The nucleic acid data:
IRESite Id: 482 Version: 0
Originaly submitted by: Martin Mokrejš
Reviewed by: Martin Mokrejš
IRESite record type:
  plasmid_with_promoter_and_putative_IRES_translationally_characterized
The shape of the nucleic acid molecule translated:
  linear
The quality of the mRNA/+RNA sequence:
  3UTR_possibly_incomplete
The mRNA/+RNA description: 
Putative in vitro BamHI run-off T7 transcript containing synthetic KMI2 IRES.
The mRNA/+RNA sequence represented in the +DNA notation:


Credibility of mRNA sequence:
  reverse_engineered_sequence_and_should_match_experiment_maybe_except_3UTR
The name of the plasmid:
pRKMI2F
The name of the promoter used to express this mRNA:
  T7
The in vivo produced transcripts are heterogeneous (due to any of promoter?/splicing?/cleavage?/breakage?):
  not tested
The in vivo produced heterogeneous transcripts occur due to alternative splicing:
  not tested
A promoter reported in cDNA corresponding to IRES sequence:
  not tested
The origin of IRES in the plasmid:
  engineered
The DNA sequence of the plasmid in (+) orientation annotated by its secondary structure:


GenBank formatted file with annotated plasmid sequence hyperlinked from vector image map:
pRKMI2F.jpg
The total number of notable open-reading frames (ORFs):
  2
Notable Open-Reading Frames (ORFs; protein coding regions) in the mRNA/+RNA sequence:
ORF
ORF position:   1
Version: 0
Originaly submitted by: Martin Mokrejš Reviewed by: Martin Mokrejš
The abbreviated name of this ORF/gene:
RLuc
The description of the protein encoded in this ORF:
Renilla luciferase
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:
  not tested
The ORF absolute position (the base range includes START and STOP codons or their equivalents):
  50-985
ORF
ORF position:   2
Version: 0
Originaly submitted by: Martin Mokrejš Reviewed by: Martin Mokrejš
The abbreviated name of this ORF/gene:
FLuc
The description of the protein encoded in this ORF:
Firefly luciferase
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:
  not tested
The ORF absolute position (the base range includes START and STOP codons or their equivalents):
  1145-2797
Remarks:
Authors did not specify how they produced T7-based transcripts. It is our guess they have used BamHI
restriction site for linearization.
Citations:
Mitchell S. A., Spriggs K. A., Bushell M., Evans J. R., Stoneley M., Le Quesne J. P., Spriggs R. V., Willis A. E. (2005) Identification of a motif that mediates polypyrimidine tract-binding protein-dependent internal ribosome entry. Genes Dev. 19(13):1556-1571
IRESs:
IRES:
Version: 0
Originaly submitted by: Martin Mokrejš Reviewed by: Martin Mokrejš
The IRES name:
  KMI2
The functional status of IRES:
  functional
The IRES absolute position (the range includes START and STOP codons or their equivalents):
  1032-1129
How IRES boundaries were determined:
experimentally_determined
5'-end of IRES relative to last base of the STOP codon of the upstream ORF:
  47
3'-end of IRES relative to last base of the STOP codon of the upstream ORF:
  144
5'-end of IRES relative to first base of the START codon of the downstream ORF:
  -113
3'-end of IRES relative to first base of the START codon of the downstream ORF:
  -16
The sequence of IRES region aligned to its secondary structure (if available):



Citations:
Mitchell S. A., Spriggs K. A., Bushell M., Evans J. R., Stoneley M., Le Quesne J. P., Spriggs R. V., Willis A. E. (2005) Identification of a motif that mediates polypyrimidine tract-binding protein-dependent internal ribosome entry. Genes Dev. 19(13):1556-1571
The translation experiments:
Translation results:
IRESite Translation Id: 534
Version: 0
Originaly submitted by: Martin Mokrejš Reviewed by: Martin Mokrejš
The translation method used to study IRES function:
in vitro
The in vitro translation system:
rabbit reticulocytes lysate
The organism used for translation:
The temperature (in degrees of Celsia):
37
The relative translation efficiency in % of this IRES:
  100.000
Name of the plasmid used as the negative control.
pRF
IRESite Id of the plasmid used as negative control.
