IRESite_Id:571 summary record describing characteristics of VEGF-A_-133

IRESite: The database of experimentally verified IRES structures

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:

AGCGCAGAGG CTTGGGGCAG CCGAGCTGCA GCGAGGCCGC GGCACTGGGG GCGAGCTGAG CGGCGGCAGC GGAGCTCTGT CGCGAGACGC AGCGACAAGG 
^1         ^11        ^21        ^31        ^41        ^51        ^61        ^71        ^81        ^91        

CAGACTATTC AGCGGACTCA CCAGCCCGGG AGTCTGTGCT CTGGGATTTG ATATTCAAAC CTCTTAATTT TTTTTTCTTA AACTGTATTG TTTTACGCTT 
^101       ^111       ^121       ^131       ^141       ^151       ^161       ^171       ^181       ^191       

TAATTTATTT TTGCTTCCTA TTCCCCTCTT AAATCGTGCC AACGGTTTGA GGAGGTTGGT TCTTCACTCC CTCAAATCAC TTCGGATTGT GGAAATCAGC 
^201       ^211       ^221       ^231       ^241       ^251       ^261       ^271       ^281       ^291       

AGACGAAAGA GGTATCAAGA GCTCCAGAGA GAAGTCAAGG AAGAGAGAGA GAGACCGGTC AGAGAGAGCG CGCTGGCGAG CGAACAGAGA GAGGGACAGG 
^301       ^311       ^321       ^331       ^341       ^351       ^361       ^371       ^381       ^391       

GGCAAAGTGA CTGACCTGCT TTTGGGGGTG ACCGCCAGAG CGCGGCGTGA GCCCTCCCCC TTGGGATCTT GCATCGGACC AGTCGCGCTG ACGGACAGAC 
^401       ^411       ^421       ^431       ^441       ^451       ^461       ^471       ^481       ^491       

AGACAGACAC CGCCCCCAGC CCCAGCGCCC ACCTCCTCGC CGGCGGGCTG CCGACGGTGG ACGCGGCGGC GAGCCGCGAG GAACCGAAGC CCGCGCCCGG 
^501       ^511       ^521       ^531       ^541       ^551       ^561       ^571       ^581       ^591       

AGGCGGGGTG GAGGGGGTCG GGGCTCGCGG GATTGCACGG AAACTTTTCG TCCAACTTCT GGGCTCTTCT CGCTCCGTAG TAGCCGTGGT CTGCGCCGCA 
^601       ^611       ^621       ^631       ^641       ^651       ^661       ^671       ^681       ^691       

GGAGACAAAC CGATCGGAGC TGGGAGAAGT GCTAGCTCGG GCCTGGAGAA GCCGGGGCCC GAGAAGAGAG GGGAGGAAGA GAAGGAAGAG GAGAGGGGGC 
^701       ^711       ^721       ^731       ^741       ^751       ^761       ^771       ^781       ^791       

CGCAGTGGGC GCTCGGCTCT CAGGAGCCGA GCTCATGGAC GGGTGAGGCG GCCGTGTGCG CAGACAGTGC TCCAGCCGCG CGCGCGCCCC AGGCCCCGGC 
^801       ^811       ^821       ^831       ^841       ^851       ^861       ^871       ^881       ^891       

CCGGGCCTCG GTTCCAGAAG GGAGAGGAGC CCGCCAAGGC GCGCAAGAGA GCGGGCTGCC TCGCAGTCCG AGCCGGAGAG GGAGCGCGAG CCGCGCCGGC 
^901       ^911       ^921       ^931       ^941       ^951       ^961       ^971       ^981       ^991       

CCCGGACGGG CCTCCGAAAC CATGAACTTT CTGCTCTCTT GGGTGCACTG GACCCTGGCT TTACTGCTGT ACCTCCACCA TGCCAAGTGG TCCCAGGCTG 
^1001      ^1011      ^1021      ^1031      ^1041      ^1051      ^1061      ^1071      ^1081      ^1091      

CACCCACGAC AGAAGGAGAG CAGAAGTCCC ATGAAGTGAT CAAGTTCATG GATGTCTACC AGCGAAGCTA CTGCCGTCCG ATTGAGACCC TGGTGGACAT 
^1101      ^1111      ^1121      ^1131      ^1141      ^1151      ^1161      ^1171      ^1181      ^1191      

