High-affinity triplex-forming oligonucleotide target sequences in mammalian genomes.
Site-specific recognition of duplex DNA by triplex-forming oligonucleotides (TFOs) provides a promising approach to manipulate mammalian genomes. A prerequisite for successful gene targeting using this approach is that the targeted gene must contain specific, high-affinity TFO target sequences (TTS). To date, TTS have been identified and characterized in only approximately 37 human or rodent genes, limiting the application of triplex-directed gene targeting. We searched the complete human and mouse genomes using an algorithm designed to identify high-affinity TTS. The resulting data set contains 1.9 million potential TTS for each species. We found that 97.8% of known human and 95.2% of known mouse genes have at least one potential high-affinity TTS in the promoter and/or transcribed gene regions. Importantly, 86.5% of known human and 83% of the known mouse genes have at least one TTS that is unique to that gene. Thus, it is possible to target the majority of human and mouse genes with specific TFOs. We found substantially more potential TTS in the promoter sequences than in the transcribed gene sequences or intergenic sequences in both genomes. We selected 12 mouse genes and 2 human genes critical for cell signaling, proliferation, and/or carcinogenesis, identified potential TTS in each, and determined TFO binding affinities to these sites in vitro. We identified at least one high-affinity, specific TFO binding site within each of these genes. Using this information, many genes involved in mammalian cell proliferation and carcinogenesis can now be targeted.
Wu
Qi
Q
Department of Carcinogenesis, Science Park-Research Division, The University of Texas M.D. Anderson Cancer Center, Smithville, Texas, USA.
Gaddis
Sara S
SS
MacLeod
Michael C
MC
Walborg
Earl F
EF
Thames
Howard D
HD
DiGiovanni
John
J
Vasquez
Karen M
KM
D000818:Animals; D004247:DNA / Q000737:chemistry*; D024202:Electrophoretic Mobility Shift Assay; D018390:Gene Targeting; D015894:Genome, Human; D006801:Humans; D008322:Mammals / Q000235:genetics*; D051379:Mice; D009841:Oligonucleotides / Q000737:chemistry*; D011401:Promoter Regions, Genetic; D017422:Sequence Analysis, DNA / Q000379:methods*
D009841:Oligonucleotides; C051802:triplex DNA; D004247:DNA