ABSTRACT
We successfully introduced two-photon-sensitive photolabile groups ([7-(diethylamino)coumarin-4-yl]methyl and p-dialkylaminonitrobiphenyl) into DNA strands and demonstrated their suitability for three-dimensional photorelease. To visualize the uncaging, we used a fluorescence readout based on double-strand displacement in a hydrogel and in neurons. Orthogonal two-photon uncaging of the two cages is possible, thus enabling complex scenarios of three-dimensional control of hybridization with light.
Subject(s)
Color , DNA/chemistry , Nucleic Acid Hybridization , PhotonsABSTRACT
We synthesized a two-photon-sensitive photocleavable linker based on the 7-diethylaminocoumarin structure and introduced it successfully into DNA strands. First, we demonstrated the inducibility of strand scissions upon irradiation at 365 nm. To verify and visualize the two-photon activity, we used a fluorescence assay based on a DNA strand displacement immobilized in a hydrogel. Additionally, we investigated its use in a new class of DNA decoys that are able to catch and release nuclear factor κB (NF-κB) by using light as an external trigger signal. In cell culture we were able to show the regulation of NF-κB-controlled transcription of green fluorescent protein.
Subject(s)
DNA Breaks , DNA/radiation effects , Oligonucleotides/chemistry , Oligonucleotides/radiation effects , Photons , Chromatography, High Pressure Liquid , Coumarins/chemistry , DNA/chemistry , Drug Delivery Systems , Electrophoretic Mobility Shift Assay , Humans , Molecular Structure , Protein Serine-Threonine Kinases/chemistry , NF-kappaB-Inducing KinaseABSTRACT
In this study the new nucleobase-caged nucleotides dT(NpHP) and dT(DEACM) are introduced. Together with two other caging groups (NDBF and NPP) this results in four layers of wavelength-selective uncaging for oligonucleotides, sequentially going from 505 to 440 nm, 365 nm, and finally to 313 nm for the photolysis reaction.