Dyads of G-Quadruplex Ligands Triggering DNA Damage Response and Tumour Cell Growth Inhibition at Subnanomolar Concentration.

Naphthalene diimide (NDI) dyads exhibiting a different substitution pattern and linker length have been synthesised and evaluated as G-quadruplex (G4) ligands, by investigating their cytotoxicity in selected cell lines. The dyads with the long C7 linker exhibit extremely low IC50 values, below 10 nm, on different cancer cell lines. Contrary, the dyads with the shorter C4 linker were much less effective, with IC values increasing up to 1 µm. Among the three dyads with the longest linker, small differences in the IC50 values emerge, suggesting that the linker length plays a more important role than the substitution pattern. We have further shown that the dyads are able to induce cellular DNA damage response, which is not limited to the telomeric regions and is likely the origin of their cytotoxicity. Both absorption titration and dynamic light scattering of the most cytotoxic dyads in the presence of hTel22 highlight their ability to induce effective G4 aggregation, acting as non-covalent cross-linking agents.

DNA aggregation induced by Mg2+ ions under different conditions.

Cations-induced DNA aggregation can modify the local structure of oligonucleotides and has potential applications in medicine and biotechnology. Here, we used atomic force microscopy to investigate λ-DNA aggregation on Mg2+ -treated glass (Mg2+ /glass) and in Mg2+ solution. Atomic force microscopy topography images showed that some DNA fragments were slightly stacked together on 10 mM Mg2+ /glass and stacked stronger on ≥50 mM Mg2+ /glass. They also showed that DNA aggregated stronger in Mg2+ solution than on Mg2+ /glass, ie, DNAs are strongly stacked and twisted at 10 mM Mg2+ , rolled together at 50 mM Mg2+ , and slightly aggregated to form small particles at 100 mM Mg2+ . At a specific condition, ie, heating λ-DNA to 92°C, cooling down to 75°C, adding Mg2+ , and vortexing the resulting solution, DNA strongly aggregated and formed pancake-like shapes at 10 and 50 mM or a large aggregate at 100 mM Mg2+ solutions. Our results may be helpful for medical applications and gene therapy using cation-DNA technology.

Binding between an amine cationic surfactant and DNA and surface properties of the resultant aggregates.

Binding between cetyltrimethylammonium bromide, a cationic surfactant, and a variety of lengths of single stranded DNA was measured using fluorescence polarization and a simple cooperative model was used to obtain dissociation constants on the order of 1 × 10-5 for the aggregates that formed. Aggregation depended on strand length where strands much shorter than 40 nucleotides (for example strands of 24-nucleotides) were too short to form the same size aggregates. Other factors such as salt concentration and temperature also affected aggregate formation: increasing either the salt concentration or performing binding at the highest temperature studied (60 °C) made it more difficult for aggregates to form. Both heating and dilution of aggregates caused the anisotropy signal to decrease, which suggested that the complexes fell apart under these conditions. Force spectroscopy of aggregate surfaces showed that both electrostatic and hydrophobic adhesive forces were present between aggregates and derivatized AFM tips. These findings can be used to better understand the stability of cationic surfactant-DNA aggregates and may provide guidance for lipid nanoparticle design used in vaccine development and therapeutics.

DNA-membrane complex formation during electroporation is DNA size-dependent.

Size of DNA molecules governs their interaction with the cell membrane during electroporation and their subsequent transport inside the cell. In order to investigate the effect of DNA size on DNA-membrane interaction during electroporation, cells are electro-pulsed with DNA molecules; 15 bp, 25 bp, 50 bp, 100 bp and 1000 bp (bp = base pairs). Within the experimental parameter space, DNA-membrane complexes or DNA aggregates are observed at the cell membrane for DNA molecules containing 25 or more base pairs. No aggregates are observed for DNA molecules containing 15 bp. For all DNA sizes, direct access to the cytoplasm is observed, however the amount translocated decays with the size. The observed dependency of DNA aggregate formation on the size of the DNA molecules is consistent with the Onsager's theory of condensation of anisotropic rod-like molecules.

The Staphylococcus aureus Extracellular Adherence Protein Eap Is a DNA Binding Protein Capable of Blocking Neutrophil Extracellular Trap Formation.

The extracellular adherence protein (Eap) of Staphylococcus aureus is a secreted protein known to exert a number of adhesive and immunomodulatory properties. Here we describe the intrinsic DNA binding activity of this multifunctional secretory factor. By using atomic force microscopy, we provide evidence that Eap can bind and aggregate DNA. While the origin of the DNA substrate (e.g., eukaryotic, bacterial, phage, and artificial DNA) seems to not be of major importance, the DNA structure (e.g., linear or circular) plays a critical role with respect to the ability of Eap to bind and condense DNA. Further functional assays corroborated the nature of Eap as a DNA binding protein, since Eap suppressed the formation of "neutrophil extracellular traps" (NETs), composed of DNA-histone scaffolds, which are thought to function as a neutrophil-mediated extracellular trapping mechanism. The DNA binding and aggregation activity of Eap may thereby protect S. aureus against a specific anti-microbial defense reaction from the host.