Transcription factor decoy oligonucleotides modified with locked nucleic acids: an in vitro study to reconcile biostability with binding affinity
Identifieur interne : 000214 ( France/Analysis ); précédent : 000213; suivant : 000215Transcription factor decoy oligonucleotides modified with locked nucleic acids: an in vitro study to reconcile biostability with binding affinity
Auteurs : Rita Crinelli ; Marzia Bianchi ; Lucia Gentilini ; Linda Palma ; Mads D. S Rensen [Danemark] ; Torsten Bryld [Danemark] ; Ravindra B. Babu [Danemark] ; Khalil Arar [France] ; Jesper Wengel [Danemark] ; Mauro Magnani [Italie]Source :
- Nucleic Acids Research [ 0305-1048 ] ; 2004.
Descripteurs français
- KwdFr :
- Cellules HeLa, Endonucleases (métabolisme), Exonucleases (métabolisme), Facteur de transcription NF-kappa B (métabolisme), Facteurs de transcription (métabolisme), Humains, Oligonucléotides, Oligonucléotides antisens (), Oligonucléotides antisens (métabolisme), Sites de fixation, Séquence consensus, Séquence nucléotidique.
- MESH :
English descriptors
- KwdEn :
- MESH :
- chemical , chemistry : Oligonucleotides, Antisense.
- chemical , metabolism : Endonucleases, Exonucleases, NF-kappa B, Oligonucleotides, Antisense, Transcription Factors.
- Base Sequence, Binding Sites, Consensus Sequence, HeLa Cells, Humans, Oligonucleotides.
Abstract
Double‐stranded oligonucleotides (ODNs) containing the consensus binding sequence of a transcription factor provide a rationally designed tool to manipulate gene expression at the transcriptional level by the decoy approach. However, modifications introduced into oligonucleotides to increase stability quite often do not guarantee that transcription factor affinity and/or specificity of recognition are retained. We have previously evaluated the use of locked nucleic acids (LNA) in the design of decoy molecules for the transcription factor κB. Oligo nucleotides containing LNA substitutions displayed high resistance to exo‐ and endonucleolytic degradation, with LNA–DNA mix‐mers being more stable than LNA–DNA–LNA gap‐mers. However, insertion of internal LNA bases resulted in a loss of affinity for the transcription factor. This latter effect apparently depended on positioning of the internal LNA substitutions. Indeed, here we demonstrate that intra‐ and inter‐strand positioning of internal LNAs has to be carefully considered to maintain affinity and achieve high stability, respectively. Unfortun ately, our data also indicate that LNA positioning is not the only parameter affecting transcription factor binding, the interference in part being dependent on the intrinsic conformational properties of this nucleotide analog. To circumvent this problem, the successful use of an α‐l‐ribo‐ configured LNA is demonstrated, indicating LNA–DNA–α‐l‐LNA molecules as promising new decoy agents.
Url:
DOI: 10.1093/nar/gkh503
Affiliations:
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Acids Res.</title><idno type="ISSN">0305-1048</idno><idno type="eISSN">1362-4962</idno><imprint><publisher>Oxford University Press</publisher><date type="published">2004</date><biblScope unit="vol">32</biblScope><biblScope unit="issue">6</biblScope><biblScope unit="page" from="1874">1874</biblScope><biblScope unit="page" to="1885">1885</biblScope></imprint><idno type="ISSN">0305-1048</idno></series></biblStruct></sourceDesc><seriesStmt><idno type="ISSN">0305-1048</idno></seriesStmt></fileDesc><profileDesc><textClass><keywords scheme="KwdEn" xml:lang="en"><term>Base Sequence</term><term>Binding Sites</term><term>Consensus Sequence</term><term>Endonucleases (metabolism)</term><term>Exonucleases (metabolism)</term><term>HeLa Cells</term><term>Humans</term><term>NF-kappa B (metabolism)</term><term>Oligonucleotides</term><term>Oligonucleotides, Antisense (chemistry)</term><term>Oligonucleotides, Antisense (metabolism)</term><term>Transcription Factors (metabolism)</term></keywords><keywords scheme="KwdFr" xml:lang="fr"><term>Cellules HeLa</term><term>Endonucleases (métabolisme)</term><term>Exonucleases (métabolisme)</term><term>Facteur de transcription NF-kappa B (métabolisme)</term><term>Facteurs de transcription (métabolisme)</term><term>Humains</term><term>Oligonucléotides</term><term>Oligonucléotides antisens ()</term><term>Oligonucléotides antisens (métabolisme)</term><term>Sites de fixation</term><term>Séquence consensus</term><term>Séquence