(395e) Using Light Illumination to Control DNA Condensation

Lee, Jr., C. T. - Presenter, University of Southern California
Le Ny, A. - Presenter, University of Southern California

A means to control DNA compaction with light illumination has been developed using the interaction of DNA with a photoresponsive cationic surfactant. The surfactant undergoes a reversible photoisomerization upon exposure to visible (trans) or UV (cis) light. As a result, surfactant binding to DNA and the resulting DNA condensation can be tuned with light. Dynamic light scattering (DLS) measurements are used to follow lambda-DNA compaction from the elongated-coil (end-to-end distance of 1.27 mm) to the compact globular (hydrodynamic radius of 120 nm) form as a function of surfactant addition and light illumination. Moreover, the light-scattering results demonstrate that the compaction process is completely photoreversible. Fluorescence microscopy with T4-DNA is used to further confirm the light-scattering results, allowing single-molecule detection of the light-controlled coil-to-globule transition. These structural studies were combined with absorbance and fluorescence spectroscopy of crystal violet in order to elucidate the binding mechanism of the photosurfactant to DNA. The results indicate that both electrostatic and hydrophobic forces are important in the compaction process. The combined results clearly show the ability to control the interaction between DNA and the complexing agent and, therefore, DNA condensation with light.

DNA condensation is relevant from the point of view of gene therapy, where it has been shown that complexation and the resulting compaction are essential to protect DNA from nuclease and to allow entry of DNA into cells, mainly by the endocytic pathway. Indeed, this method is preferred over viral delivery vectors, due to the potential for immunological responses associated with the use of viruses. However, while complexing agents can increase cellular uptake as a result of DNA neutralization and compaction, the tight binding of these same agents to DNA may also generally preclude or greatly reduce nuclear uptake, resulting in lower transfection efficiencies compared to viral vectors. This occurs because in the condensed state, the interaction of DNA with intracellular molecules such as importin or transportin, which transport DNA into the nucleus, are largely prevented. Thus, the development of these photoreversible DNA complexes could greatly increase transfection efficiency.