CTTCCAGGAG TACCCCGACG AGATAGAGTA CATCTTCAAG CCGTCCTGTG TGCCGCTGAT GCGCTGTGCA GGCTGCTGTA ACGATGAAGC CCTGGAGTGC 
^1201      ^1211      ^1221      ^1231      ^1241      ^1251      ^1261      ^1271      ^1281      ^1291      

GTGCCCACGT CAGAGAGCAA CATCACCATG CAGATCATGC GGATCAAACC TCACCAAAGC CAGCACATAG GAGAGATGAG CTTCCTACAG CACAGCAGAT 
^1301      ^1311      ^1321      ^1331      ^1341      ^1351      ^1361      ^1371      ^1381      ^1391      

GTGAATGCAG ACCAAAGAAA GACAGAACAA AGCCAGAAAA ATGTGACAAG CCAAGGCGGT GAGCCAGGCT GCAGGAAGGA GCCTCCCTCA GGGTTTCGGG 
^1401      ^1411      ^1421      ^1431      ^1441      ^1451      ^1461      ^1471      ^1481      ^1491      

AACCAGACCT CTCACCGGAA AGACCGATTA ACCATGTCAC CACCACGCCA TCATCGTCAC CGTTGACAGA ACAGTCCTTA ATCCAGAAAG CCTGACATGA 
^1501      ^1511      ^1521      ^1531      ^1541      ^1551      ^1561      ^1571      ^1581      ^1591      

AGGAAGAGGA GACTCTTCGA GGAGCACTTT GGGTCCGGAG GGCGAGACTC CGGCAGACGC ATTCCCGGGC AGGTGACCAA GCACGGTCCC TCGTGGGACT 
^1601      ^1611      ^1621      ^1631      ^1641      ^1651      ^1661      ^1671      ^1681      ^1691      

GGATTCGCCA TTTTCTTATA TCTGCTGCTA AATCGCCAAG CCCGGAAGAT TAGGGTTGTT TCTGGGATTC CTGTAGACAC ACCCACCCAC ATACACACAT 
^1701      ^1711      ^1721      ^1731      ^1741      ^1751      ^1761      ^1771      ^1781      ^1791      

ATATATATAT TATATATATA AATAAATATA TATGTTTTAT ATATAAAATA TATATATATT CTTTTTTTTA AATTAACTCT GCTAATGTTA TTGGTGTCTT 
^1801      ^1811      ^1821      ^1831      ^1841      ^1851      ^1861      ^1871      ^1881      ^1891      

CACTGGATAT GTTTGACTGC TGTGGACTTG TGTTGGGAGG AGGATGTCCT CACTCGGATG CCGACACGGG AGACAATGGG ATGAAAGGCT TCAGTGTGGT 
^1901      ^1911      ^1921      ^1931      ^1941      ^1951      ^1961      ^1971      ^1981      ^1991      

CTGAGAGAGG CCGAAGTCCT TTTGCCTGCC GGGGAGCAAG CAAGGCCAGG GCACGGGGGC ACATTGGCTC ACTTCCAGAA ACACGACAAA CCCATTCCTG 
^2001      ^2011      ^2021      ^2031      ^2041      ^2051      ^2061      ^2071      ^2081      ^2091      

GCCCTGAGTC AAGAGGACAG AGAGACAGAT GATGACAGAG AAAGAGATAA AGATGCCGGT TCCAACCAGA AGTTTGGGGA GCCTCAGGAC ATGGCATGCT 
^2101      ^2111      ^2121      ^2131      ^2141      ^2151      ^2161      ^2171      ^2181      ^2191      

TTGTGGATCC CCATGATAGT CTACAAAAGC ACCCCGCCCC TCTGGGCACT GCCTGGAAGA ATCGGGAGCC TGGCCAGCCT TCAGCTCGCT CCTCCACTTC 
^2201      ^2211      ^2221      ^2231      ^2241      ^2251      ^2261      ^2271      ^2281      ^2291      

