Specific inhibition of c-fos proto-oncogene expression by triple-helix-forming oligonucleotides.
The promoter region of the c-fos oncogene 5' flanking sequence contains enhancer elements crucial for binding nuclear factors that regulate transcription following cell proliferation and differentiation. Single-stranded deoxyoligonucleotides were chosen for modulation of c-fos protooncogene expression because of their high-affinity binding to specific nucleotide sequences. We designed two oligonucleotides that form a triple-helix complex on the retinoblastoma gene product-responsible element of the c-fos oncogene. Modification of the DNA triplex with dimethyl sulfate and affinity cleaving assays demonstrate that the predicted oligonucleotides form a DNA triplex structure with the c-fos promoter in a sequence-specific manner. Tumorigenic and non-tumorigenic fibroblasts were transiently transfected with fos-CAT plasmid modified with alkylating triplex-forming oligonucleotide reagents. A dramatic depression of CAT activity was found when the cross-linked triple helix complex at the retinoblastoma gene product-related site of the c-fos promoter was used. These experiments suggest that transcription of individual genes can be selectively modulated in cell culture by sequence specific triplex formation in regulatory enhancer sequences.
Lavrovsky
Y
Y
Rockefeller University, New York, New York 10021, USA.
Mastyugin
V
V
Stoltz
R A
RA
Abraham
N G
NG
D000477:Alkylating Agents; D002478:Cells, Cultured; D015500:Chloramphenicol O-Acetyltransferase / Q000037:antagonists & inhibitors; D005786:Gene Expression Regulation / Q000502:physiology*; D017930:Genes, Reporter; D016161:Genes, Retinoblastoma; D016762:Genes, fos; D015335:Molecular Probes; D009690:Nucleic Acid Conformation; D009841:Oligonucleotides / Q000737:chemistry*; D011401:Promoter Regions, Genetic; D013463:Sulfuric Acid Esters
D000477:Alkylating Agents; D015335:Molecular Probes; D009841:Oligonucleotides; D013463:Sulfuric Acid Esters; D015500:Chloramphenicol O-Acetyltransferase; C007482:dimethyl sulfate