Sensing of transcription factor binding via cyanine dye pair fluorescence lifetime changes.
Summary of "Sensing of transcription factor binding via cyanine dye pair fluorescence lifetime changes."
We designed and synthesized sensors for imaging transcription factor-DNA interactions using a complementary pair of 21-base pair long oligonucleotides (ODNs) carrying two internucleoside phosphate-linked cyanine fluorophores that can either engage in Förster's resonance energy transfer (FRET) with fluorescence emission or assemble into a ground state quenched dimer with short fluorescence lifetimes (FL). Cyanine fluorophores were linked to ODNs within the NF-κB binding site. These sensors were tested in the presence of recombinant p50 and p65 NF-κB proteins or constitutively NF-κB activating HeLa cell lysates. By using a coherent light excitation source we followed changes in fluorescence lifetime of the donor (Cy5.5) at the donor's excitation and emission light wavelengths, as well as the acceptor (800CW or Cy7 cyanine fluorophores) in FRET mode. We observed increases in the donor lifetime in both emitting (0.08-0.15 ns) and non-emitting quenched (0.21 ns) sensors in response to protein binding. The measurements of lifetimes in FRET mode in quenched pair-carrying ODN duplex sensors showed significant differences in FL of the acceptor cyanine fluorophore between NF-κB-containing and NF-κB-free samples but not in control sensors with ODN sequences that have decreased binding affinity to NF-κB. We anticipate that the observed effects will be instrumental for developing sensors enabling non-invasive imaging in cells that undergo activation of NF-κB.
The Laboratory of Molecular Imaging Probes, S6-434, Department of Radiology, University of Massachusetts Medical School, 55 Lake Ave North, Worcester, MA 01655, USA. Alexei.Bogdanov@umassmed.edu.
This article was published in the following journal.
Name: Molecular bioSystems
Medical and Biotech [MESH] Definitions
Transcription Factor Dp1
A transcription factor that possesses DNA-binding and E2F-binding domains but lacks a transcriptional activation domain. It is a binding partner for E2F TRANSCRIPTION FACTORS and enhances the DNA binding and transactivation function of the DP-E2F complex.
Core Binding Factor Beta Subunit
A non-DNA binding transcription factor that is a subunit of core binding factor. It forms heterodimeric complexes with CORE BINDING FACTOR ALPHA SUBUNITS, and regulates GENETIC TRANSCRIPTION of a variety of GENES involved primarily in CELL DIFFERENTIATION and CELL CYCLE progression.
Transcription Factor Tfiia
An RNA POLYMERASE II specific transcription factor. It may play a role in transcriptional activation of gene expression by interacting with the TATA-BOX BINDING PROTEIN component of TRANSCRIPTION FACTOR TFIID.
Core Binding Factors
Heterodimeric transcription factors containing a DNA-binding alpha subunits, (CORE BINDING FACTOR ALPHA SUBUNITS), along with a non-DNA-binding beta subunits, CORE BINDING FACTOR BETA SUBUNIT. Core Binding Factor regulates GENETIC TRANSCRIPTION of a variety of GENES involved primarily in CELL DIFFERENTIATION and CELL CYCLE progression.
Core Binding Factor Alpha 2 Subunit
A transcription factor that dimerizes with the cofactor CORE BINDING FACTOR BETA SUBUNIT to form core binding factor. It contains a highly conserved DNA-binding domain known as the runt domain. Runx1 is frequently mutated in human LEUKEMIAS.
In this report we propose a lifetime-based sensing (LBS) for the detection of hyaluronidase (HA-ase). First, we heavily label hyaluronan macromolecules (HAs) with fluorescein amine. The fluorescein la...
The potential use of CdTe quantum dots as luminescence nano-probes for lifetime fluorescence nano-thermometry is demonstrated. The maximum thermal sensitivity achievable is strongly dependent on the q...
Two-photon (2P) optical properties of cyanine dyes were evaluated using a 2P fluorescence spectrophotometer with 1.55 μm excitation. We report the 2P characteristics of common NIR polymethine dyes, i...
A novel long lifetime fluorescence reporter based on 9-aminoacridine was designed, the lifetime of which can be modulated in a defined manner when in proximity to a tryptophan residue enabling fluores...
Quorum-sensing bacteria communicate via small molecules called autoinducers to coordinate collective behaviors. Because quorum sensing controls virulence factor expression in many clinically relevant...
The purpose of this study is to evaluate the genotype of CTCF, a proven transcription factor, in patients with infantile hemangiomas and to monitor tumor growth. The investigators aim to...
Endolymphatic Sac Tumors in a Population of Patients With Von Hippel-Lindau Disease:The Natural History and Pathobiology, and a Prospective Non-Randomized Clinical Trial of Hearing Preservation Surgery in Patients With Early Stage Endolymphatic Sac Tumors
The von Hippel Lindau (VHL) gene has recently been identified as the genetic defect resulting in a syndrome of multiple neoplasias. Patients with VHL disease develop retinal angiomata, re...
OBJECTIVES: I. Determine the efficacy and toxic effects of recombinant human insulin-like growth factor I (rhIGF-I) on carbohydrate tolerance, insulin action, insulin secretion, hyperandro...
The aim of the study is the comparison of two different leads in their capabilities to detect episodes and duration of paroxysmal atrial fibrillation (AF) and atrial tachyarrhythmia (AT),...
The aim of this study is to investigate the safety and feasibility of daily subcutaneous injections of recombinant IGF1 complexed with IGF binding protein 3 (SomatoKine-INSMED) as a treatm...