TGAGGGGCCT AGGAGGCCTC CCACAGGTGT CCCGGCAAGA GAAGACACGG TGGTGGAAGA AGAGGCCTGG TAATGGCCCC TCCTCCTGGG ACCCCTTCGT 
^2301      ^2311      ^2321      ^2331      ^2341      ^2351      ^2361      ^2371      ^2381      ^2391      

CCTCTCCTTA CCCCACCTCC TGGGTACAGC CCAGGAGGAC CTTGTGTGAT CAGACCATTG AAACCACTAA TTCTGTCCCC AGGAGACTTG GCTGTGTGTG 
^2401      ^2411      ^2421      ^2431      ^2441      ^2451      ^2461      ^2471      ^2481      ^2491      

TGAGTGGCTT ACCCTTCCTC ATCTTCCCTT CCCAAGGCAC AGAGCAATGG GGCAGGACCC GCAAGCCCCT CACGGAGGCA GAGAAAAGAG AAAGTGTTTT 
^2501      ^2511      ^2521      ^2531      ^2541      ^2551      ^2561      ^2571      ^2581      ^2591      

ATATACGGTA CTTATTTAAT AGCCCTTTTT AATTAGAAAT TAAAACAGTT AATTTAATTA AAGAGTAGGG TTTTTTTCAG TATTCTTGGT TAATATTTAA 
^2601      ^2611      ^2621      ^2631      ^2641      ^2651      ^2661      ^2671      ^2681      ^2691      

TTTCAACTAT TTATGAGATG TATCTCTCGC TCTCTCTTAT TTGTACTTGT GTGTGTGTGT GTGTGTGTGT GTGTGTGTGT GTGTGTGTGT GTATGAAATC 
^2701      ^2711      ^2721      ^2731      ^2741      ^2751      ^2761      ^2771      ^2781      ^2791      

TGTGTTTCCA ATCTCTCTCT CCCAGATCGG TGACAGTCAC TAGCTTGTCC TGAGAAGATA TTTAATTTTG CTAACACTCA GCTCTGCCCT CCCTTGTCCC 
^2801      ^2811      ^2821      ^2831      ^2841      ^2851      ^2861      ^2871      ^2881      ^2891      

CACCACACAT TCCTTTGAAA TAAGGTTTCA ATATACATTT ACATACTATA TATATATTTG GCAACTTGTG TTTGTATATA AATATATATA TATATATATA 
^2901      ^2911      ^2921      ^2931      ^2941      ^2951      ^2961      ^2971      ^2981      ^2991      

TGTTTATGTA TATATGTGAT TCTGATAAAA TAGACATTGC TATTCTGTTT TTTATATGTA AAAACAAAAC AAGAAAAATA GAGAATTCTA CATACTAAAT 
^3001      ^3011      ^3021      ^3031      ^3041      ^3051      ^3061      ^3071      ^3081      ^3091      

CTCTCTCCTT TTTTAATTTT AATATTTGTT ATCATTTATT TATTGGTGCT ACTGTTTATC CGTAATAATT GTGGGGGAAA AAGATATTAA CATCACGTCT 
^3101      ^3111      ^3121      ^3131      ^3141      ^3151      ^3161      ^3171      ^3181      ^3191      

TTGTCTCTAG AGCAGTTTTC CGAGATATTC CGTAGTACAT ATTTATTTTT AAACAGCAAC AAAGAAATAC AGATATATCT TAAAAAAAAA AGCATTTTGT 
^3201      ^3211      ^3221      ^3231      ^3241      ^3251      ^3261      ^3271      ^3281      ^3291      

ATTAAAGAAT TGAATTCTGA TCTC
^3301      ^3311      ^3321

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):

CCAGGCCCCG GCCCGGGCCT CGGTTCCAGA AGGGAGAGGA GCCCGCCAAG GCGCGCAAGA GAGCGGGCTG CCTCGCAGTC CGAGCCGGAG AGGGAGCGCG 
^889       ^899       ^909       ^919       ^929       ^939       ^949       ^959       ^969       ^979       

AGCCGCGCCG GCCCCGGACG GGCCTCCGAA ACC
^989       ^999       ^1009      ^1019

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:

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: 2019-03-18 09:32:49 GMT+1