nucléotidique</term></keywords><keywords scheme="MESH" type="chemical" qualifier="chemistry" xml:lang="en"><term>Oligonucleotides, Antisense</term></keywords><keywords scheme="MESH" type="chemical" qualifier="metabolism" xml:lang="en"><term>Endonucleases</term><term>Exonucleases</term><term>NF-kappa B</term><term>Oligonucleotides, Antisense</term><term>Transcription Factors</term></keywords><keywords scheme="MESH" qualifier="métabolisme" xml:lang="fr"><term>Endonucleases</term><term>Exonucleases</term><term>Facteur de transcription NF-kappa B</term><term>Facteurs de transcription</term><term>Oligonucléotides antisens</term></keywords><keywords scheme="MESH" xml:lang="en"><term>Base Sequence</term><term>Binding Sites</term><term>Consensus Sequence</term><term>HeLa Cells</term><term>Humans</term><term>Oligonucleotides</term></keywords><keywords scheme="MESH" xml:lang="fr"><term>Cellules HeLa</term><term>Humains</term><term>Oligonucléotides</term><term>Oligonucléotides antisens</term><term>Sites de fixation</term><term>Séquence consensus</term><term>Séquence nucléotidique</term></keywords></textClass></profileDesc></teiHeader><front><div type="abstract" xml:lang="en">Double‐stranded oligonucleotides (ODNs) containing the consensus binding sequence of a transcription factor provide a rationally designed tool to manipulate gene expression at the transcriptional level by the decoy approach. However, modifications introduced into oligonucleotides to increase stability quite often do not guarantee that transcription factor affinity and/or specificity of recognition are retained. We have previously evaluated the use of locked nucleic acids (LNA) in the design of decoy molecules for the transcription factor κB. Oligo nucleotides containing LNA substitutions displayed high resistance to exo‐ and endonucleolytic degradation, with LNA–DNA mix‐mers being more stable than LNA–DNA–LNA gap‐mers. However, insertion of internal LNA bases resulted in a loss of affinity for the transcription factor. This latter effect apparently depended on positioning of the internal LNA substitutions. Indeed, here we demonstrate that intra‐ and inter‐strand positioning of internal LNAs has to be carefully considered to maintain affinity and achieve high stability, respectively. Unfortun ately, our data also indicate that LNA positioning is not the only parameter affecting transcription factor binding, the interference in part being dependent on the intrinsic conformational properties of this nucleotide analog. To circumvent this problem, the successful use of an α‐l‐ribo‐ configured LNA is demonstrated, indicating LNA–DNA–α‐l‐LNA molecules as promising new decoy agents.</div></front></TEI><affiliations><list><country><li>Danemark</li><li>France</li><li>Italie</li></country><region><li>Île-de-France</li></region><settlement><li>Paris</li></settlement></list><tree><noCountry><name sortKey="Bianchi, Marzia" sort="Bianchi, Marzia" uniqKey="Bianchi M" first="Marzia" last="Bianchi">Marzia Bianchi</name><name sortKey="Crinelli, Rita" sort="Crinelli, Rita" uniqKey="Crinelli R" first="Rita" last="Crinelli">Rita Crinelli</name><name sortKey="Gentilini, Lucia" sort="Gentilini, Lucia" uniqKey="Gentilini L" first="Lucia" last="Gentilini">Lucia Gentilini</name><name sortKey="Palma, Linda" sort="Palma, Linda" uniqKey="Palma L" first="Linda" last="Palma">Linda Palma</name></noCountry><country name="Danemark"><noRegion><name sortKey="S Rensen, Mads D" sort="S Rensen, Mads D" uniqKey="S Rensen M" first="Mads D." last="S Rensen">Mads D. S Rensen</name></noRegion><name sortKey="Babu, Ravindra B" sort="Babu, Ravindra B" uniqKey="Babu R" first="Ravindra B." last="Babu">Ravindra B. Babu</name><name sortKey="Bryld, Torsten" sort="Bryld, Torsten" uniqKey="Bryld T" first="Torsten" last="Bryld">Torsten Bryld</name><name sortKey="Wengel, Jesper" sort="Wengel, Jesper" uniqKey="Wengel J" first="Jesper" last="Wengel">Jesper Wengel</name></country><country name="France"><region name="Île-de-France"><name sortKey="Arar, Khalil" sort="Arar, Khalil" uniqKey="Arar K" first="Khalil" last="Arar">Khalil Arar</name></region></country><country name="Italie"><noRegion><name sortKey="Magnani, Mauro" sort="Magnani, Mauro" uniqKey="Magnani M" first="Mauro" last="Magnani">Mauro Magnani</name></noRegion></country></tree></affiliations></record>Pour manipuler ce document sous Unix (Dilib)
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