IRESite_Id:110 summary record describing characteristics of Apaf-1 IRES

IRESite: The database of experimentally verified IRES structures

The nucleic acid data:

IRESite Id: `110`	Version: `14`
Originaly submitted by: Martin Mokrejš	Submission date: `2006-01-12 00:00:00`
Reviewed by: Martin Mokrejš	Last change: `2015-04-08 23:31:45`

IRESite record type:

natural_transcript

The shape of the nucleic acid molecule translated: `linear`	The quality of the mRNA/+RNA sequence: `hopefully_full-length_mRNA`
The abbreviated name of the virus/gene coding for this mRNA/+RNA molecule: `Apaf-1`	The genetic origin of this natural mRNA/+RNA: `nuclear`

The GenBankId GI:# number of the most similar mRNA/+RNA sequence to this one.
164692476

The mRNA/+RNA description:

Homo sapiens apoptotic protease activating factor 1 (Apaf-1) mRNA, complete cds. Similar to C. elegans cell
death gene ced-4.

The mRNA/+RNA sequence represented in the +DNA notation:

ACTTCTACGC GCGCGGGCAT GAGCCGTGGC AGGAGTGCGC GGCGGCAGCG GTGGCCGCCC CTTGGGGCTT GGGGTGTGTT TATTTGCATA AGCGGGCGCG 
^1         ^11        ^21        ^31        ^41        ^51        ^61        ^71        ^81        ^91        

CGCCGTCCGG GCTGGGTGGA TCCGGCGGGA TTTGACTGCT CCGCTGTCCA GAGGCGGAGA AGAAGAGGTA GCGAGTGGAC GTGACTGCTC TATCCCGGGC 
^101       ^111       ^121       ^131       ^141       ^151       ^161       ^171       ^181       ^191       

AAAAGGGATA GAACCAGAGG TGGGGAGTCT GGGCAGTCGG CGACCCGCGA AGACTTGAGG TGCCGCAGCG GCATCCGGAG TAGCGCCGGG CTCCCTCCGG 
^201       ^211       ^221       ^231       ^241       ^251       ^261       ^271       ^281       ^291       

GGTGCAGCCG CCGTCGGGGG AAGGGCGCCA CAGGCCGGGA AGACCTCCTC CCTTTGTGTC CAGTAGTGGG GTCCACCGGA GGGCGGCCCG TGGGCCGGGC 
^301       ^311       ^321       ^331       ^341       ^351       ^361       ^371       ^381       ^391       

CTCACCGCGG CGCTCCGGGA CTGTGGGGTC AGGCTGCGTT GGGTGGACGC CCACCTCGCC AACCTTCGGA GGTCCCTGGG GGTCTTCGTG CGCCCCGGGG 
^401       ^411       ^421       ^431       ^441       ^451       ^461       ^471       ^481       ^491       

CTGCAGAGAT CCAGGGGAGG CGCCTGTGAG GCCCGGACCT GCCCCGGGGC GAAGGGTATG TGGCGAGACA GAGCCCTGCA CCCCTAATTC CCGGTGGAAA 
^501       ^511       ^521       ^531       ^541       ^551       ^561       ^571       ^581       ^591       

ACTCCTGTTG CCGTTTCCCT CCACCGGCCT GGAGTCTCCC AGTCTTGTCC CGGCAGTGCC GCCCTCCCCA CTAAGACCTA GGCGCAAAGG CTTGGCTCAT 
^601       ^611       ^621       ^631       ^641       ^651       ^661       ^671       ^681       ^691       

GGTTGACAGC TCAGAGAGAG AAAGATCTGA GGGAAGATGG ATGCAAAAGC TCGAAATTGT TTGCTTCAAC ATAGAGAAGC TCTGGAAAAG GACATCAAGA 
^701       ^711       ^721       ^731       ^741       ^751       ^761       ^771       ^781       ^791       

CATCCTACAT CATGGATCAC ATGATTAGTG ATGGATTTTT AACAATATCA GAAGAGGAAA AAGTAAGAAA TGAGCCCACT CAACAGCAAA GAGCAGCTAT 
^801       ^811       ^821       ^831       ^841       ^851       ^861       ^871       ^881       ^891       

GCTGATTAAA ATGATACTTA AAAAAGATAA TGATTCCTAC GTATCATTCT ACAGTGCTCT ACTACATGAA GGATATAAAG ATCTTGCTGC CCTTCTCCAT 
^901       ^911       ^921       ^931       ^941       ^951       ^961       ^971       ^981       ^991       

