ability to expand from an initial infection of 1% of hypoxic cells to spread through an entire population. In all cases, virus infection induced classical cytopathic effects and apoptotic cell death. When VSV was used to treat tumors established in nude mice, we found VSV replication in hypoxic areas of these tumors. This occurred whether the virus was administered intratumorally or intravenously. These results show for the first time that VSV has an inherent capacity for infecting and killing hypoxic cancer cells. This ability could represent a critical advantage over existing therapies in treating established tumors.

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Host and Viral Translational Mechanisms during Cricket Paralysis Virus Infection

Journal of Virology ◽

10.1128/jvi.02006-09 ◽

2009 ◽

Vol 84 (2) ◽

pp. 1124-1138 ◽

Cited By ~ 52

Author(s):

Julianne L. Garrey ◽

Yun-Young Lee ◽

Hilda H. T. Au ◽

Martin Bushell ◽

Eric Jan

Keyword(s):

Protein Synthesis ◽

Viral Protein ◽

Rna Virus ◽

Virus Production ◽

Translation Initiation Factor ◽

Initiation Factor ◽

S2 Cells ◽

Structural Protein ◽

Viral Protein Synthesis ◽

Paralysis Virus

ABSTRACT The dicistrovirus is a positive-strand single-stranded RNA virus that possesses two internal ribosome entry sites (IRES) that direct translation of distinct open reading frames encoding the viral structural and nonstructural proteins. Through an unusual mechanism, the intergenic region (IGR) IRES responsible for viral structural protein expression mimics a tRNA to directly recruit the ribosome and set the ribosome into translational elongation. In this study, we explored the mechanism of host translational shutoff in Drosophila S2 cells infected by the dicistrovirus, cricket paralysis virus (CrPV). CrPV infection of S2 cells results in host translational shutoff concomitant with an increase in viral protein synthesis. CrPV infection resulted in the dissociation of eukaryotic translation initiation factor 4G (eIF4G) and eIF4E early in infection and the induction of deIF2α phosphorylation at 3 h postinfection, which lags after the initial inhibition of host translation. Forced dephosphorylation of deIF2α by overexpression of dGADD34, which activates protein phosphatase I, did not prevent translational shutoff nor alter virus production, demonstrating that deIF2α phosphorylation is dispensable for host translational shutoff. However, premature induction of deIF2α phosphorylation by thapsigargin treatment early in infection reduced viral protein synthesis and replication. Finally, translation mediated by the 5′ untranslated region (5′UTR) and the IGR IRES were resistant to impairment of eIF4F or eIF2 in translation extracts. These results support a model by which the alteration of the deIF4F complex contribute to the shutoff of host translation during CrPV infection, thereby promoting viral protein synthesis via the CrPV 5′UTR and IGR IRES.

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KSHV lytic mRNA is efficiently translated in the absence of eIF4F

10.1101/356162 ◽

2018 ◽

Author(s):

Eric S. Pringle ◽

Carolyn-Ann Robinson ◽

Nicolas Crapoulet ◽

Andrea L-A. Monjo ◽

Katrina Bouzanis ◽

...

Keyword(s):

