A promoter reported in cDNA corresponding to IRES sequence: no
The total number of notable open-reading frames (ORFs): 1
Summary of possible issues when IRES cDNA is experimentally transcribed in vivo:
Summary of experiments studying integrity of the in vivo transcripts in a particular host:
Integrity (uniformity) of mRNA tested using Northern-blot: homogeneous_population_of_molecules_confirmed
Integrity (uniformity) of mRNA tested using RNase protection: not_tested
Integrity (uniformity) of mRNA tested using 5'-RACE: not_tested
Integrity (uniformity) of mRNA tested using primer extension : not_tested
Integrity (uniformity) of mRNA tested using RT-PCR: not_tested
Integrity (uniformity) of mRNA tested using real-time quantitative polymerase chain reaction (rtqPCR): not_tested
Integrity (uniformity) of mRNA tested using RNAi: not_tested
Integrity (uniformity) of mRNA tested using S1 nuclease mapping: not_tested
Cryptic promoter presence was confirmed by expression from a promoter-less plasmid: no_promoter_confirmed
Cryptic promoter presence was confirmed in an experimental setup involving inducible promoter: not_tested
Integrity (uniformity) of mRNA molecules or possible promoter presence expressed in vivo was tested using another method, please specify in Remarks: not_tested
Summary of experiments studying integrity of the in vivo transcripts in a particular host:
Integrity (uniformity) of mRNA tested using Northern-blot: not_tested
Integrity (uniformity) of mRNA tested using RNase protection: not_tested
Integrity (uniformity) of mRNA tested using 5'-RACE: not_tested
Integrity (uniformity) of mRNA tested using primer extension : not_tested
Integrity (uniformity) of mRNA tested using RT-PCR: not_tested
Integrity (uniformity) of mRNA tested using real-time quantitative polymerase chain reaction (rtqPCR): not_tested
Integrity (uniformity) of mRNA tested using RNAi: homogeneous_population_of_molecules_confirmed
Integrity (uniformity) of mRNA tested using S1 nuclease mapping: not_tested
Cryptic promoter presence was confirmed by expression from a promoter-less plasmid: not_tested
Cryptic promoter presence was confirmed in an experimental setup involving inducible promoter: not_tested
Integrity (uniformity) of mRNA molecules or possible promoter presence expressed in vivo was tested using another method, please specify in Remarks: not_tested
The abbreviated name of this ORF/gene: polyprotein
The description of the protein encoded in this ORF: genome polyprotein
The translational frameshift (ribosome slippage) involved: 0
The ribosome read-through involved: no
The alternative forms of this protein occur by the alternative initiation of translation: no
The ORF absolute position (the base range includes START and STOP codons or their equivalents): 611-7063
Remarks:
The sequence of the HRV2 IRES was inferred from pR-HRV-F plasmid sequence provided by N. S. Magnuson which was
actually provided from the lab of A. Willis (who copied the sequence from R. Jacksons plasmid pXLJ(10-605)).
However, A. Willis utilized PvuII/NcoI sites of the plasmid whereas N. S. Magnuson utilized SpeI/NcoI sites
(it is unclear which HRV-2 sequence used was thus shorter).
Cammas et al. (2007) excluded both splicing and cryptic promoter issues with the pR-HRV-F plasmid in DNA
transfections (Supplemental Figure S2).
The IRES name: HRV-2 Warning: please make ires_name same as the gene_name and optionally append to it coordinates. E.g. when gene/virus name is EMCV-R use EMCV-R_-222_to_-1 or EMCV-R_1-456, etc. but not Emcv-R-... or EMCV-222_to_-1. Please keep case of letters as well. This rewards when searching through the database.
The IRES absolute position (the range includes START and STOP codons or their equivalents): 11-614
Conclusion: strongly_supported_IRES
How IRES boundaries were determined: experimentally_determined
The sequence of IRES region aligned to its secondary structure (if available):
Remarks:
The 5'-UTR tested was confirmed to be functional as IRES and without cryptic promoter activity. No integrity
of the transcripts was studied in case of in vivo DNA plasmid transfections. But, direct RNA
transfections have confirmed IRES activity (Stoneley et al. (2000), Figure 6C). The IRES sequence is
terminated here after the naturally occurring NcoI site (ccATGg).
ITAF fullname: poly(rC)-binding protein 2 (39 kDa)
ITAF description (long): Required for poliovirus IRES activity (Blyn et al. 1996 and 1997) and replication (Toyoda et al., 2007). Its
amount is not limiting in rabbit reticulocyte lysates (RRL) (Hunt and Jackson 1999).
3.2.2. Organisms or in vitro systems where this ITAF was functionally studied:
Organism or in vitro system where ITAF was shown:
Necessity of ITAF for translation in this particular organism or system: required_but_available_internally
Method used to demonstrate ITAF effect: in_vivo
The organism where action of this ITAF was studied:
The interaction was confirmed by several methods using HeLa cell nuclear extracts (NE) with 32P-labeled RNAs.
Data in Figure 1. In Supplemental Figure S1 are describe HRV-2 mutant IRESs lacking the binding activity.