GATGGCATTC CTGTTGTCTC TTCTTCCAGT GGTAAAGATT CAGTTAGTGG AATAACTTCG TATGTAAGGA CAGTCCTGTG TGAAGGTGGA GTACCACAGA 
^1001      ^1011      ^1021      ^1031      ^1041      ^1051      ^1061      ^1071      ^1081      ^1091      

GGCCAGTTGT TTTTGTCACA AGGAAGAAGC TGGTGAATGC AATTCAGCAG AAGCTCTCCA AATTGAAAGG TGAACCAGGA TGGGTCACCA TACATGGAAT 
^1101      ^1111      ^1121      ^1131      ^1141      ^1151      ^1161      ^1171      ^1181      ^1191      

GGCAGGCTGT GGGAAGTCTG TATTAGCTGC AGAAGCTGTT AGAGATCATT CCCTTTTAGA AGGTTGTTTC CCAGGGGGAG TGCATTGGGT TTCAGTTGGG 
^1201      ^1211      ^1221      ^1231      ^1241      ^1251      ^1261      ^1271      ^1281      ^1291      

AAACAAGACA AATCTGGGCT TCTGATGAAA CTGCAGAATC TTTGCACACG GTTGGATCAG GATGAGAGTT TTTCCCAGAG GCTTCCACTT AATATTGAAG 
^1301      ^1311      ^1321      ^1331      ^1341      ^1351      ^1361      ^1371      ^1381      ^1391      

AGGCTAAAGA CCGTCTCCGC ATTCTGATGC TTCGCAAACA CCCAAGGTCT CTCTTGATCT TGGATGATGT TTGGGACTCT TGGGTGTTGA AAGCTTTTGA 
^1401      ^1411      ^1421      ^1431      ^1441      ^1451      ^1461      ^1471      ^1481      ^1491      

CAGTCAGTGT CAGATTCTTC TTACAACCAG AGACAAGAGT GTTACAGATT CAGTAATGGG TCCTAAATAT GTAGTCCCTG TGGAGAGTTC CTTAGGAAAG 
^1501      ^1511      ^1521      ^1531      ^1541      ^1551      ^1561      ^1571      ^1581      ^1591      

GAAAAAGGAC TTGAAATTTT ATCCCTTTTT GTTAATATGA AGAAGGCAGA TTTGCCAGAA CAAGCTCATA GTATTATAAA AGAATGTAAA GGCTCTCCCC 
^1601      ^1611      ^1621      ^1631      ^1641      ^1651      ^1661      ^1671      ^1681      ^1691      

TTGTAGTATC TTTAATTGGT GCACTTTTAC GTGATTTTCC CAATCGCTGG GAGTACTACC TCAAACAGCT TCAGAATAAG CAGTTTAAGA GAATAAGGAA 
^1701      ^1711      ^1721      ^1731      ^1741      ^1751      ^1761      ^1771      ^1781      ^1791      

ATCTTCGTCT TATGATTATG AGGCTCTAGA TGAAGCCATG TCTATAAGTG TTGAAATGCT CAGAGAAGAC ATCAAAGATT ATTACACAGA TCTTTCCATC 
^1801      ^1811      ^1821      ^1831      ^1841      ^1851      ^1861      ^1871      ^1881      ^1891      

CTTCAGAAGG ACGTTAAGGT GCCTACAAAG GTGTTATGTA TTCTCTGGGA CATGGAAACT GAAGAAGTTG AAGACATACT GCAGGAGTTT GTAAATAAGT 
^1901      ^1911      ^1921      ^1931      ^1941      ^1951      ^1961      ^1971      ^1981      ^1991      

CTCTTTTATT CTGTGATCGG AATGGAAAGT CGTTTCGTTA TTATTTACAT GATCTTCAAG TAGATTTTCT TACAGAGAAG AATTGCAGCC AGCTTCAGGA 
^2001      ^2011      ^2021      ^2031      ^2041      ^2051      ^2061      ^2071      ^2081      ^2091      

TCTACATAAG AAGATAATCA CTCAGTTTCA GAGATATCAC CAGCCGCATA CTCTTTCACC AGATCAGGAA GACTGTATGT ATTGGTACAA CTTTCTGGCC 
^2101      ^2111      ^2121      ^2131      ^2141      ^2151      ^2161      ^2171      ^2181      ^2191      

