scholarly journals Sequence analysis of the 5′ untranslated region of swine vesicular disease virus reveals block deletions between the end of the internal ribosomal entry site and the initiation codon

2005 ◽  
Vol 86 (10) ◽  
pp. 2753-2761 ◽  
Author(s):  
Andrew E. Shaw ◽  
Scott M. Reid ◽  
Nick J. Knowles ◽  
Geoffrey H. Hutchings ◽  
Ginette Wilsden ◽  
...  
Keyword(s):  
Sequence Analysis ◽  
Disease Virus ◽  
Untranslated Region ◽  
Internal Ribosomal Entry Site ◽  
Initiation Codon ◽  
Entry Site ◽  
Swine Vesicular Disease Virus ◽  
Ribosomal Entry ◽  
Vesicular Disease

Swine vesicular disease virus (SVDV) is a picornavirus closely related to the human pathogen coxsackievirus B5. In common with other picornaviruses, the 5′ untranslated region (5′ UTR) of SVDV contains an internal ribosomal entry site (IRES) that plays an important role in cap-independent translation. The aim of this study was to use RT-PCR and sequencing to characterize a fragment of the 5′ UTR encompassing the entire IRES. Sequence analysis demonstrated high nucleotide identities within the IRES between 33 representative SVDV isolates. These data support the choice of this region as a diagnostic target and provide information for the improvement of laboratory-based molecular assays to detect SVDV. In contrast to the relative conservation of the IRES element, there was considerable nucleotide variability in the spacer region located between the cryptic AUG at the 3′ end of the IRES and the initiation codon of the polyprotein. Interestingly, 11 SVDV isolates had block deletions of between 6 and 125 nt in this region. Nine of these isolates were of recent European origin and were phylogenetically closely related. In vitro growth studies showed that selected isolates with these deletions had a significantly reduced plaque diameter and grew to a significantly lower titre relative to an isolate with a full-length 5′ UTR. Further work is required to define the significance of these deletions and to assess whether they impact on the pathogenesis of SVD.

scholarly journals The non-primate hepacivirus 5′ untranslated region possesses internal ribosomal entry site activity

2013 ◽  
Vol 94 (12) ◽  
pp. 2657-2663 ◽  
Author(s):  
Hazel Stewart ◽  
Cheryl Walter ◽  
Dale Jones ◽  
Sinead Lyons ◽  
Peter Simmonds ◽  
...  
Keyword(s):  
Untranslated Region ◽  
Internal Ribosomal Entry Site ◽  
Rna Replication ◽  
Entry Site ◽  
Subgenomic Replicon ◽  
Stem Loop ◽  
Long Range Interactions ◽  
Ribosomal Entry ◽  
Internal Translation ◽  
And Function

The 5′ untranslated region (5′UTR) of the recently described non-primate hepacivirus (NPHV) contains a region with sequence homology to the internal ribosomal entry site (IRES) of hepatitis C virus (HCV) and GB virus B (GBV-B). Here, we demonstrated internal translation initiation by the NPHV 5′UTR in a bicistronic vector. An RNA stem–loop upstream of the NPHV IRES was structurally distinct from corresponding regions in HCV and GBV-B, and was not required for IRES function. Insertion of the NPHV stem–loop into the corresponding region of the HCV 5′UTR within the HCV subgenomic replicon significantly impaired RNA replication, indicating that long-range interactions between the 5′UTR and cis-acting downstream elements within the NPHV genome are not interchangeable with those of HCV. Despite similarities in IRES structure and function between hepaciviruses, replication elements in the NPHV 5′UTR appear functionally distinct from those of HCV.

Ribosomal deficiencies in Diamond-Blackfan anemia impair translation of transcripts essential for differentiation of murine and human erythroblasts

Blood ◽  
2012 ◽  
Vol 119 (1) ◽  
pp. 262-272 ◽  
Author(s):  
Rastislav Horos ◽  
Hanna IJspeert ◽  
Dagmar Pospisilova ◽  
Regine Sendtner ◽  
Charlotte Andrieu-Soler ◽  
...  
Keyword(s):  
Translation Initiation ◽  
Rna Binding ◽  
Fetal Liver ◽  
Internal Ribosomal Entry Site ◽  
Ribosomal Subunit ◽  
Translation Efficiency ◽  
Entry Site ◽  
Diamond Blackfan Anemia ◽  
Ribosomal Entry

