IRESite_Id:491 summary record describing characteristics of AQP4 IRES

IRESite: The database of experimentally verified IRES structures

The nucleic acid data:

IRESite Id: `491`	Version: `5`
Originaly submitted by: Václav Vopálenský
Reviewed by: Martin Mokrejš	Last change: `2009-10-10 02:09:04`

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: `AQP4`	The genetic origin of this natural mRNA/+RNA: `nuclear`

The GenBankId GI:# number of the most similar mRNA/+RNA sequence to this one.
1072052

Synonyms of the gene name:

Synonym: MIWC

Synonym: HMIWC2

Synonym: MGC22454

The mRNA/+RNA description:

Human mercurial-insensitive water channel mRNA, form 1, complete cds.

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

cctttctagG GACAGTTTGG ATAATTTTCC GTGAAACTGG ATGATCTTTG AATATTAAAT GAAAGGAGAC TAGAATAACA GTCTCTTGAC TATTCATGAA 
^1         ^11        ^21        ^31        ^41        ^51        ^61        ^71        ^81        ^91        

GGAGCTTTAG CAGGAGCATT CTTTCTTGGT GTGCAAATCA CGCCTGGCAG CCCCTGCCCA CAGTACCCGA AGATGGCAGG CCTTGTTCCC TTCACCTAAA 
^101       ^111       ^121       ^131       ^141       ^151       ^161       ^171       ^181       ^191       

TTCATAAACC TGGGTGTAGT GGCTTCTGAT GCTGATTTGT TTCTCTTTTC AGTAAGTGTG GACCTTTGTG TACCAGAGAG AACATCATGG TGGctttcaa 
^201       ^211       ^221       ^231       ^241       ^251       ^261       ^271       ^281       ^291       

aggggtctgg actcaagctt tctggaaagc agtcacagcg gaatttctgg ccatgcttat ttttgttctc ctcagcctgg gatccaccat caactggggt 
^301       ^311       ^321       ^331       ^341       ^351       ^361       ^371       ^381       ^391       

ggaacagaaa agcctttacc ggtcgacatg gtgctcatct ccctttgctt tggactcagc attgcaacca tggtgcagtg ctttggccat atcagcggtg 
^401       ^411       ^421       ^431       ^441       ^451       ^461       ^471       ^481       ^491       

gccacatcaa ccctgcagtg actgtggcca tggtgtgcac caggaagatc agcatcgcca agtctgtctt ctacatcgca gcccagtgcc tgggggccat 
^501       ^511       ^521       ^531       ^541       ^551       ^561       ^571       ^581       ^591       

cattggagca ggaatcctct atctggtcac acctcccagt gtggtgggag gcctgggagt caccatggtt catggaaatc ttaccgctgg tcatggtctc 
^601       ^611       ^621       ^631       ^641       ^651       ^661       ^671       ^681       ^691       

ctggttgagt tgataatcac atttcaattg gtgtttacta tctttgccag ctgtgattcc aaacggactg atgtcactgg ctcaatagct ttagcaattg 
^701       ^711       ^721       ^731       ^741       ^751       ^761       ^771       ^781       ^791       

ggttttctgt tgcaattgga catttatttg caatcaatta tactggtgcc agcatgaatc ccgcccgatc ctttggacct gcagttatca tgggaaattg 
^801       ^811       ^821       ^831       ^841       ^851       ^861       ^871       ^881       ^891       

ggaaaaccat tggatatatt gggttgggcc catcatagga gctgtcctcg ctggtgccct ttatgagtat gtcttctgtc cagatgttga attcaaacgt 
^901       ^911       ^921       ^931       ^941       ^951       ^961       ^971       ^981       ^991       

cgttttaaag aagccttcag caaagctgcc cagcaaacaa aaggaagcta catggaggtg gaggacaaca ggagtcaggc aaagacggat gacctgattc 
^1001      ^1011      ^1021      ^1031      ^1041      ^1051      ^1061      ^1071      ^1081      ^1091      

taaaacttgg agtggtgcat gtgattgacg ttgaccgggg agaggagaag aaggggaaag accaatctgg agaggtattg tcttcagtat gactagaaga 
^1101      ^1111      ^1121      ^1131      ^1141      ^1151      ^1161      ^1171      ^1181      ^1191      