TATCACATGG CCAGTGCCAA GATGCACAAG GAACTTTGTG CTTTAATGTT TTCCCTGGAT TGGATTAAAG CAAAAACAGA ACTTGTAGGC CCTGCTCATC 
^2201      ^2211      ^2221      ^2231      ^2241      ^2251      ^2261      ^2271      ^2281      ^2291      

TGATTCATGA ATTTGTGGAA TACAGACATA TACTAGATGA AAAGGATTGT GCAGTCAGTG AGAATTTTCA GGAGTTTTTA TCTTTAAATG GACACCTTCT 
^2301      ^2311      ^2321      ^2331      ^2341      ^2351      ^2361      ^2371      ^2381      ^2391      

TGGACGACAG CCATTTCCTA ATATTGTACA ACTGGGTCTC TGTGAGCCGG AAACTTCAGA AGTTTATCAG CAAGCTAAGC TGCAGGCCAA GCAGGAGGTC 
^2401      ^2411      ^2421      ^2431      ^2441      ^2451      ^2461      ^2471      ^2481      ^2491      

GATAATGGAA TGCTTTACCT GGAATGGATA AACAAAAAAA ACATCACGAA TCTTTCCCGC TTAGTTGTCC GCCCCCACAC AGATGCTGTT TACCATGCCT 
^2501      ^2511      ^2521      ^2531      ^2541      ^2551      ^2561      ^2571      ^2581      ^2591      

GCTTTTCTGA GGATGGTCAG AGAATAGCTT CTTGTGGAGC TGATAAAACC TTACAGGTGT TCAAAGCTGA AACAGGAGAG AAACTTCTAG AAATCAAGGC 
^2601      ^2611      ^2621      ^2631      ^2641      ^2651      ^2661      ^2671      ^2681      ^2691      

TCATGAGGAT GAAGTGCTTT GTTGTGCATT CTCTACAGAT GACAGATTTA TAGCAACCTG CTCAGTGGAT AAAAAAGTGA AGATTTGGAA TTCTATGACT 
^2701      ^2711      ^2721      ^2731      ^2741      ^2751      ^2761      ^2771      ^2781      ^2791      

GGGGAACTAG TACACACCTA TGATGAGCAC TCAGAGCAAG TCAATTGCTG CCATTTCACC AACAGTAGTC ATCATCTTCT CTTAGCCACT GGGTCAAGTG 
^2801      ^2811      ^2821      ^2831      ^2841      ^2851      ^2861      ^2871      ^2881      ^2891      

ACTGCTTCCT CAAACTTTGG GATTTGAATC AAAAAGAATG TCGAAATACC ATGTTTGGTC ATACAAATTC AGTCAATCAC TGCAGATTTT CACCAGATGA 
^2901      ^2911      ^2921      ^2931      ^2941      ^2951      ^2961      ^2971      ^2981      ^2991      

TAAGCTTTTG GCTAGTTGTT CAGCTGATGG AACCTTAAAG CTTTGGGATG CGACATCAGC AAATGAGAGG AAAAGCATTA ATGTGAAACA GTTCTTCCTA 
^3001      ^3011      ^3021      ^3031      ^3041      ^3051      ^3061      ^3071      ^3081      ^3091      

AATTTGGAGG ACCCTCAAGA GGATATGGAA GTGATAGTGA AGTGTTGTTC GTGGTCTGCT GATGGTGCAA GGATAATGGT GGCAGCAAAA AATAAAATCT 
^3101      ^3111      ^3121      ^3131      ^3141      ^3151      ^3161      ^3171      ^3181      ^3191      

TTTTGTGGAA TACAGACTCA CGTTCAAAGG TGGCTGATTG CAGAGGACAT TTAAGTTGGG TTCATGGTGT GATGTTTTCT CCTGATGGAT CATCATTTTT 
^3201      ^3211      ^3221      ^3231      ^3241      ^3251      ^3261      ^3271      ^3281      ^3291      

GACATCTTCT GATGACCAGA CAATCAGGCT CTGGGAGACA AAGAAAGTAT GTAAGAACTC TGCTGTAATG TTAAAGCAAG AAGTAGATGT TGTGTTTCAA 
^3301      ^3311      ^3321      ^3331      ^3341      ^3351      ^3361      ^3371      ^3381      ^3391      

GAAAATGAAG TGATGGTCCT TGCAGTTGAC CATATAAGAC GTCTGCAACT CATTAATGGA AGAACAGGTC AGATTGATTA TCTGACTGAA GCTCAAGTTA 
^3401      ^3411      ^3421      ^3431      ^3441      ^3451      ^3461      ^3471      ^3481      ^3491      

