Ittig, D.; Liu, Shi-Xia; Renneberg, Dorte; Schümperli, Daniel; Leumann, Christian (2004). Nuclear antisense effects in cyclophilin A pre-mRNA splicing by oligonucleotides: A comparison of tricyclo-DNA with LNA. Nucleic acids research, 32(1), pp. 346-353. Oxford University Press 10.1093/nar/gkh187
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The nuclear antisense properties of a series of tricyclo(tc)-DNA oligonucleotide 9-15mers, targeted against the 3' and 5' splice sites of exon 4 of cyclophilin A (CyPA) pre-mRNA, were evaluated in HeLa cells and compared with those of corresponding LNA-oligonucleotides. While the 9mers showed no significant antisense effect, the 11-15mers induced exon 4 skipping and exon 3+4 double skipping to about an equal extent upon lipofectamine mediated transfection in a sequence and dose dependent manner, as revealed by a RT-PCR assay. The antisense efficacy of the tc-oligonucleotides was found to be superior to that of the LNA-oligonucleotides in all cases by a factor of at least 4-5. A tc-oligonucleotide 15mer completely abolished CyPA mRNA production at 0.2‘M concentration. The antisense effect was confirmed by western blot analysis which revealed a reduction of CyPA protein to 13% of its normal level. Fluorescence microscopic investigations with a fluorescein labeled tc-15mer revealed a strong propensity for homogeneous nuclear localization of this backbone type after lipofectamine mediated transfection, while the corresponding lna 15mer showed a less clear cellular distribution pattern. Transfection without lipid carrier showed no significant internalization of both tc- and LNA-oligonucleotides. The obtained results confirm the power of tricyclo-DNA for nuclear antisense applications. Morover, CyPA may become an interesting therapeutic target due to its important role in the early steps of the viral replication of HIV-1.
Item Type: |
Journal Article (Original Article) |
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Division/Institute: |
08 Faculty of Science > Department of Chemistry, Biochemistry and Pharmaceutical Sciences (DCBP) 08 Faculty of Science > Department of Biology > Institute of Cell Biology |
UniBE Contributor: |
Liu, Shi-Xia, Renneberg, Dorte, Schümperli, Daniel, Leumann, Christian |
Subjects: |
500 Science > 570 Life sciences; biology 500 Science > 540 Chemistry |
ISSN: |
0305-1048 |
Publisher: |
Oxford University Press |
Language: |
English |
Submitter: |
Christian Leumann |
Date Deposited: |
16 Jun 2014 11:08 |
Last Modified: |
05 Dec 2022 14:34 |
Publisher DOI: |
10.1093/nar/gkh187 |
Uncontrolled Keywords: |
analysis Antisense backbone Cells cellular Cyclophilin A distribution double Fluorescein fluorescence HeLa Hela Cells Hiv-1 LNA oligonucleotide Oligonucleotides properties Protein reduction REPLICATION site Transfection tricyclo-DNA |
BORIS DOI: |
10.7892/boris.53215 |
URI: |
https://boris.unibe.ch/id/eprint/53215 |
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