Molecular behavior of DNA in a cell-sized compartment coated by lipids

The behavior of long DNA molecules in a cell-sized confined space was investigated. We prepared water-in-oil droplets covered by phospholipids, which mimic the inner space of a cell, following the encapsulation of DNA molecules with unfolded coil and folded globule conformations. Microscopic observation revealed that the adsorption of coiled DNA onto the membrane surface depended on the size of the vesicular space. Globular DNA showed a cell-size-dependent unfolding transition after adsorption on the membrane. Furthermore, when DNA interacted with a two-phase membrane surface, DNA selectively adsorbed on the membrane phase, such as an ordered or disordered phase, depending on its conformation. We discuss the mechanism of these trends by considering the free energy of DNA together with a polyamine in the solution. The free energy of our model was consistent with the present experimental data. The cooperative interaction of DNA and polyamines with a membrane surface leads to the size-dependent behavior of molecular systems in a small space. These findings may contribute to a better understanding of the physical mechanism of molecular events and reactions inside a cell.

Figure 1

(a) Schematic illustrations of a cell-sized droplet that encapsulates DNAs. (b) Long-axis length ($L$) of unfolded coiled T4 DNA together with a typical microscopic image. The solution was 10 mM Tris-HCl. $N=50$. (c, d) Behavior of coiled DNA with SPD at $\le 0.5$ mM in droplets with DOPC membrane. DNA diffused in an aqueous phase in a small droplet (c), whereas DNA adsorbed on the surface in a large droplet (d). SPD concentrations in (c) and (d) were the same (0.2 mM).

Figure 2

(a) Long-axis length ($L$) of folded globular T4 DNA together with a typical microscopic image. The solutions were 10 mM Tris-HCl with 2.5 mM SPD. $N=50$. (b–d) Behavior of globular DNA with SPD at $\ge 1.0$ mM in droplets with a DOPC membrane. Typical images of DNA before (b) and after (c) incubation for 2 h, using a laser scanning microscope. (d) An aggregation structure of DNA on the membrane surface after 2 h of incubation, which was observed with a fluorescent microscope. SPD concentrations in (c) and (d) were the same (2.5 mM).

Figure 3

Phase diagram of DNA behavior in droplets as a function of droplet radius $R$ and SPD concentration $C_{\text{SPD}}$. Crosses and circles indicate coil and globular conformations of DNA, respectively. Gray region corresponds to the adsorption of DNA on the membrane surface.

Figure 4

(a) Schematic illustration of the adsorption of SPD on the membrane. (b) Number of absorbed SPD molecules at thermal equilibrium per lattice cell, ${\varphi }_{\text{s,equ}}$, as a function of $R$ for different adsorption energies of SPD, ${\epsilon }_{\text{ads}}^{\text{SPD}}$, for 5 $k_{\text{B}}T$, 10 $k_{\text{B}}T$ and SPD concentrations of 0.2 mM, 0.5 mM, 1.0 mM, and 2.5 mM [see Eq. (3)]. (c) Behavior of coiled DNA with SPD of $\le 0.5$ mM in droplets. Open circles and solid lines represent the experimental data and the theoretical fitting, respectively. (d) Behavior of globular DNA with SPD of $\ge 1.0\mathrm{mM}$ in droplets. Open circles and solid lines represent the experimental data and the theoretical fitting, respectively [19].

Figure 5

Selective localization of DNA on phase-separated membrane interfaces, where the membranes were stained with rho-PE, which partitions into the disordered phase. (a) Coiled DNA with 0.2 mM SPD was localized in the ordered phase of the membrane. (b) Globular DNA with 1.0 mM SPD was distributed in the disordered phase, and exhibited Brownian motion. The membranes were composed of DPPC and cholesterol (DPPC/cholesterol = 1:1) [35].