Cardiovirus 2A Protein Associates with 40S but Not 80S Ribosome Subunits during Infection

Groppo, Rachel; Palmenberg, Ann C.

doi:10.1128/jvi.00185-07

Cited by 41 publications

(45 citation statements)

References 27 publications

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“…Polysome analysis was performed as described previously (35). Polysomal fractionation experiments using 500 mM KCl were done as described previously (18).…”

Section: Methodsmentioning

confidence: 99%

“…To determine whether the 80S ribosomes that accumulate on silencing of DHX29 are translation-competent monosomes assembled on the mRNA or mRNA-free ribosomes, we analyzed polysome profiles in the presence of high salt (0.5 M KCl), which disrupts mRNA-free 80S complexes that are not engaged in translation (17,18). Polysome profiles from DHX29-silenced cells showed a dramatically reduced abundance of 80S ribosomes, due to their dissociation into individual 40S and 60S subunits (Fig.…”

Section: Dhx29 Down-regulation Disrupts Polysomes and Inhibits Translmentioning

confidence: 99%

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The helicase protein DHX29 promotes translation initiation, cell proliferation, and tumorigenesis

Parsyan

Shahbazian²,

Martineau³

et al. 2009

Proc. Natl. Acad. Sci. U.S.A.

104

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Translational control plays an important role in cell growth and tumorigenesis. Cap-dependent translation initiation of mammalian mRNAs with structured 5 UTRs requires the DExH-box protein, DHX29, in vitro. Here we show that DHX29 is important for translation in vivo. Down-regulation of DHX29 leads to impaired translation, resulting in disassembly of polysomes and accumulation of mRNA-free 80S monomers. DHX29 depletion also impedes cancer cell growth in culture and in xenografts. Thus, DHX29 is a bona fide translation initiation factor that potentially can be exploited as a target to inhibit cancer cell growth.I nitiation is a tightly regulated rate-limiting step in the translation of eukaryotic mRNAs. Ribosome recruitment to the mRNA commences with binding of translation initiation factor 4F (eIF4F) to the 7-methyl guanosine cap structure, which is present at the 5Ј end of all nuclear-encoded eukaryotic mRNAs (1). eIF4F (comprising the cap-binding protein eIF4E, the DEAD-box RNA helicase eIF4A and eIF4G, a scaffold for binding eIF4E and eIF4A) binds to the cap, unwinds (with the aid of eIF4A) the cap-proximal region of the mRNA, and, through interaction with the ribosome-bound eIF3, recruits the 40S ribosomal subunit to the mRNA (2-4). The 40S subunit then scans the 5Ј UTR in a 5Ј to 3Ј direction until it encounters an initiation codon. A subsequent joining of the 60S ribosomal subunit and release of eIFs result in formation of an elongationcompetent 80S ribosome.Secondary structures in 5ЈUTRs of mRNAs are thought to become unwound to allow ribosomal complexes to move along the mRNA in search of the initiation codon. Thus, in addition to its role in the initial attachment of ribosomal complexes to mRNA, eIF4A is believed to assist ribosomal complexes during scanning (5). Recent observations suggest that the process of eukaryotic initiation requires additional members of the DEAD/DExH-box protein family; for instance, a DEAD-box protein, yeast Ded1, and its mammalian homologue, DDX3, are biochemically and genetically implicated in translation initiation on long structured 5ЈUTRs (6), and another DExH-box protein, DHX29, strongly stimulates cap-dependent initiation on mRNAs with structured 5ЈUTRs in vitro (7). Here we studied the importance of DHX29 for translation in vivo and characterized it as a novel factor required for cell proliferation. Results DHX29 Is a Ubiquitously Expressed Cytoplasmic Protein That Associ-ates with the 40S Ribosomal Subunit. DHX29 has been found to interact with the 40S ribosomal subunit in vitro and to associate with 40S ribosomal complexes in a rabbit reticulocyte lysate (7). To determine how general these findings are, we examined the distribution of DHX29 in polysome preparations from HeLa cells using 2 commercial DHX29 antibodies (Fig. 1A). The specificity of these antibodies to human DHX29 was confirmed by immunoblotting against the purified native protein [supporting information (SI) Fig. S1] and recombinant DHX29, as well as by the loss of immunoreactivity following DHX29 RNAi ...

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“…Polysome analysis was performed as described previously (35). Polysomal fractionation experiments using 500 mM KCl were done as described previously (18).…”

Section: Methodsmentioning

confidence: 99%

Section: Dhx29 Down-regulation Disrupts Polysomes and Inhibits Translmentioning

confidence: 99%

The helicase protein DHX29 promotes translation initiation, cell proliferation, and tumorigenesis

Parsyan

Shahbazian²,

Martineau³

et al. 2009

Proc. Natl. Acad. Sci. U.S.A.

