The nucleic acid data:
IRESite Id: 571 Version: 5
Originaly submitted by: Martin Mokrejš Submission date: 2009-03-17 01:31:35
Reviewed by: Martin Mokrejš Last change: 2009-09-03 13:40:02
IRESite record type:
  natural_transcript
The shape of the nucleic acid molecule translated:
  linear
The quality of the mRNA/+RNA sequence:
  our_best_guess
The abbreviated name of the virus/gene coding for this mRNA/+RNA molecule:
  VEGF-A
The genetic origin of this natural mRNA/+RNA:
  nuclear
The GenBankId GI:# number of exactly this mRNA/+RNA sequence:
160358806
Synonyms of the gene name:
Synonym: SP163
The mRNA/+RNA description: 
Mus musculus vascular endothelial growth factor A (Vegfa), transcript variant 3, mRNA. Poly(A)-tail sequence
was dropped.
The mRNA/+RNA sequence represented in the +DNA notation:


Credibility of mRNA sequence:
  reverse_engineered_fragment_and_the_rest_is_a_guess
The organism containing this mRNA with IRES segment in its genome:
Mus musculus C57BL/6
A promoter reported in cDNA corresponding to IRES sequence:
  yes
The total number of notable open-reading frames (ORFs):
  2
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:
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:
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:
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
The organism used:
Homo sapiens NIH/3T3 (ATCC CRL-1658)
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:
VEGF-A
The description of the protein encoded in this ORF:
vascular endothelial growth factor A isoform 3 precursor (initiated from CUG codon)
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:
  yes
The ORF absolute position (the base range includes START and STOP codons or their equivalents):
  488-1462
ORF
ORF position:   2
Version: 0
Originaly submitted by: Martin Mokrejš Reviewed by: Martin Mokrejš
The abbreviated name of this ORF/gene:
VEGF-A
The description of the protein encoded in this ORF:
vascular endothelial growth factor A (initiated from AUG codon)
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:
  yes
The ORF absolute position (the base range includes START and STOP codons or their equivalents):
  1022-1462
Remarks:
It is not clear how this VEGF-A region corresponds to the region used by A.-C. Prats and O. Elroy-Stein.
However, Baranick et al. picked the SP163 variant because it was reported to be more efficient than the
"native" 5'-UTR IRES in the past.

Bert et al. (2006) tested VEGF IRES in HeLa cells using promoter-less plasmids with or without the enhancer
left in (Figure 2). They showed there is a cryptic promoter activatable when the enhancer is left in.

Young et al. (2008) in Figure 2C show cryptic promoter activity in the "IRES".

It could Baranick et al. (2008) failed to catch the promoter in HeLa cells because of their PCR-primer
combination designed to reveal splicing issues (Supplementary Figure 7).
Citations:
Baranick B. T., Lemp N. A., Nagashima J., Hiraoka K., Kasahara N., Logg C. R. (2008) Splicing mediates the activity of four putative cellular internal ribosome entry sites. Proc. Natl. Acad. Sci. U. S. A. 105(12):4733-4738
Stein I., Itin A., Einat P., Skaliter R., Grossman Z., Keshet E. (1998) Translation of vascular endothelial growth factor mRNA by internal ribosome entry: implications for translation under hypoxia. Mol. Cell. Biol. 18(6):3112-3119
Bert AG, Grépin R, Vadas MA, Goodall GJ (2006) Assessing IRES activity in the HIF-1alpha and other cellular 5' UTRs. RNA. 12(6):1074-1083
IRESs:
IRES:
Version: 6 Last change: 2009-09-03 13:54:41
Originaly submitted by: Martin Mokrejš Reviewed by: Martin Mokrejš
The IRES name:
  VEGF-A_-133_-1
The IRES absolute position (the range includes START and STOP codons or their equivalents):
  889-1021
Conclusion:
  disproved_IRES
How IRES boundaries were determined:
experimentally_determined
The sequence of IRES region aligned to its secondary structure (if available):


Remarks:
Bert et al. (2006) reported that in direct RNA transfection the VEGF IRES is only 5.4x above the negative
control while EMCV IRES was 221x above (Figure 4).

The "IRES" was found to be not functional in vivo (Figures 1 and 2) while no issues with aberrant splicing
were reported (Baranick et al., 2008, Figure 3D).

The "IRES" is not functional in direct RNA transfection assay nor in (Young et al., 2008, Figure 3). From
Figure 2B one can gather that there in NIH3T3 plasmid transfected cells there is some "IRES" activity or the
VEGF "IRES" is well leaky-scanned so that some additional FLuc is synthesized from bicistronic messages.
Citations:
Baranick B. T., Lemp N. A., Nagashima J., Hiraoka K., Kasahara N., Logg C. R. (2008) Splicing mediates the activity of four putative cellular internal ribosome entry sites. Proc. Natl. Acad. Sci. U. S. A. 105(12):4733-4738
Young RM, Wang SJ, Gordan JD, Ji X, Liebhaber SA, Simon MC (2008) Hypoxia-mediated selective mRNA translation by an internal ribosome entry site-independent mechanism. J. Biol. Chem. 283(24):16309-16319
Bert AG, Grépin R, Vadas MA, Goodall GJ (2006) Assessing IRES activity in the HIF-1alpha and other cellular 5' UTRs. RNA. 12(6):1074-1083
Last change to the database: 2015-04-16 16:45:23 GMT+1