Switchable interfaces, also known as smart interfaces, can alter their macroscopic properties in response to external stimuli. Compared to an artificial switchable interface, DNA-based switchable biointerfaces have high diversity, uniformity, reproducibility, and functionality and are easily designed and developed with atomic precision because the sequence of the DNA strand strictly governs the structural and active properties of its assembly. Moreover, various structures such as double strands based on the Watson-Crick base-pairing rule, G-quadruplexes, i-Motifs, triplexes, and parallel-stranded duplexes exist between or among DNA strands to enrich the structures of DNA biointerfaces. In this article, the design, stimulus responses, and applications of switchable DNA biointerfaces were discussed in terms of single-switch, dual-response, and sequential operation. The applications related to sensing, imaging, delivery, logic gates, and nanomechines were introduced in terms of the design and construction of DNA biointerfaces. Future directions and challenges were also outlined for this rapidly emerging field.