tcgcactgaa agccagacaa gactccttag aactgtcctc agatttcctt ccgcccatta aggaaacaga tttgttataa attagaatgt gcaggtttgt 
^1201      ^1211      ^1221      ^1231      ^1241      ^1251      ^1261      ^1271      ^1281      ^1291      

tgtttcatgt catattactc agtctaaaca ataaatattt cataatttac aaggaggaac ggaagaaacc tattgtgaat tccaaatcta aaaaaagaaa 
^1301      ^1311      ^1321      ^1331      ^1341      ^1351      ^1361      ^1371      ^1381      ^1391      

tatttttaaa atgttcttaa gcaaatatat acctatttta tctagttacc tttcattaac aaccaatttt aaccgtgtgt caagatttgg ttaagtcttg 
^1401      ^1411      ^1421      ^1431      ^1441      ^1451      ^1461      ^1471      ^1481      ^1491      

cctgacagaa ctcaaagaca gctctatcag cttattcctt ctctactgga atattggtat agtcaattct attgaatatt atccttttta atgcccaagg 
^1501      ^1511      ^1521      ^1531      ^1541      ^1551      ^1561      ^1571      ^1581      ^1591      

taatgtggac taaagcccag aaatttgaaa agaatattca gaaatccttc ccaaatcata agggccc
^1601      ^1611      ^1621      ^1631      ^1641      ^1651      ^1661

Credibility of mRNA sequence:

end-to-end_sequence_reverse_engineered_and_should_match_experiment

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

Homo sapiens HEK 293T/17 (ATCC CRL-11268)

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:
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):
homogeneous_population_of_molecules_confirmed

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

The organism used:

Homo sapiens HEK293 (ATCC CRL-1573)

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

ORF

ORF position:

1

Version: `2`	Last change: `2008-07-16 23:11:53`
Originaly submitted by: Václav Vopálenský	Reviewed by: Martin Mokrejš

The abbreviated name of this ORF/gene:
AQP4

The description of the protein encoded in this ORF:
mercurial-insensitive water channel

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):

287-1192

Remarks:

For creation of this IRESite record we have used AQP4 5'-UTR used through experiments (as kindly provided by
Dr. Martin Holcik) and merged it on its right with ORF and 3'-UTR from mRNA sequence of AQP4 (GI:1072052).
Holcik et al. (2007) actually amplified the AQP4 5'-UTR sequence from HEK 293T/17 (ATCC CRL-11268) cells.
There are few single nucleotide insertions/deletions between the two sequences. Other variants of AQP4
messages lack the leftmost cca 251 bp while first exon is within first 669bp.

Citations:

Baird S. D., Lewis S. M., Turcotte M., Holcik M. (2007) A search for structurally similar cellular internal ribosome entry sites. Nucleic. Acids. Res. 35(14):4664-4677

IRESs:

IRES:

Version: `1`	Last change: `2008-07-16 23:11:53`
Originaly submitted by: Václav Vopálenský	Reviewed by: Martin Mokrejš
The IRES name: `AQP4`

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

10-293

Conclusion:
weakly_supported_IRES

How IRES boundaries were determined:
guessed

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

GGACAGTTTG GATAATTTTC CGTGAAACTG GATGATCTTT GAATATTAAA TGAAAGGAGA CTAGAATAAC AGTCTCTTGA CTATTCATGA AGGAGCTTTA 
((.((((((( (........) )...)))))) ....(((((( (......... ......(((( ((........ )))))).((( ....))).(( (((((((((. 
^10        ^20        ^30        ^40        ^50        ^60        ^70        ^80        ^90        ^100       

GCAGGAGCAT TCTTTCTTGG TGTGCAAATC ACGCCTGGCA GCCCCTGCCC ACAGTACCCG AAGATGGCAG GCCTTGTTCC CTTCACCTAA ATTCATAAAC 
...))))).. ))))))..(( ((((.....) ))))).((.. ...))..... ........)) ))))).(((. ....)))..) )))((((((. .......... 
^110       ^120       ^130       ^140       ^150       ^160       ^170       ^180       ^190       ^200       