GCTGCTGTTG CTTAAGTCCA CATCTTCAGT ACATTGCATT TGGAGATGAA AATGGAGCCA TTGAGATTTT AGAACTTGTA AACAATAGAA TCTTCCAGTC 
^3501      ^3511      ^3521      ^3531      ^3541      ^3551      ^3561      ^3571      ^3581      ^3591      

CAGGTTTCAG CACAAGAAAA CTGTATGGCA CATCCAGTTC ACAGCCGATG AGAAGACTCT TATTTCAAGT TCTGATGATG CTGAAATTCA GGTATGGAAT 
^3601      ^3611      ^3621      ^3631      ^3641      ^3651      ^3661      ^3671      ^3681      ^3691      

TGGCAATTGG ACAAATGTAT CTTTCTACGA GGCCATCAGG AAACAGTGAA AGACTTTAGA CTCTTGAAAA ATTCAAGACT GCTTTCTTGG TCATTTGATG 
^3701      ^3711      ^3721      ^3731      ^3741      ^3751      ^3761      ^3771      ^3781      ^3791      

GAACAGTGAA GGTATGGAAT ATTATTACTG GAAATAAAGA AAAAGACTTT GTCTGTCACC AGGGTACAGT ACTTTCTTGT GACATTTCTC ACGATGCTAC 
^3801      ^3811      ^3821      ^3831      ^3841      ^3851      ^3861      ^3871      ^3881      ^3891      

CAAGTTTTCA TCTACCTCTG CTGACAAGAC TGCAAAGATC TGGAGTTTTG ATCTCCTTTT GCCACTTCAT GAATTGAGGG GCCACAACGG CTGTGTGCGC 
^3901      ^3911      ^3921      ^3931      ^3941      ^3951      ^3961      ^3971      ^3981      ^3991      

TGCTCTGCCT TCTCTGTGGA CAGTACCCTG CTGGCAACGG GAGATGACAA TGGAGAAATC AGGATATGGA ATGTCTCAAA CGGTGAGCTT CTTCATTTGT 
^4001      ^4011      ^4021      ^4031      ^4041      ^4051      ^4061      ^4071      ^4081      ^4091      

GTGCTCCGCT TTCAGAAGAA GGAGCTGCTA CCCATGGAGG CTGGGTGACT GACCTTTGCT TTTCTCCAGA TGGCAAAATG CTTATCTCTG CTGGAGGATA 
^4101      ^4111      ^4121      ^4131      ^4141      ^4151      ^4161      ^4171      ^4181      ^4191      

TATTAAGTGG TGGAACGTTG TCACTGGGGA ATCCTCACAG ACCTTCTACA CAAATGGAAC CAATCTTAAG AAAATACACG TGTCCCCTGA CTTCAAAACA 
^4201      ^4211      ^4221      ^4231      ^4241      ^4251      ^4261      ^4271      ^4281      ^4291      

TATGTGACTG TGGATAATCT TGGTATTTTA TATATTTTAC AGACTTTAGA ATAAAATAGT TAAGCATTAA TGTAGTTGAA CTTTTTAAAT TTTTGAATTG 
^4301      ^4311      ^4321      ^4331      ^4341      ^4351      ^4361      ^4371      ^4381      ^4391      

GAAAAAAATT CTAATGAAAC CCTGATATCA ACTTTTTATA AAGCTCTTAA TTGTTGTGCA GTATTGCATT CATTA
^4401      ^4411      ^4421      ^4431      ^4441      ^4451      ^4461      ^4471

Credibility of mRNA sequence:

only_fragment_published_or_from_author_and_the_rest_is_a_guess

The organism containing this mRNA with IRES segment in its genome:

Homo sapiens

A promoter reported in cDNA corresponding to IRES sequence:
unclear 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:
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:
heterogeneous_population_of_molecules_found

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 organism used:

Homo sapiens HeLa (ATCC CCL-2)

Notable Open-Reading Frames (ORFs; protein coding regions) in the mRNA/+RNA sequence:

ORF

ORF position:

1

Version: `3`	Last change: `2009-09-01 19:57:05`
Originaly submitted by: Martin Mokrejš	Reviewed by: Martin Mokrejš

The abbreviated name of this ORF/gene:
Apaf-1

The description of the protein encoded in this ORF:
apoptotic protease activating factor 1

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):

737-4354

Remarks:

The best really existing mRNA sequence in Genbank is AK307509 (GI:164692476) which has longer 5'-UTR by 159bp
(obtained by oligo-capping approach) compared to entry AF013263 (GI:2330014). The below mentioned 7kb long
sequence is a chimera of Apaf1 mRNA with another gene since position 5244:


>emb|CR603841.1| UniGene info full-length cDNA clone CS0DI077YE15 of Placenta Cot 25-normalized of Homo
sapiens Length=1596

AF013263  5244  GGTCTTGAACTCTTGGCCTCAAGTAATCCTCCTGCCTCAGCCTCCCAAAGTGTTGGGATT  5303
                ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Sbjct     1     GGTCTTGAACTCTTGGCCTCAAGTAATCCTCCTGCCTCAGCCTCCCAAAGTGTTGGGATT  60

[cut]

AF013263  6384  TCCTTAGAATACCCAAATCATAATTTTATTTGTACACACTGTTAGGGGCTCATCTCATGT  6443
                |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Sbjct     1141  TCCTTAGAATACCCAAATCATAATTTTATTTGTACACACTGTTAGGGGCTCATCTCATG-  1199

AF013263  6444  AGG  6446
                |||
Sbjct     1200  AGG  1202

[cut]

AF013263  6617  GAGGATTCCTTGAGCCCTGGAGTTTGAGTCCAGCCTGGGTGACATAGCAAGACCCTGTCT  6676
                ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Sbjct     1199  GAGGATTCCTTGAGCCCTGGAGTTTGAGTCCAGCCTGGGTGACATAGCAAGACCCTGTCT  1258

[cut]

AF013263  6977  gaatttaaaaaatttttgtaaaaataaaattcacaaaa  7014
                ||||||||||||||||||||||||||||||||||||||
Sbjct     1559  GAATTTAAAAAATTTTTGTAAAAATAAAATTCACAAAA  1596




The Apaf1-mRNA region in this chimeric message of AF013263 ends at the position 4283:

>gb|EF560718.1| UniGene infoGene info Homo sapiens clone DKFZp781B1145 APAF1 protein (APAF1) mRNA, complete
cds Length=3744

AF013263   573  GGAAGATGGATGCAAAAGCTCGAAATTGTTTGCTTCAACATAGAGAAGCTCTGGAAAAGG  632
                ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Sbjct      1    GGAAGATGGATGCAAAAGCTCGAAATTGTTTGCTTCAACATAGAGAAGCTCTGGAAAAGG  60

[cut]

AF013263  4231  TGATATCAACTTTTTATAAAGCTCTTAATTGTTGTGCAGTATTGCATTCATTA  4283
                |||||||||||||||||||||||||||||||||||||||||||||||||||||
Sbjct     3692  TGATATCAACTTTTTATAAAGCTCTTAATTGTTGTGCAGTATTGCATTCATTA  3744




Interestingly, several mRNAS incomplete at the 3'-end terminate just at position 4162 which is a STOP codon:

>emb|AJ243004.1| UniGene infoGene info Homo sapiens mRNA for apoptotic protease activating factor 1
Length=3618

[cut]

AF013263  4111  TGTGACTGTGGATAATCTTGGTATTTTATATATTTTACAGACTTTAGAATAA  4162
                ||||||||||||||||||||||||||||||||||||||||||||||||||||
Sbjct     3567  TGTGACTGTGGATAATCTTGGTATTTTATATATTTTACAGACTTTAGAATAA  3618



In conclusion, we have used sequence in AK307509 for the 5'-UTR and most of the CDS and the record EF560718
to get up to the 3'-end.


Several alternatively spliced messages exist but the splice junctions are in the CDS, thus not important to
the IRES story.




Integrity of the transcripts was verified by Northern blot using randomly primed cDNA probes against Fluc.
Shorter transcript fragment of 1.3kb was detected (also found in pRmycF transcripts containing c-myc). No more
sensitive testing of integrity of transcripts produced was done, e.g. RT-PCR of the in vivo produced
transcription products or direct mRNA transfection or tests based on a promoter-less plasmid. [Coldwell et
al., 2000]


It appears the very two 3'-most bases of the 5'-UTR of Apaf-1 were not cloned into the pRAF plasmid vector
because they had to be replaced by 'cc' bases of the cloning NcoI site 'ccATGg'. Please refer to the
annotation of pRAF plasmid (IRESiteID:341).

