Peptide nucleic acids directed to the promoter of the alpha-chain of the interleukin-2 receptor.
Two 10-mer oligopyrimidine peptide nucleic acids (PNAs) were designed to interfere with IL-2R alpha promoter expression by binding to the regulatory sequences overlapping SRF and NF-kappa B transcription factor sites. Specific complexes were formed on each target sequence, and clearly involved (1) Hoogsteen hydrogen bonds as shown by experiments in which the purine strand of a single or double-stranded target was substituted with 7-deazadeoxyguanosine, (2) P-loop formation on double-helical DNA as evidenced by susceptibility to a single-strand-specific nuclease. When formed on a single-stranded DNA target, these highly stable complexes were responsible for efficient physical blockage of T7 DNA polymerase elongation on the template DNA containing the target oligopurine sequence. On a double-stranded target, these complexes only formed at low ionic strength and were slowly dissociated at physiological ionic strength (pH 6.5) with a t1/2 of 6.5-7 h. The salt-dependent instability of preformed complexes on a plasmid target was probably the critical factor responsible for their lack of significant sequence-specific effect on IL-2R alpha promoter activity inside living cells.
Praseuth
D
D
Laboratoire de Biophysique, INSERM U.201-CNRS URA 481, Muséum National d'Histoire Naturelle, Paris, France. praseuth@mnhn.fr
Grigoriev
M
M
Guieysse
A L
AL
Pritchard
L L
LL
Harel-Bellan
A
A
Nielsen
P E
PE
Hélène
C
C
D001665:Binding Sites; D018983:DNA Footprinting; D004278:DNA, Superhelical / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000187:drug effects; D004259:DNA-Directed DNA Polymerase; D004591:Electrophoresis, Polyacrylamide Gel; D017930:Genes, Reporter / Q000235:genetics; D006160:Guanosine Triphosphate / Q000031:analogs & derivatives / Q000378:metabolism; D006860:Hydrogen Bonding; D009690:Nucleic Acid Conformation; D019384:Nucleic Acid Synthesis Inhibitors; D009838:Oligodeoxyribonucleotides / Q000378:metabolism / Q000494:pharmacology*; D020135:Peptide Nucleic Acids; D010455:Peptides / Q000494:pharmacology*; D010957:Plasmids / Q000378:metabolism; D015375:Receptors, Interleukin-2 / Q000378:metabolism*; D012492:Salts / Q000494:pharmacology; D015719:Single-Strand Specific DNA and RNA Endonucleases / Q000378:metabolism; D014157:Transcription Factors / Q000187:drug effects / Q000378:metabolism; D014158:Transcription, Genetic / Q000187:drug effects
D004278:DNA, Superhelical; D004268:DNA-Binding Proteins; D019384:Nucleic Acid Synthesis Inhibitors; D009838:Oligodeoxyribonucleotides; D020135:Peptide Nucleic Acids; D010455:Peptides; D015375:Receptors, Interleukin-2; D012492:Salts; D014157:Transcription Factors; C078138:peptide nucleic acid, T10-lysine; D006160:Guanosine Triphosphate; C034970:bacteriophage T7 induced DNA polymerase; D004259:DNA-Directed DNA Polymerase; D015719:Single-Strand Specific DNA and RNA Endonucleases