CTGGGTGTAG TGGCTTCTGA TGCTGATTTG TTTCTCTTTT CAGTAAGTGT GGACCTTTGT GTACCAGAGA GAACATCATG GTGG
.))))))..( ((((...... .)).(((((( ((.(((.... .......... .......... ........)) )))))..... ....
^210       ^220       ^230       ^240       ^250       ^260       ^270       ^280       ^290

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:

AQP4 IRES represents almost the full length 5' UTR from human AQP4 mRNA (from nt -275 to nt -1 of the original
GenBank sequence) and the first 7 nts (+1 to +7) from AQP4 ORF.

Citations:

Baird S. D., Lewis S. M., Turcotte M., Holcik M. (2007) A search for structurally similar cellular internal ribosome entry sites. Nucleic. Acids. Res. 35(14):4664-4677

IRES trans-acting factor (ITAFS):

IRES trans-acting factor (ITAF):

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

Type of the interaction between ITAF and the RNA subject to translation:
direct_interaction_with_rna

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

3.1.2. 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_but_available_internally

Method used to demonstrate ITAF effect:
in_vitro In vitro system used to demonstrate ITAF effect:
other

Remarks:

HEK 293T cell extracts were used to demonstrate in vitro binding to AQP4 RNA. Data from Figure 8A.

Citations:

Baird S. D., Lewis S. M., Turcotte M., Holcik M. (2007) A search for structurally similar cellular internal ribosome entry sites. Nucleic. Acids. Res. 35(14):4664-4677

IRES trans-acting factor (ITAF):

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

Type of the interaction between ITAF and the RNA subject to translation:
direct_interaction_with_rna

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).

3.2.2. 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:
both In vitro system used to demonstrate ITAF effect:
other

The organism where action of this ITAF was studied:

Homo sapiens HEK 293T/17 (ATCC CRL-11268)

Remarks:

Deletion of the AQP4 5' UTR polypyrimidine tract causes a loss of PTB binding and a complete loss of IRES
activity, but does not disrupt the binding of other ITAFs.
Overexpression of PTB enhances AQP4 IRES activity approximately 2-fold.

HEK 293T cell extracts were used to demonstrate in vitro binding to AQP4 RNA. Data from Figure 8A/B/C.

Citations:

Baird S. D., Lewis S. M., Turcotte M., Holcik M. (2007) A search for structurally similar cellular internal ribosome entry sites. Nucleic. Acids. Res. 35(14):4664-4677

Regions with experimentally determined secondary structures:

A region with the experimentally determined secondary structure:

IRESite 2D Struct Id: 24

Version: `0`
Originaly submitted by: Václav Vopálenský	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):
10-295

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

GGACAGTTTG GATAATTTTC CGTGAAACTG GATGATCTTT GAATATTAAA TGAAAGGAGA CTAGAATAAC AGTCTCTTGA CTATTCATGA AGGAGCTTTA 
((.((((((( (........) )...)))))) ....(((((( (......... ......(((( ((........ )))))).((( ....))).(( (((((((((. 
^10        ^20        ^30        ^40        ^50        ^60        ^70        ^80        ^90        ^100       

GCAGGAGCAT TCTTTCTTGG TGTGCAAATC ACGCCTGGCA GCCCCTGCCC ACAGTACCCG AAGATGGCAG GCCTTGTTCC CTTCACCTAA ATTCATAAAC 
...))))).. ))))))..(( ((((.....) ))))).((.. ...))..... ........)) ))))).(((. ....)))..) )))((((((. .......... 
^110       ^120       ^130       ^140       ^150       ^160       ^170       ^180       ^190       ^200       

CTGGGTGTAG TGGCTTCTGA TGCTGATTTG TTTCTCTTTT CAGTAAGTGT GGACCTTTGT GTACCAGAGA GAACATCATG GTGGct
.))))))..( ((((...... .)).(((((( ((.(((.... .......... .......... ........)) )))))..... ......
^210       ^220       ^230       ^240       ^250       ^260       ^270       ^280       ^290

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:

2D structure of AQP4 from Fig 5B.

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