Gene Expression ◽

Protein Synthesis ◽

Host Cell ◽

Translation Initiation ◽

Viral Protein ◽

Viral Proteins ◽

Lytic Replication ◽

Cell Protein ◽

Viral Protein Synthesis ◽

Viral Mrnas

ABSTRACTHerpesvirus genomes are decoded by host RNA polymerase II, generating messenger ribonucleic acids (mRNAs) that are post-transcriptionally modified and exported to the cytoplasm. These viral mRNAs have 5 ′ -m7GTP caps and poly(A) tails that should permit assembly of canonical eIF4F cap-binding complexes to initiate protein synthesis. However, we have shown that chemical disruption of eIF4F does not impede KSHV lytic replication, suggesting that alternative translation initiation mechanisms support viral protein synthesis. Here, using polysome profiling analysis, we confirmed that eIF4F disassembly did not affect the efficient translation of viral mRNAs during lytic replication, whereas a large fraction of host mRNAs remained eIF4F-dependent. Lytic replication altered multiple host translation initiation factors (TIFs), causing caspase-dependent cleavage of eIF2α and eIF4G1 and decreasing levels of eIF4G2 and eIF4G3. Non-eIF4F TIFs NCBP1, eIF4E2 and eIF4G2 associated with actively translating messenger ribonucleoprotein (mRNP) complexes during KSHV lytic replication, but their depletion by RNA silencing did not affect virion production, suggesting that the virus does not exclusively rely on one of these alternative TIFs for efficient viral protein synthesis. METTL3, an N6-methyladenosine (m6A) methyltransferase that modifies mRNAs and influences translational efficiency, was dispensable for early viral gene expression and genome replication but required for late gene expression and virion production. METTL3 was also subject to caspase-dependent degradation during lytic replication, suggesting that its positive effect on KSHV late gene expression may be indirect. Taken together, our findings reveal extensive remodelling of TIFs during lytic replication, which may help sustain efficient viral protein synthesis in the context of host shutoff.IMPORTANCEViruses use host cell protein synthesis machinery to create viral proteins. Herpesviruses have evolved a variety of ways to gain control over this host machinery to ensure priority synthesis of viral proteins and diminished synthesis of host proteins with antiviral properties. We have shown that a herpesvirus called KSHV disrupts normal cellular control of protein synthesis. A host cell protein complex called eIF4F starts translation of most cellular mRNAs, but we observed it is dispensable for efficient synthesis of viral proteins. Several proteins involved in alternative modes of translation initiation were likewise dispensable. However, an enzyme called METTL3 that modifies mRNAs is required for efficient synthesis of certain late KSHV proteins and productive infection. We observed caspase-dependent degradation of several host cell translation initiation proteins during infection, suggesting that the virus alters pools of available factors to favour efficient viral protein synthesis at the expense of host protein synthesis.

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Paramyxovirus-Induced Shutoff of Host and Viral Protein Synthesis: Role of the P and V Proteins in Limiting PKR Activation

Journal of Virology ◽

10.1128/jvi.02023-07 ◽

2007 ◽

Vol 82 (2) ◽

pp. 828-839 ◽

Cited By ~ 36

Author(s):

Maria D. Gainey ◽

Patrick J. Dillon ◽

Kimberly M. Clark ◽

Mary J. Manuse ◽

Griffith D. Parks

Keyword(s):

Protein Synthesis ◽

Hela Cells ◽

Viral Protein ◽

Time Course ◽

Translation Initiation Factor ◽

Host Protein ◽

Initiation Factor ◽

Viral Protein Synthesis ◽

Viral Mrnas ◽

V Protein

ABSTRACT The paramyxovirus simian virus 5 (SV5) establishes highly productive persistent infections of epithelial cells without inducing a global inhibition of translation. Here we show that an SV5 mutant (the P/V-CPI− mutant) with substitutions in the P subunit of the viral polymerase and the accessory V protein also establishes highly productive infections like wild-type (WT) SV5 but that cells infected with the P/V-CPI− mutant show an overall shutdown of both host and viral translation at late times postinfection. Reduced host and viral protein synthesis with the P/V-CPI− virus was not due to lower levels of mRNA or caspase-dependent apoptosis and correlated with phosphorylation of the translation initiation factor eIF-2α. WT SV5 was a poor activator of the eIF-2α kinase protein kinase R (PKR). By contrast, the P/V-CPI− mutant induced PKR phosphorylation, which correlated with the time course of translation inhibition but was independent of interferon signaling. In HeLa cells that expressed the PKR inhibitor influenza A virus NS1 or reovirus sigma3, the rate of host protein synthesis at late times after infection with the P/V-CPI− mutant was restored to ∼50% that of control HeLa cells. By contrast, the rates of P/V-CPI− viral protein synthesis in HeLa cells expressing NS1 or sigma3 were dramatically enhanced, between 5- and 20-fold, while levels of viral mRNA were increased only slightly (NS1-expressing cells) or remained constant (sigma3-expressing cells). Similar results were found using HeLa cells where PKR levels were reduced due to knockdown by small interfering RNA. Expression of either the WT P or the WT V protein from the genome of the P/V-CPI− mutant resulted in lower levels of PKR activation and rates of host and viral protein synthesis that closely matched those seen with WT SV5. Despite higher rates of translation, cells infected with the V- or P-complemented virus accumulated viral mRNAs to lower levels than that seen with the parental P/V-CPI− mutant. We present a model in which the paramyxovirus P/V gene products limit induction of PKR by limiting the synthesis of aberrant viral mRNAs and double-stranded RNA and thus prevent the shutdown of translation by a mechanism that differs from that of other PKR inhibitors such as NS1 and sigma3.

