Internal ribosome entry segment-mediated translation during apoptosis: the role of IRES-trans-acting factors

ABSTRACT To investigate the role of the hepatitis C virus internal ribosome entry site (HCV IRES) domain IV in translation initiation and regulation, two chimeric IRES elements were constructed to contain the reciprocal domain IV in the otherwise HCV and classical swine fever virus IRES elements. This permitted an examination of the role of domain IV in the control of HCV translation. A specific inhibitor of the HCV IRES, vitamin B12, was shown to inhibit translation directed by all IRES elements which contained domain IV from the HCV and the GB virus B IRES elements, whereas the HCV core protein could only suppress translation from the wild-type HCV IRES. Thus, the mechanisms of translation inhibition by vitamin B12 and the core protein differ, and they target different regions of the IRES.

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Poliovirus Internal Ribosome Entry Segment Structure Alterations That Specifically Affect Function in Neuronal Cells: Molecular Genetic Analysis

Journal of Virology ◽

10.1128/jvi.76.21.10617-10626.2002 ◽

2002 ◽

Vol 76 (21) ◽

pp. 10617-10626 ◽

Cited By ~ 37

Author(s):

Cécile E. Malnou ◽

Tuija A. A. Pöyry ◽

Richard J. Jackson ◽

Katherine M. Kean

Keyword(s):

Secondary Structure ◽

Neuronal Cells ◽

Neuronal Cell ◽

In Vitro Translation ◽

Internal Ribosome Entry ◽

Cell Extracts ◽

Internal Ribosome Entry Segment ◽

Domain V ◽

Bulge Loop

ABSTRACT Translation of poliovirus RNA is driven by an internal ribosome entry segment (IRES) present in the 5′ noncoding region of the genomic RNA. This IRES is structured into several domains, including domain V, which contains a large lateral bulge-loop whose predicted secondary structure is unclear. The primary sequence of this bulge-loop is strongly conserved within enteroviruses and rhinoviruses: it encompasses two GNAA motifs which could participate in intrabulge base pairing or (in one case) could be presented as a GNRA tetraloop. We have begun to address the question of the significance of the sequence conservation observed among enterovirus reference strains and field isolates by using a comprehensive site-directed mutagenesis program targeted to these two GNAA motifs. Mutants were analyzed functionally in terms of (i) viability and growth kinetics in both HeLa and neuronal cell lines, (ii) structural analyses by biochemical probing of the RNA, and (iii) translation initiation efficiencies in vitro in rabbit reticulocyte lysates supplemented with HeLa or neuronal cell extracts. Phenotypic analyses showed that only viruses with both GNAA motifs destroyed were significantly affected in their growth capacities, which correlated with in vitro translation defects. The phenotypic defects were strongly exacerbated in neuronal cells, where a temperature-sensitive phenotype could be revealed at between 37 and 39.5°C. Biochemical probing of mutated domain V, compared to the wild type, demonstrated that such mutations lead to significant structural perturbations. Interestingly, revertant viruses possessed compensatory mutations which were distant from the primary mutations in terms of sequence and secondary structure, suggesting that intradomain tertiary interactions could exist within domain V of the IRES.

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Connexins and Cap-independent translation: Role of internal ribosome entry sites

Brain Research ◽

10.1016/j.brainres.2012.05.065 ◽

2012 ◽

Vol 1487 ◽

pp. 99-106 ◽

Cited By ~ 6

Author(s):

Mahboob Ul-Hussain ◽

Rolf Dermietzel ◽

Georg Zoidl

Keyword(s):

Internal Ribosome Entry ◽

Internal Ribosome Entry Sites ◽

Independent Translation

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The 5′ Untranslated Region of Protein Kinase Cδ Directs Translation by an Internal Ribosome Entry Segment That Is Most Active in Densely Growing Cells and during Apoptosis

Molecular and Cellular Biology ◽

10.1128/mcb.22.17.6089-6099.2002 ◽

2002 ◽

Vol 22 (17) ◽

pp. 6089-6099 ◽

Cited By ~ 13

Author(s):

Bronwyn C. Morrish ◽

Martin G. Rumsby

Keyword(s):

Protein Kinase ◽

Untranslated Region ◽

Initiation Factor ◽

Luciferase Reporter ◽

Serum Starvation ◽

Proliferating Cells ◽

Internal Ribosome Entry ◽

Internal Ribosome Entry Segment ◽

Eukaryotic Initiation Factor 4G ◽

Protein Kinase Cδ

ABSTRACT Protein kinase Cδ (PKCδ) is a member of the PKC family of phospholipid-dependent serine/threonine kinases and is involved in cell proliferation, apoptosis, and differentiation. Previous studies have suggested that different PKC isoforms might be translationally regulated. We report here that the 395-nt-long 5′ untranslated region (5′ UTR) of PKCδ is predicted to form very stable secondary structures with free energies (ΔG values) of around −170 kcal/mol. The 5′ UTR of PKCδ can significantly repress luciferase translation in rabbit reticulocyte lysate but does not repress luciferase translation in a number of transiently transfected cell lines. By using a bicistronic luciferase reporter, we show that the 5′ UTR of PKCδ contains a functional internal ribosome entry segment (IRES). The activity of the PKCδ IRES is greatest in densely growing cells and during apoptosis, when total protein synthesis and levels of full-length eukaryotic initiation factor 4G are reduced. However, the IRES activity of the 5′ UTR of PKCδ is not enhanced during serum starvation, another condition shown to inhibit cap-dependent translation, suggesting that its potency is dependent on specific cellular conditions. Accumulating data suggest that PKCδ has a function as proliferating cells reach high density and in early and later events of apoptosis. Our studies suggest a mechanism whereby PKCδ synthesis can be maintained under these conditions when cap-dependent translation is inhibited.