Cammas et al. (2007) confirmed either a splicing or cryptic promoter issue with the pRAF plasmid in DNA
transfections (Supplemental Figure S2) in RNAi based assay in which they were not able to inhibit the Apaf-1
IRES activity like of EMCV, HRV IRES. They repeated results of Sherril et al. (2004) showing that either a
splicing issue or a promoter is functional in HeLa cells when Bcl2 IRES is placed into the pRF vector.
Further, they show that in direct mRNA transfection Apaf-1 IRES activity is only 2x above the empty vector
control (capped, poly(A)-tailed messages) and thus conclude as not functional.

Citations:

Coldwell M. J., Mitchell S. A., Stoneley M., MacFarlane M., Willis A. E. (2000) Initiation of Apaf-1 translation by internal ribosome entry. Oncogene. 19(7):899-905

Cammas A., Pileur F., Bonnal S., Lewis S. M., Leveque N., Holcik M., Vagner S. (2007) Cytoplasmic relocalization of heterogeneous nuclear ribonucleoprotein A1 controls translation initiation of specific mRNAs. Mol. Biol. Cell. 18(12):5048-5059

IRESs:

IRES:

Version: `7`	Last change: `2009-09-01 19:57:05`
Originaly submitted by: Martin Mokrejš	Reviewed by: Martin Mokrejš
The IRES name: `Apaf-1`

The IRES absolute position (the range includes START and STOP codons or their equivalents):

504-734

Conclusion:
putative_IRES

How IRES boundaries were determined:
experimentally_determined

The sequence of IRES region aligned to its secondary structure (if available):

CAGAGATCCA GGGGAGGCGC CTGTGAGGCC CGGACCTGCC CCGGGGCGAA GGGTATGTGG CGAGACAGAG CCCTGCACCC CTAATTCCCG GTGGAAAACT 
)........( (((((((.(( ((((...((( ...((.(((( (((...)).. ))))).)))) )...)))).) ))))...))) ))..(((((. .........( 
^504       ^514       ^524       ^534       ^544       ^554       ^564       ^574       ^584       ^594       

CCTGTTGCCG TTTCCCTCCA CCGGCCTGGA GTCTCCCAGT CTTGTCCCGG CAGTGCCGCC CTCCCCACTA AGACCTAGGC GCAAAGGCTT GGCTCATGGT 
(..(.(.(.. ((((.(((.. (((((((... ((.((...(( (((((..... .......... ......)).) ))))...)). ))..))))). )).(((.... 
^604       ^614       ^624       ^634       ^644       ^654       ^664       ^674       ^684       ^694       

TGACAGCTCA GAGAGAGAAA GATCTGAGGG A
.......))) ..))).)))) )).).))))) )
^704       ^714       ^724       ^734

There is no Vienna RNA package installed on the server or some error/warning messages were output. Due to that maybe we cannot prepare 2D structures for display. The error/warning message was:

)........((((((((.((((((...(((...((.(((((((...))..))))).)))))...)))).)))))...)))))..(((((..........((..(.(.(..((((.(((..(((((((...((.((...(((((((.....................)).)))))...)).))..))))).)).(((...........)))..))).)))))).).))))))
ERROR: unbalanced brackets in make_pair_table

STDOUT was:

Remarks:

The -233 to -1 (due to the cloning approach possibly only -233 to -3) fragment retains 75% activity (Fig. 6B)
of the full-length 5'-UTR (1-577b). Apaf-1 IRES is functional in HeLa, HepG2 cells, lower activity was seen in
MCF7, HK293, COS7, MRC5 cells. No activity seen in SY5Y and Balb/c cells.

Citations:

Coldwell M. J., Mitchell S. A., Stoneley M., MacFarlane M., Willis A. E. (2000) Initiation of Apaf-1 translation by internal ribosome entry. Oncogene. 19(7):899-905

RNA:protein interactions:

The RNA:protein interaction:

Version: `0`
Originaly submitted by: Martin Mokrejš	Reviewed by: Martin Mokrejš

The description of the protein interacting with the RNA:
Heterogeneous nuclear ribonucleoprotein (hnRNP) A1

The organism where this RNA:protein interaction occurs:

Homo sapiens HeLa (ATCC CCL-2)

Remarks:

The interaction was confirmed by several methods using HeLa cell nuclear extracts (NE) with 32P-labeled RNAs.
Data in Figure 1.