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Vaccinia Virus Protein Synthesis Has a Low Requirement for the Intact Translation Initiation Factor eIF4F, the Cap-Binding Complex, within Infected Cells

Journal of Virology ◽

10.1128/jvi.72.11.8813-8819.1998 ◽

1998 ◽

Vol 72 (11) ◽

pp. 8813-8819 ◽

Cited By ~ 21

Author(s):

Jacqueline Mulder ◽

Morwenna E. M. Robertson ◽

Rachael A. Seamons ◽

Graham J. Belsham

Keyword(s):

Protein Synthesis ◽

Vaccinia Virus ◽

Translation Initiation Factor ◽

Initiation Factor ◽

Virus Protein ◽

Entry Site ◽

Internal Ribosome Entry ◽

Cap Binding Complex ◽

Infected Cells ◽

Virus Mrnas

ABSTRACT The role of the cap-binding complex, eIF4F, in the translation of vaccinia virus mRNAs has been analyzed within infected cells. Plasmid DNAs, which express dicistronic mRNAs containing a picornavirus internal ribosome entry site, produced within vaccinia virus-infected cells both β-glucuronidase and a cell surface-targeted single-chain antibody (sFv). Cells expressing sFv were selected from nonexpressing cells, enabling analysis of protein synthesis specifically within the transfected cells. Coexpression of poliovirus 2A or foot-and-mouth disease virus Lb proteases, which cleaved translation initiation factor eIF4G, greatly inhibited cap-dependent protein (β-glucuronidase) synthesis. Under these conditions, internal ribosome entry site-directed expression of sFv continued and cell selection was maintained. Furthermore, vaccinia virus protein synthesis persisted in the selected cells containing cleaved eIF4G. Thus, late vaccinia virus protein synthesis has a low requirement for the intact cap-binding complex eIF4F. This may be attributed to the short unstructured 5′ noncoding regions of the vaccinia virus mRNAs, possibly aided by the presence of poly(A) at both 5′ and 3′ termini.

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Rotavirus Nonstructural Protein NSP3 Is Not Required for Viral Protein Synthesis

Journal of Virology ◽

10.1128/jvi.00437-06 ◽

2006 ◽

Vol 80 (18) ◽

pp. 9031-9038 ◽

Cited By ~ 59

Author(s):

Hilda Montero ◽

Carlos F. Arias ◽

Susana Lopez

Keyword(s):

Protein Synthesis ◽

Viral Protein ◽

Nonstructural Protein ◽

Consensus Sequence ◽

Initiation Factor ◽

Cell Protein ◽

Viral Protein Synthesis ◽

Viral Mrnas ◽

Initiation Of Translation ◽

Cellular Mrnas

ABSTRACT Initiation is the rate-limiting step in protein synthesis and therefore an important target for regulation. For the initiation of translation of most cellular mRNAs, the cap structure at the 5′ end is bound by the translation factor eukaryotic initiation factor 4E (eIF4E), while the poly(A) tail, at the 3′ end, is recognized by the poly(A)-binding protein (PABP). eIF4G is a scaffold protein that brings together eIF4E and PABP, causing the circularization of the mRNA that is thought to be important for an efficient initiation of translation. Early in infection, rotaviruses take over the host translation machinery, causing a severe shutoff of cell protein synthesis. Rotavirus mRNAs lack a poly(A) tail but have instead a consensus sequence at their 3′ ends that is bound by the viral nonstructural protein NSP3, which also interacts with eIF4GI, using the same region employed by PABP. It is widely believed that these interactions lead to the translation of rotaviral mRNAs, impairing at the same time the translation of cellular mRNAs. In this work, the expression of NSP3 in infected cells was knocked down using RNA interference. Unexpectedly, under these conditions the synthesis of viral proteins was not decreased, while the cellular protein synthesis was restored. Also, the yield of viral progeny increased, which correlated with an increased synthesis of viral RNA. Silencing the expression of eIF4GI further confirmed that the interaction between eIF4GI and NSP3 is not required for viral protein synthesis. These results indicate that NSP3 is neither required for the translation of viral mRNAs nor essential for virus replication in cell culture.

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Virus-Mediated Compartmentalization of the Host Translational Machinery

mBio ◽

10.1128/mbio.01463-14 ◽

2014 ◽

Vol 5 (5) ◽

Cited By ~ 37

Author(s):

Emily A. Desmet ◽

Lynne J. Anguish ◽

John S. L. Parker

Keyword(s):