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The role of PERK and GCN2 in basal and hydrogen peroxide-regulated translation from the hepatitis C virus internal ribosome entry site

Virus Genes ◽

10.1007/s11262-011-0629-1 ◽

2011 ◽

Vol 43 (2) ◽

pp. 208-214 ◽

Cited By ~ 5

Author(s):

Samantha C. Jack ◽

Shiu-Wan Chan

Keyword(s):

Hydrogen Peroxide ◽

Hepatitis C Virus ◽

Hepatitis C ◽

Internal Ribosome Entry Site ◽

Entry Site ◽

Internal Ribosome Entry ◽

Virus Internal Ribosome Entry

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Age-Dependent Poliovirus Replication in the Mouse Central Nervous System Is Determined by Internal Ribosome Entry Site-Mediated Translation

Journal of Virology ◽

10.1128/jvi.80.6.2589-2595.2006 ◽

2006 ◽

Vol 80 (6) ◽

pp. 2589-2595 ◽

Cited By ~ 7

Author(s):

Steven Kauder ◽

Sherry Kan ◽

Vincent R. Racaniello

Keyword(s):

Spinal Cord ◽

Internal Ribosome Entry Site ◽

Species Specificity ◽

Entry Site ◽

Internal Ribosome Entry ◽

Adult Mice ◽

Poliovirus Receptor ◽

Mouse Cells ◽

Brain And Spinal Cord

ABSTRACT Mouse cells are not permissive for the replication of human rhinovirus type 2 (HRV2). To determine the role of the HRV2 internal ribosome entry site (IRES) in determining species specificity, a recombinant poliovirus (P1/HRV2) was constructed by substituting the poliovirus IRES with the IRES from HRV2. This recombinant virus replicated in all human and murine cell lines examined, demonstrating that the HRV2 IRES does not limit viral replication in transformed murine cells. P1/HRV2 replicated in the brain and spinal cord in neonatal but not adult mice transgenic for the poliovirus receptor, CD155. Passage of P1/HRV2 in mice led to selection of a virus that caused paralysis in neonatal mice. To determine the relationship between HRV2 IRES-mediated translation and replication of P1/HRV2 in mice, recombinant human adenoviruses were used to express bicistronic mRNAs in murine organs. The results demonstrate that the HRV2 IRES mediates translation in organs of neonatal but not adult mice. These findings show that HRV2 IRES-mediated translation is a determinant of virus replication in the murine brain and spinal cord and suggest that the IRES determines the species specificity of HRV2 infection.

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The Apaf-1 Internal Ribosome Entry Segment Attains the Correct Structural Conformation for Function via Interactions with PTB and unr

Molecular Cell ◽

10.1016/s1097-2765(03)00093-5 ◽

2003 ◽

Vol 11 (3) ◽

pp. 757-771 ◽

Cited By ~ 153

Author(s):

Sally A. Mitchell ◽

Keith A. Spriggs ◽

Mark J. Coldwell ◽

Richard J. Jackson ◽

Anne E. Willis

Keyword(s):

Internal Ribosome Entry ◽

Structural Conformation ◽

Internal Ribosome Entry Segment

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A dynamic RNA loop in an IRES affects multiple steps of elongation factor-mediated translation initiation

eLife ◽

10.7554/elife.08146 ◽

2015 ◽

Vol 4 ◽

Cited By ~ 17

Author(s):

Marisa D Ruehle ◽

Haibo Zhang ◽

Ryan M Sheridan ◽

Somdeb Mitra ◽

Yuanwei Chen ◽

...

Keyword(s):

Elongation Factor ◽

Model Systems ◽

Initiation Mechanism ◽

Translation Elongation ◽

80S Ribosome ◽

Internal Ribosome Entry ◽

Elongation Cycle ◽

Different Types ◽

Rna Pseudoknot

Internal ribosome entry sites (IRESs) are powerful model systems to understand how the translation machinery can be manipulated by structured RNAs and for exploring inherent features of ribosome function. The intergenic region (IGR) IRESs from the Dicistroviridae family of viruses are structured RNAs that bind directly to the ribosome and initiate translation by co-opting the translation elongation cycle. These IRESs require an RNA pseudoknot that mimics a codon-anticodon interaction and contains a conformationally dynamic loop. We explored the role of this loop and found that both the length and sequence are essential for translation in different types of IGR IRESs and from diverse viruses. We found that loop 3 affects two discrete elongation factor-dependent steps in the IRES initiation mechanism. Our results show how the IRES directs multiple steps after 80S ribosome placement and highlights the often underappreciated significance of discrete conformationally dynamic elements within the context of structured RNAs.

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