  231
The relative translation efficiency in % of the negative control:
  10.000
The size (length) of intercistronic region in the negative control:
67
The effect of 5'-cap analogs on translation:
not tested
Rapamycin affects translation:
not tested
Type of RNA subject to translation:
  exogenous_RNA_with_GpppG_cap_without_polyA_tail
Remarks:
Data from Figure 2I. It is suspicious that authors used as the control pRF-based transcripts instead of
pRKMI(1/2)F based transcripts without supplemented PTB proteins. However, the data showing that unr and La
protein do not bind to pRKMI(1/2)F transcripts so results for these two supplemented proteins confirm that
they most probably wanted to write in the figure legend that they used as the control RRL with same plasmids
but without supplemented PTB.
Citations:
Mitchell S. A., Spriggs K. A., Bushell M., Evans J. R., Stoneley M., Le Quesne J. P., Spriggs R. V., Willis A. E. (2005) Identification of a motif that mediates polypyrimidine tract-binding protein-dependent internal ribosome entry. Genes Dev. 19(13):1556-1571
IRES trans-acting factor (ITAFS):
IRES trans-acting factor (ITAF):
Version: 0
Originaly submitted by: Martin Mokrejš Reviewed by: Martin Mokrejš
Type of the interaction between ITAF and the RNA subject to translation:
direct_interaction_with_rna
OPTIONAL: The interacting RNA base range (if any):
1033-1076
ITAF protein characteristics:
Version: 0
Originaly submitted by: Martin Mokrejš Reviewed by: Václav Vopálenský
ITAF abbreviated name:
PTB-1
ITAF fullname:
polypyrimidine tract-binding protein isoform 1
ITAF description (long):
polypyrimidine tract-binding protein isoform 1 binds dsRNA with (CCU)n motif where n is at least 3. By SELEX approach it was found it binds to 5'-CAGCCUGGUGCCUCUCUUUCGG-3' (Singh et al. (1995) Science 268:1173-1176) but also UCUU or UCUUC within pyrimidine-rich sequence (Perez et al. (1997) RNA3:1334-1347).
3.1.2. 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_and_must_be_supplemented
Method used to demonstrate ITAF effect:
in_vitro
In vitro system used to demonstrate ITAF effect:
rabbit reticulocytes lysate
Remarks:
Data from Figure 2I.
Citations:
Mitchell S. A., Spriggs K. A., Bushell M., Evans J. R., Stoneley M., Le Quesne J. P., Spriggs R. V., Willis A. E. (2005) Identification of a motif that mediates polypyrimidine tract-binding protein-dependent internal ribosome entry. Genes Dev. 19(13):1556-1571
IRES trans-acting factor (ITAF):
Version: 0
Originaly submitted by: Martin Mokrejš Reviewed by: Martin Mokrejš
Type of the interaction between ITAF and the RNA subject to translation:
direct_interaction_with_rna
OPTIONAL: The interacting RNA base range (if any):
1033-1076
ITAF protein characteristics:
Version: 0
Originaly submitted by: Martin Mokrejš Reviewed by: Martin Mokrejš
ITAF abbreviated name:
PTB-2
ITAF fullname:
polypyrimidine tract-binding protein isoform 2
ITAF description (long):
polypyrimidine tract-binding protein isoform 2 binds (CCU)n motif
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_and_must_be_supplemented
Method used to demonstrate ITAF effect:
in_vitro
In vitro system used to demonstrate ITAF effect:
rabbit reticulocytes lysate
Remarks:
Data from Figure 2I.
Citations:
Mitchell S. A., Spriggs K. A., Bushell M., Evans J. R., Stoneley M., Le Quesne J. P., Spriggs R. V., Willis A. E. (2005) Identification of a motif that mediates polypyrimidine tract-binding protein-dependent internal ribosome entry. Genes Dev. 19(13):1556-1571
IRES trans-acting factor (ITAF):
Version: 1 Last change: 2008-07-12 14:24:06
Originaly submitted by: Martin Mokrejš Reviewed by: Martin Mokrejš
Type of the interaction between ITAF and the RNA subject to translation:
no_direct_interaction_with_rna
ITAF protein characteristics:
Version: 0
Originaly submitted by: Martin Mokrejš Reviewed by: Martin Mokrejš
ITAF abbreviated name:
PTB-4
ITAF fullname:
polypyrimidine tract-binding protein isoform 4
ITAF description (long):
polypyrimidine tract-binding protein isoform 4 binds (CCU)n motif
3.3.2. 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:
no_effect_on_translation
Method used to demonstrate ITAF effect:
in_vitro
In vitro system used to demonstrate ITAF effect:
rabbit reticulocytes lysate
Remarks:
Data from Figure 2I.