Protein Synthesis ◽

Translation Initiation ◽

Rna Binding ◽

Viral Proteins ◽

Translation Initiation Factor ◽

Initiation Factor ◽

Viral Translation ◽

Cytoplasmic Inclusions ◽

Translational Machinery ◽

Translational Apparatus

ABSTRACTViruses require the host translational apparatus to synthesize viral proteins. Host stress response mechanisms that suppress translation, therefore, represent a significant obstacle that viruses must overcome. Here, we report a strategy whereby the mammalian orthoreoviruses compartmentalize the translational machinery within virus-induced inclusions known as viral factories (VF). VF are the sites of reovirus replication and assembly but were thought not to contain ribosomes. It was assumed viral mRNAs exited the VF to undergo translation by the cellular machinery, and proteins reentered the factory to participate in assembly. Here, we used ribopuromycylation to visualize active translation in infected cells. These studies revealed that active translation occurs within VF and that ribosomal subunits and proteins required for translation initiation, elongation, termination, and recycling localize to the factory. Interestingly, we observed components of the 43S preinitiation complex (PIC) concentrating primarily at factory margins, suggesting a spatial and/or dynamic organization of translation within the VF. Similarly, the viral single-stranded RNA binding protein σNS localized to the factory margins and had a tubulovesicular staining pattern that extended a short distance from the margins of the factories and colocalized with endoplasmic reticulum (ER) markers. Consistent with these colocalization studies, σNS was found to associate with both eukaryotic translation initiation factor 3 subunit A (eIF3A) and the ribosomal subunit pS6R. Together, these findings indicate that σNS functions to recruit 43S PIC machinery to the primary site of viral translation within the viral factory. Pathogen-mediated compartmentalization of the translational apparatus provides a novel mechanism by which viruses might avoid host translational suppression.IMPORTANCEViruses lack biosynthetic capabilities and depend upon the host for protein synthesis. This dependence requires viruses to evolve mechanisms to coerce the host translational machinery into synthesizing viral proteins in the face of ongoing cellular stress responses that suppress global protein synthesis. Reoviruses replicate and assemble within cytoplasmic inclusions called viral factories. However, synthesis of viral proteins was thought to occur in the cytosol. To identify the site(s) of viral translation, we undertook a microscopy-based approach using ribopuromycylation to detect active translation. Here, we report that active translation occurs within viral factories and that translational factors are compartmentalized within factories. Furthermore, we find that the reovirus nonstructural protein σNS associates with 43S preinitiation complexes at the factory margins, suggesting a role for σNS in translation. Together, virus-induced compartmentalization of the host translational machinery represents a strategy for viruses to spatiotemporally couple viral protein synthesis with viral replication and assembly.

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Distinct Structural Features ofCaprin-1 Mediate Its Interaction with G3BP-1 and Its Induction of Phosphorylation of Eukaryotic Translation InitiationFactor 2α, Entry to Cytoplasmic Stress Granules, and Selective Interaction with a Subset of mRNAs

Molecular and Cellular Biology ◽

10.1128/mcb.02300-06 ◽

2007 ◽

Vol 27 (6) ◽

pp. 2324-2342 ◽

Cited By ~ 139

Author(s):

Samuel Solomon ◽

Yaoxian Xu ◽

Bin Wang ◽

Muriel D. David ◽

Peter Schubert ◽

...

Keyword(s):

Protein Synthesis ◽

Cellular Proliferation ◽

Stress Granules ◽

Cell Types ◽

Structural Features ◽

Translation Initiation Factor ◽

Initiation Factor ◽

Eukaryotic Translation ◽

And Migration ◽

Carboxy Terminal

ABSTRACT Caprin-1 is a ubiquitously expressed, well-conserved cytoplasmic phosphoprotein that is needed for normal progression through the G1-S phase of the cell cycle and occurs in postsynaptic granules in dendrites of neurons. We demonstrate that Caprin-1 colocalizes with RasGAP SH3 domain binding protein-1 (G3BP-1) in cytoplasmic RNA granules associated with microtubules and concentrated in the leading and trailing edge of migrating cells. Caprin-1 exhibits a highly conserved motif, F(M/I/L)Q(D/E)Sx(I/L)D that binds to the NTF-2-like domain of G3BP-1. The carboxy-terminal region of Caprin-1 selectively bound mRNA for c-Myc or cyclin D2, this binding being diminished by mutation of the three RGG motifs and abolished by deletion of the RGG-rich region. Overexpression of Caprin-1 induced phosphorylation of eukaryotic translation initiation factor 2α (eIF-2α) through a mechanism that depended on its ability to bind mRNA, resulting in global inhibition of protein synthesis. However, cells lacking Caprin-1 exhibited no changes in global rates of protein synthesis, suggesting that physiologically, the effects of Caprin-1 on translation were limited to restricted subsets of mRNAs. Overexpression of Caprin-1 induced the formation of cytoplasmic stress granules (SG). Its ability to bind RNA was required to induce SG formation but not necessarily its ability to enter SG. The ability of Caprin-1 or G3BP-1 to induce SG formation or enter them did not depend on their association with each other. The Caprin-1/G3BP-1 complex is likely to regulate the transport and translation of mRNAs of proteins involved with synaptic plasticity in neurons and cellular proliferation and migration in multiple cell types.

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