Using AFM to probe the complexation of DNA with anionic lipids mediated by Ca2+: the role of surface pressure

Abstract

Complexation of DNA with lipids is currently being developed as an alternative to classical vectors based on viruses. Most of the research to date focuses on cationic lipids owing to their spontaneous complexation with DNA. Nonetheless, recent investigations have revealed that cationic lipids induce a large number of adverse effects on DNA delivery. Precisely, the lower cytotoxicity of anionic lipids accounts for their use as a promising alternative. However, the complexation of DNA with anionic lipids (mediated by cations) is still in early stages and is not yet well understood. In order to explore the molecular mechanisms underlying the complexation of anionic lipids and DNA we proposed a combined methodology based on the surface pressure-area isotherms, Gibbs elasticity and Atomic Force Microscopy (AFM). These techniques allow elucidation of the role of the surface pressure in the complexation and visualization of the interfacial aggregates for the first time. We demonstrate that the DNA complexes with negatively charged model monolayers (DPPC/DPPS 4 : 1) only in the presence of Ca(2+), but is expelled at very high surface pressures. Also, according to the Gibbs elasticity plot, the complexation of lipids and DNA implies a whole fluidisation of the monolayer and a completely different phase transition map in the presence of DNA and Ca(2+). AFM imaging allows identification for the first time of specific morphologies associated with different packing densities. At low surface coverage, a branched net like structure is observed whereas at high surface pressure fibers formed of interfacial aggregates appear. In summary, Ca(2+) mediates the interaction between DNA and negatively charged lipids and also the conformation of the ternary system depends on the surface pressure. Such observations are important new generic features of the interaction between DNA and anionic lipids.

Like-charge attraction and opposite-charge decomplexation between polymers and DNA molecules

“…[12]. Therein, the approach was applied to the case of a polymer interacting with a like-charged plane wall in order to explain the adsorption of DNA molecules onto negatively charged lipid membranes [10]. In the present problem, the test-charge approach allows us to recover the cylindrical symmetry dictated by the geometry of the DNA molecule.…”

“…The biological functions of this anionic molecule are intimately related to its electrostatic coupling with the surrounding macromolecular charges. In this context, the like-charge attraction phenomenon plays a capital role in DNA packing around histones [1], the condensation of polymer solutions [2][3][4][5][6][7][8][9], DNA adsorption on anionic substrates [10][11][12][13], and gene delivery techniques [14]. The characterization of like-charge attraction is thus of paramount importance for understanding and controlling in vivo and in vitro biological processes involving DNA molecules.…”

Improved DNA condensation, stability, and transfection with alkyl sulfonyl-functionalized PAMAM G2

“…Penetration of the dendriplexes into the lipid monolayer could result in alterations of the lateral packing in the monolayer (Luque-Caballero et al 2014). In order to investigate this in more detail, we measured the surface dilatational moduli of the adsorbed dendriplexes onto the DPPC monolayer.…”

“…By deposition with a microsyringe, a fixed amount of DPPC is spread onto the drop surface to provide a surface pressure of 20 mN/m. After evaporation of the solvent and equilibration of the surface pressure, the buffer subphase is exchanged by simultaneously injecting the new solution and extracting the bulk subphase (Maldonado-Valderrama et al 2014;Wege et al 2002). After complete exchange of the subphase, we monitor the surface pressure at constant surface area for 20 min.…”

Cytotoxicity and behavior of polystyrene latex nanoparticles to budding yeast

“…In the presence of Mg 2+ and Ca 2+ , extremely strong interactions are often observed in biological membranes [25]. To investigate the effect of cationic species in an electrolyte dispersion medium on the cell viability and NP location, the cytotoxicity tests and CLSM measurements were carried out in different ionic strength dispersion media.…”