104

View full text Add to dashboard Cite

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“…A variety of mechanisms operating during distinct steps of viral reproduction have been implicated, but the relevant differences between various picornavirus representatives attracted relatively little attention. Now that we know how diverse are the mechanisms used by various picornaviruses to affect the translation of host mRNA (34,35,48,59,78,95), impair nucleocytoplasmic transport (6,9,10,25,36,37,50,64,65), suppress innate immunity (7,26,32,38,62,68,83,91), or affect the apoptotic machinery of the target cells (5,11,15,40,44,54,69,73,75,86), we should not be surprised by the possible variability of the viral effects on plasma membrane integrity. The relevant data are, however, rather scarce.…”

Section: Discussionmentioning

confidence: 99%

Interactions between Viral and Prokaryotic Pathogens in a Mixed Infection with Cardiovirus and Mycoplasma

Lidsky

Romanova

Kolesnikova

et al. 2009

J Virol

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In the natural environment, animal and plant viruses often share ecological niches with microorganisms, but the interactions between these pathogens, although potentially having important implications, are poorly investigated. The present report demonstrates, in a model system, profound mutual effects of mycoplasma and cardioviruses in animal cell cultures. In contrast to mycoplasma-free cells, cultures contaminated with Mycoplasma hyorhinis responded to infection with encephalomyocarditis virus (EMCV), a picornavirus, but not with poliovirus (also a picornavirus), with a strong activation of a DNase(s), as evidenced by the TUNEL (terminal deoxynucleotidyltransferase-mediated dUTP-biotin nick end labeling) immunofluorescence assay and electrophoretic analysis of host DNA. This degradation was reminiscent of that observed upon apoptosis but was caspase independent, judging by the failure of the specific pan-caspase inhibitor Q-VD-OPh to prevent it. The electrophoretic mobility of the enzyme responsible for DNA degradation and dependence of its activity on ionic conditions strongly suggested that it was represented by a DNase(s) of mycoplasma origin. In cells not infected with EMCV, the relevant DNase was dormant. The possibility is discussed that activation of the mycoplasma DNase might be linked to a relatively early increase in permeability of plasma membrane of the infected cells caused by EMCV. This type of unanticipated virus-mycoplasma "cooperation" may exemplify the complexity of pathogen-host interactions under conditions when viruses and microorganisms are infecting the same host. In the course of the present study, it was also demonstrated that pan-caspase inhibitor zVAD(OMe).fmk strongly suppressed cardiovirus polyprotein processing, illustrating an additional pitfall in investigations of viral effects on the apoptotic system of host cells.

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“…The protein influences 4EBP1 pathways in certain cell types, and moreover, 2A-deficient viruses can be rescued by chemical inhibitors of mTOR and phosphatidylinositol 3-kinase (PI3K), elements required for cap-dependent but not virus-dependent translation (19). During infection, a portion of 2A is found in association with 40S, but not 60S or 80S ribosomal subunits, though no determined mechanism yet links these observations (18).…”

mentioning

confidence: 99%

“…Mutagenic mapping has identified a ribosome protein-like nuclear localization signal (NLS) and a C-proximal eIF4E binding site, which partially overlaps the scission cassette sequences and are common to all known cardioviruses (17). Similar mutations, tested during infection, link the activities of 2A (EMCV) to virus-induced shutdown of cap-dependent translation (4,18). The protein influences 4EBP1 pathways in certain cell types, and moreover, 2A-deficient viruses can be rescued by chemical inhibitors of mTOR and phosphatidylinositol 3-kinase (PI3K), elements required for cap-dependent but not virus-dependent translation (19).…”

mentioning

confidence: 99%

Binding Interactions between the Encephalomyocarditis Virus Leader and Protein 2A

Petty

Basta

Bacot‐Davis

et al. 2014

J Virol

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The leader (L) and 2A proteins of cardioviruses are the primary antihost agents produced during infection. For encephalomyocarditis virus (EMCV), the prototype of the genus Cardiovirus, these proteins interact independently with key cellular partners to bring about inhibition of active nucleocytoplasmic trafficking and cap-dependent translation, respectively. L and 2A also bind each other and require this cooperation to achieve their effects during infection.

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Cardiovirus 2A Protein Associates with 40S but Not 80S Ribosome Subunits during Infection

Cited by 41 publications

References 27 publications

The helicase protein DHX29 promotes translation initiation, cell proliferation, and tumorigenesis

The helicase protein DHX29 promotes translation initiation, cell proliferation, and tumorigenesis

Interactions between Viral and Prokaryotic Pathogens in a Mixed Infection with Cardiovirus and Mycoplasma

Binding Interactions between the Encephalomyocarditis Virus Leader and Protein 2A

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