Abstract Diamond-Blackfan anemia (DBA) is associated with developmental defects and profound anemia. Mutations in genes encoding a ribosomal protein of the small (eg, RPS19) or large (eg, RPL11) ribosomal subunit are found in more than half of these patients. The mutations cause ribosomal haploinsufficiency, which reduces overall translation efficiency of cellular mRNAs. We reduced the expression of Rps19 or Rpl11 in mouse erythroblasts and investigated mRNA polyribosome association, which revealed deregulated translation initiation of specific transcripts. Among these were Bag1, encoding a Hsp70 cochaperone, and Csde1, encoding an RNA-binding protein, and both were expressed at increased levels in erythroblasts. Their translation initiation is cap independent and starts from an internal ribosomal entry site, which appeared sensitive to knockdown of Rps19 or Rpl11. Mouse embryos lacking Bag1 die at embryonic day 13.5, with reduced erythroid colony forming cells in the fetal liver, and low Bag1 expression impairs erythroid differentiation in vitro. Reduced expression of Csde1 impairs the proliferation and differentiation of erythroid blasts. Protein but not mRNA expression of BAG1 and CSDE1 was reduced in erythroblasts cultured from DBA patients. Our data suggest that impaired internal ribosomal entry site–mediated translation of mRNAs expressed at increased levels in erythroblasts contributes to the erythroid phenotype of DBA.

scholarly journals Construction of a full-length infectious cDNA clone of swine vesicular disease virus strain NET/1/92 and analysis of new antigenic variants derived from it

2000 ◽  
Vol 81 (11) ◽  
pp. 2763-2769 ◽  
Author(s):  
J. M. J. Rebel ◽  
C. H. Leendertse ◽  
A. Dekker ◽  
F. van Poelwijk ◽  
R. J. M. Moormann
Keyword(s):  
Amino Acid ◽  
Disease Virus ◽  
Cdna Clone ◽  
Parent Strain ◽  
Biological Properties ◽  
Infectious Cdna Clone ◽  
Full Length ◽  
Swine Vesicular Disease Virus ◽  
Vesicular Disease

The Dutch swine vesicular disease virus (SVDV) isolate NET/1/92 was one of the first isolates belonging to a new SVDV antigenic group. This strain was completely sequenced and was shown to have 93% similarity with the UKG/27/72 isolate. To enable antigenicity, replication, maturation and pathogenicity studies of NET/1/92, an infectious full-length cDNA clone, designated pSVD146, was prepared. The in vitro and in vivo biological properties of the virus derived from pSVD146 were studied by analysing antigenicity, plaque morphology, growth curves and virulence in pigs. The epitopes of newly prepared monoclonal antibodies were roughly mapped by fusion-PCR. Fine mapping of epitopes at the amino acid level was achieved by introducing single amino acid mutations in pSVD146. Two new amino acids important in epitope formation were located in VP1; one was mapped in the C-terminal end and the second is thought to be located in the H–I loop. Growth curve and plaque sizes in vitro were similar between virus derived from pSVD146 and the parent wild-type virus. In virulence studies in pigs, the lesions score, neutralization titres and the seroconversion rates were comparable between virus derived from pSVD146 and the parent strain. Since virus derived from pSVD146 had the same biological properties as the parent strain NET/1/92, the full-length infectious cDNA clone pSVD146 will be very useful in studies of the antigenicity, virulence, pathogenesis, maturation and replication of SVDV.

scholarly journals An Enzymatic Footprinting Analysis of the Interaction of 40S Ribosomal Subunits with the Internal Ribosomal Entry Site of Hepatitis C Virus

2000 ◽  
Vol 74 (14) ◽  
pp. 6242-6250 ◽  
Author(s):  
Victoria G. Kolupaeva ◽  
Tatyana V. Pestova ◽  
Christopher U. T. Hellen
Keyword(s):  
Hepatitis C Virus ◽  
Hepatitis C ◽  
Internal Ribosomal Entry Site ◽  
Initiation Codon ◽  
Entry Site ◽  
Interaction Sites ◽  
Ribosomal Entry ◽  
Binding Cleft ◽  
Domain Iii ◽  
Mrna Binding

ABSTRACT Hepatitis C virus translation is initiated on a ∼330-nucleotide (nt)-long internal ribosomal entry site (IRES) at the 5′ end of the genome. In this process, a 43S preinitiation complex (comprising a 40S ribosomal subunit, eukaryotic initiation factor 3 (eIF3), and a ternary [eIF2-GTP-initiator tRNA] complex) binds the IRES in a precise manner so that the initiation codon is placed at the ribosomal P site. This binding step involves specific interactions between the IRES and different components of the 43S complex. The 40S subunit and eIF3 can bind to the IRES independently; previous analyses revealed that eIF3 binds specifically to an apical half of IRES domain III. Nucleotides in the IRES that are involved in the interaction with the 40S subunit were identified by RNase footprinting and mapped to the basal half of domain III and in domain IV. Interaction sites were identified in locations that have been found to be essential for IRES function, including (i) the apical loop residues GGG266-268 in subdomain IIId and (ii) the pseudoknot. Extensive protection from RNase cleavage also occurred downstream of the pseudoknot in domain IV, flanking both sides of the initiation codon and corresponding in length to that of the mRNA-binding cleft of the 40S subunit. These results indicate that the 40S subunit makes multiple interactions with the IRES and suggest that only nucleotides in domain IV are inserted into the mRNA-binding cleft of the 40S subunit.

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