A CRISPR-Cas12a-Responsive DNA Tetrahedron

Abstract Functionalizing DNA nanostructures for stimuli-responsive reconfiguration has been a key area of development in DNA nanotechnology. Here, we demonstrate programmable control over the structural integrity of a tetrahedral DNA nanostructure using an external DNA stimulus that is modulated through an intermediate CRISPR-Cas12a reaction. We designed toehold-containing DNA tetrahedra that can be disassembled by a DNA or RNA invading strand. Activation of the Cas12a enzyme by double-stranded DNA provides precise control over the disassembly of the DNA tetrahedra by digesting the DNA invading strand but not the RNA, enabling highly responsive and programmable DNA architectures. Further, targeted activation of Cas12a by conserved genomic regions of three bacterial pathogens, namely Listeria monocytogenes, Campylobacter jejuni, and Shiga toxin-producing Escherichia coli, provides biologically relevant responsiveness and demonstrates the platform’s potential for future applications. Our results show that the DNA tetrahedra are observed in the presence of the genomic target but not observed in its absence due to disassembly, demonstrating a highly programmable and responsive molecular level architecture.