Citations:

Regions with experimentally determined secondary structures:

A region with the experimentally determined secondary structure:

IRESite 2D Struct Id: 26

Version: `1`	Last change: `2009-09-01 19:57:05`
Originaly submitted by: Václav Vopálenský	Reviewed by: Václav Vopálenský
The function of the 2D structure: `IRES`	The 2D structure causes frameshift: `no`

The absolute position of the experimentally mapped region (the range includes START and STOP codons or their equivalents):
160-739

The underlying nucleic acid sequence and structure of the mapped region:

AAGAAGAGGT AGCGAGTGGA CGTGACTGCT CTATCCCGGG CAAAAGGGAT AGAACCAGAG GTGGGGAGTC TGGGCAGTCG GCGACCCGCG AAGACTTGAG 
......(((( ..((...((. (((((((((( (((((((... .....))))) ))).(((((. ........)) )))))))))) .)).))..)) ...))))... 
^160       ^170       ^180       ^190       ^200       ^210       ^220       ^230       ^240       ^250       

GTGCCGCAGC GGCATCCGGA GTAGCGCCGG GCTCCCTCCG GGGTGCAGCC GCCGTCGGGG GAAGGGCGCC ACAGGCCGGG AAGACCTCCT CCCTTTGTGT 
.....((((( ....(((((. ((.(((((.. ..((((((.( .((((....) ))).).)))) ))..))))). )).(((((.( (((((((((( (((...(((. 
^260       ^270       ^280       ^290       ^300       ^310       ^320       ^330       ^340       ^350       

CCAGTAGTGG GGTCCACCGG AGGGCGGCCC GTGGGCCGGG CCTCACCGCG GCGCTCCGGG ACTGTGGGGT CAGGCTGCGT TGGGTGGACG CCCACCTCGC 
((.((.(((( (((((.((.. ..)).((((. ...))))))) )))))).))) ))))...))) )....((((( ((((.((.(( .((((((... .)))))).)) 
^360       ^370       ^380       ^390       ^400       ^410       ^420       ^430       ^440       ^450       

CAACCTTCGG AGGTCCCTGG GGGTCTTCGT GCGCCCCGGG GCTGCAGAGA TCCAGGGGAG GCGCCTGTGA GGCCCGGACC TGCCCCGGGG CGAAGGGTAT 
)).))).... .)))))).)) )))))))).) ).)))))))) )))))..... ...((((((( (.((((((.. .(((...((. (((((((... ))..))))). 
^460       ^470       ^480       ^490       ^500       ^510       ^520       ^530       ^540       ^550       

GTGGCGAGAC AGAGCCCTGC ACCCCTAATT CCCGGTGGAA AACTCCTGTT GCCGTTTCCC TCCACCGGCC TGGAGTCTCC CAGTCTTGTC CCGGCAGTGC 
)))))...)) )).))))).. .)))))..(( (((....... ...((..(.( .(..((((.( ((..(((((( (...((.((. ..(((((((. .......... 
^560       ^570       ^580       ^590       ^600       ^610       ^620       ^630       ^640       ^650       

CGCCCTCCCC ACTAAGACCT AGGCGCAAAG GCTTGGCTCA TGGTTGACAG CTCAGAGAGA GAAAGATCTG AGGGAAGATG
.......... )).))))).. .)).))..)) ))).)).((( .......... .)))..))). )))))).).) ))))))....
^660       ^670       ^680       ^690       ^700       ^710       ^720       ^730

There is no Vienna RNA package installed on the server or some error/warning messages were output. Due to that maybe we cannot prepare 2D structures for display. The error/warning message was:

WARNING: bases 539 and 553 (CC) can't pair!

STDOUT was:

AAGAAGAGGUAGCGAGUGGACGUGACUGCUCUAUCCCGGGCAAAAGGGAUAGAACCAGAGGUGGGGAGUCUGGGCAGUCGGCGACCCGCGAAGACUUGAGGUGCCGCAGCGGCAUCCGGAGUAGCGCCGGGCUCCCUCCGGGGUGCAGCCGCCGUCGGGGGAAGGGCGCCACAGGCCGGGAAGACCUCCUCCCUUUGUGUCCAGUAGUGGGGUCCACCGGAGGGCGGCCCGUGGGCCGGGCCUCACCGCGGCGCUCCGGGACUGUGGGGUCAGGCUGCGUUGGGUGGACGCCCACCUCGCCAACCUUCGGAGGUCCCUGGGGGUCUUCGUGCGCCCCGGGGCUGCAGAGAUCCAGGGGAGGCGCCUGUGAGGCCCGGACCUGCCCCGGGGCGAAGGGUAUGUGGCGAGACAGAGCCCUGCACCCCUAAUUCCCGGUGGAAAACUCCUGUUGCCGUUUCCCUCCACCGGCCUGGAGUCUCCCAGUCUUGUCCCGGCAGUGCCGCCCUCCCCACUAAGACCUAGGCGCAAAGGCUUGGCUCAUGGUUGACAGCUCAGAGAGAGAAAGAUCUGAGGGAAGAUG
......((((..((...((.(((((((((((((((((........)))))))).(((((.........)))))))))))).)).))..))...))))........(((((....(((((.((.(((((....((((((.(.((((....)))).).))))))..))))).)).(((((.((((((((((((((...(((.((.((.(((((((((.((....)).((((....))))))))))))).)))))))...))))....(((((((((.((.((.((((((....)))))).)))).))).....)))))).)))))))))).)).)))))))))))))........((((((((.((((((...(((...((.(((((((...))..))))).)))))...)))).)))))...)))))..(((((..........((..(.(.(..((((.(((..(((((((...((.((...(((((((.....................)).)))))...)).))..))))).)).(((...........)))..))).)))))).).))))))).... (-194.30)

Rendering structure of Apaf-1 mRNA 580 nt long with energy of -194.30 kcal/mol as calculated by RNAeval using VARNA Java applet with some IRESite improvements (see VARNA modified by IRESite). Hold left mouse button to move structure parts, hold right mouse button to move whole structure, use mouse wheel to zoom. Right mouse-click opens a menu to export into JPG/SVG and many other options.

You need a Java-enabled browser so that modified varsion of VARNA could be started. See http://www.iresite.org/VARNA/ for more details.

Remarks:

Structure from Fig. 4E.

4.1.1. Enzymes used to characterize at least partially the 2D structure.

Enzyme or a combination of enzymes used in a single experiment with respective buffer:

ss_experiment_with_enzyme_id: 48

The temperature (in degrees of Celsia):
22 The enzymatic method used to determine the 2D structure:
ribonuclease V1

Enzyme or a combination of enzymes used in a single experiment with respective buffer:

Version: 0

pH `7.00`	Li⁺ [mM] `0`	Na⁺ [mM] `0`	K⁺ [mM] `100.00`	Mg²⁺ [mM] `10.00`	Ca²⁺ [mM] `0`
Cl^- [mM] `120.00`	Tris [mM] `10.00`	BSA [mM] `0`	HEPES [mM] `0`	EGTA [mM] `0`	EDTA [mM] `0`
cacodylate [mM] `0`

4.1.2. Chemicals used to characterize at least partially the 2D structure.

Chemical reagent used with its respective buffer:

ss_experiment_with_chemical_id: 20

The temperature (in degrees of Celsia):
0 The chemical reagent used to determine the 2D structure:
DMS

Chemical reagent used with its respective buffer:

Version: 0

pH `7.00`	Li⁺ [mM] `0`	Na⁺ [mM] `0`	K⁺ [mM] `100.00`	Mg²⁺ [mM] `10.00`	Ca²⁺ [mM] `0`
Cl^- [mM] `100.00`	Tris [mM] `10.00`	BSA [mM] `0`	HEPES [mM] `0`	EGTA [mM] `0`	EDTA [mM] `0`
cacodylate [mM] `0`

Other buffer components and their relative concentrations:

20mM acetate (CH3COO)-

Chemical reagent used with its respective buffer:

ss_experiment_with_chemical_id: 21

The temperature (in degrees of Celsia):
0 The chemical reagent used to determine the 2D structure:
kethoxal

Chemical reagent used with its respective buffer:

Version: 0

pH `7.00`	Li⁺ [mM] `0`	Na⁺ [mM] `0`	K⁺ [mM] `100.00`	Mg²⁺ [mM] `10.00`	Ca²⁺ [mM] `0`
Cl^- [mM] `100.00`	Tris [mM] `10.00`	BSA [mM] `0`	HEPES [mM] `0`	EGTA [mM] `0`	EDTA [mM] `0`
cacodylate [mM] `0`

Other buffer components and their relative concentrations:

20mM acetate (CH3COO)-

Citations:

Mitchell S. A., Spriggs K. A., Coldwell M. J., Jackson R. J., Willis A. E. (2003) The Apaf-1 internal ribosome entry segment attains the correct structural conformation for function via interactions with PTB and unr. Mol. Cell. 11(3):757-771

Last change to the database: 2019-03-18 09:32:49 GMT+1