Using Well-Defined DNA Nanostructures To Study the Influence of DNA Clustering and Presentation on SNA Cellular Uptake

Spherical nucleic acids (SNAs) are examples of how nucleic acid structures can impact important biological functions. Herein, we explore how well-defined DNA nanostructures assembled on the surface of preformed SNAs can influence important processes like cellular uptake. Three different DNA nanostructures, which vary in clustering and/or topology, were studied with three different cell lines (NIH-3T3, HaCaT, RAW 264.7). All three structures exhibited higher cellular uptake than conventional SNAs, with one structure (TX motif SNA) exhibiting a 5-fold increase after 4 h of incubation. Increased DNA clustering and DNA crossover numbers correlate with enhanced Ca 2+ binding and, ultimately, higher uptake primarily through clathrin- and macropinocytosis-mediated pathways (caveolae-mediated pathways have been observed with traditional ssSNAs). Ca 2+ content within SNA structures facilitates uptake by making the structure less negatively charged and increasing interactions with Ca 2+ -binding proteins. This work shows how structural manipulation of the SNA shell can control and optimize its biological function.