Citations:
Mitchell S. A., Spriggs K. A., Bushell M., Evans J. R., Stoneley M., Le Quesne J. P., Spriggs R. V., Willis A. E. (2005) Identification of a motif that mediates polypyrimidine tract-binding protein-dependent internal ribosome entry. Genes Dev. 19(13):1556-1571
IRES trans-acting factor (ITAF):
Version: 0
Originaly submitted by: Martin Mokrejš Reviewed by: Martin Mokrejš
Type of the interaction between ITAF and the RNA subject to translation:
direct_interaction_with_rna
OPTIONAL: The interacting RNA base range (if any):
1033-1076
ITAF protein characteristics:
Version: 1 Last change: 2008-07-11 23:17:15
Originaly submitted by: Martin Mokrejš Reviewed by: Martin Mokrejš
ITAF abbreviated name:
nPTB
ITAF fullname:
polypyrimidine tract-binding protein isoform n
ITAF description (long):
neuronal polypyrimidine tract-binding protein isoform binds (CCU)n motif
3.4.2. 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_and_must_be_supplemented
Method used to demonstrate ITAF effect:
in_vitro
In vitro system used to demonstrate ITAF effect:
rabbit reticulocytes lysate
Remarks:
Data from Figure 2I.
Citations:
Mitchell S. A., Spriggs K. A., Bushell M., Evans J. R., Stoneley M., Le Quesne J. P., Spriggs R. V., Willis A. E. (2005) Identification of a motif that mediates polypyrimidine tract-binding protein-dependent internal ribosome entry. Genes Dev. 19(13):1556-1571
IRES trans-acting factor (ITAF):
Version: 0
Originaly submitted by: Martin Mokrejš Reviewed by: Martin Mokrejš
Type of the interaction between ITAF and the RNA subject to translation:
unknown
ITAF protein characteristics:
Version: 0
Originaly submitted by: Martin Mokrejš Reviewed by: Václav Vopálenský
ITAF abbreviated name:
Unr
ITAF fullname:
upstream of N-ras
ITAF description (long):
Unr protein is known to bind to gaagaaguaa
3.5.2. 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:
no_effect_on_translation
Method used to demonstrate ITAF effect:
in_vitro
In vitro system used to demonstrate ITAF effect:
rabbit reticulocytes lysate
Remarks:
Data from Figure 2I.
Citations:
Mitchell S. A., Spriggs K. A., Bushell M., Evans J. R., Stoneley M., Le Quesne J. P., Spriggs R. V., Willis A. E. (2005) Identification of a motif that mediates polypyrimidine tract-binding protein-dependent internal ribosome entry. Genes Dev. 19(13):1556-1571
IRES trans-acting factor (ITAF):
Version: 0
Originaly submitted by: Martin Mokrejš Reviewed by: Martin Mokrejš
Type of the interaction between ITAF and the RNA subject to translation:
unknown
ITAF protein characteristics:
Version: 2 Last change: 2009-08-29 12:19:15
Originaly submitted by: Martin Mokrejš Reviewed by: Martin Mokrejš
ITAF abbreviated name:
La
ITAF fullname:
La autoantigen
ITAF description (long):
La autoantigen (p52), 52 kDa RNA binding protein, predominantly localized to nucleus, unwinds the dsRNA in ATP-dependent manner, forms a dimer
3.6.2. 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:
no_effect_on_translation
Method used to demonstrate ITAF effect:
in_vitro
In vitro system used to demonstrate ITAF effect:
rabbit reticulocytes lysate
Remarks:
Data from Figure 2I.
Citations:
Mitchell S. A., Spriggs K. A., Bushell M., Evans J. R., Stoneley M., Le Quesne J. P., Spriggs R. V., Willis A. E. (2005) Identification of a motif that mediates polypyrimidine tract-binding protein-dependent internal ribosome entry. Genes Dev. 19(13):1556-1571
Regions with experimentally determined secondary structures:
A region with the experimentally determined secondary structure:
IRESite 2D Struct Id: 20
Version: 0
Originaly submitted by: Martin Mokrejš Reviewed by: Martin Mokrejš
The function of the 2D structure:
IRES
The 2D structure causes frameshift:
unknown
The absolute position of the experimentally mapped region (the range includes START and STOP codons or their equivalents):
1032-1147
The underlying nucleic acid sequence and structure of the mapped region:



Remarks:
Data from Figure 2A(ii).
Citations:
Mitchell S. A., Spriggs K. A., Bushell M., Evans J. R., Stoneley M., Le Quesne J. P., Spriggs R. V., Willis A. E. (2005) Identification of a motif that mediates polypyrimidine tract-binding protein-dependent internal ribosome entry. Genes Dev. 19(13):1556-1571
Last change to the database: 2015-04-16 16:45:23 GMT+1