Delivery and imaging of functionalized DNA origami in cells
Résumé
Human cells package over two meters of genetic information into the microscopic space of the nucleus in the form of chromatin (a complex between DNA and histone proteins). This packaging or “folding” of the genome is critical for the regulation of gene expression, maintenance of genomic stability, and defining cell fate during development. Change to the spatial organization of chromatin is implicated in a host of neurological disorders as well as in aging and cancer. Remodeling of chromatin via post-translational modifications allows for epigenetic regulation of transcription in healthy and diseased cells. Thus, tools to engineer chromatin epigenetics in a controlled manner are crucial to study the cause-consequence relationship between epigenetics, chromatin organization and gene expression. Previous approaches to chromatin engineering have significant limitations, namely the inability to perform multiple functions (visualize, modify, or detect downstream products) simultaneously. Our work seeks to develop and apply DNA origami nanodevices (DOs) as multi-functional platforms to probe and engineer chromatin epigenetics. We propose to leverage the versatility of DO and its potential as a platform for multiple functional elements to visualize, modify, and sense the transcriptional output of genomic regions in mammalian cell nuclei. We have taken an interdisciplinary approach to develop and test four different DNA origami structures (26-helix bundle, 14-helix bundle, 13-helix bundle, and 8-helix bundle) functionalized with fluorophores as well as an anti-RNA Polymerase II (Pol2) antibody. We show that electroporation enables delivery of these structures into cells and further, 8-helix bundle structures functionalized with RNA Pol2 antibodies are successfully piggybacked into the nucleus. Super-resolution imaging further demonstrates the delivery of single DNA origami structures into the cell nucleus. These results open the door for DNA origami as a multifunctional platform for targeting, modifying, and visualizing the genome.