Molecular self-assembly is a promising approach to the preparation of nanostructures. DNA, in particular, shows great potential to be a superb molecular system. Synthetic DNA molecules have been programmed to assemble into a wide range of nanostructures. It is generally believed that rigidities of DNA nanomotifs (tiles) are essential for programmable self-assembly of well defined nanostructures. Recently, we have shown that adequate conformational flexibility could be exploited for assembling 3D objects, including tetrahedra, dodecahedra, and buckyballs, out of DNA three-point star motifs. In the current study, we have integrated tensegrity principle into this concept to assemble well defined, complex nanostructures in both 2D and 3D. A symmetric five-pointstar motif (tile) has been designed to assemble into icosahedra or large nanocages depending on the concentration and flexibility of the DNA tiles. In both cases, the DNA tiles exhibit significant flexibilities and undergo substantial conformational changes, either symmetrically bending out of the plane or asymmetrically bending in the plane. In contrast to the complicated natures of the assembled structures, the approach presented here is simple and only requires three different component DNA strands. These results demonstrate that conformational flexibility could be explored to generate complex DNA nanostructures. The basic concept might be further extended to other biomacromolecular systems, such as RNA and proteins.icosahedron ͉ three-dimensional ͉ polyhedron ͉ cryo-EM ͉ molecular cages M olecular self-assembly provides a bottom-up approach to the preparation of nanostructures (1-3). DNA, in particular, shows great potential to be a superb molecular system (4). In the last 20 years, DNA has been explored as building blocks for nanoconstructions, including preparation of periodic and aperiodic 2D nanopatterns (5-8) and 3D polyhedra (9-14). Most of the branched DNA structures are intrinsically flexible and are not suitable building blocks for construction of well defined geometric structures. How to overcome the conformational flexibility of branched DNA structures is a major challenge in structural DNA nanotechnology. In the last decade, a series of rigid structural motifs have been successfully engineered that lead to the rapid evolution of structural DNA nanotechnology (4). However, with more experience and knowledge, it is possible to controllably introduce the conformational flexibility to prepare complex DNA nanostructures (15). In our recent study of 3D self-assembly of DNA three-point-star tiles (16), we found that DNA tetrahedra could be readily assembled, and the tetrahedra are well behaved during sample characterizations. In contrast, DNA dodecahedra and buckyballs have significantly lower assembly yields and are prone to deformation. This phenomenon can be explained by the geometrical differences of these structures. Tetrahedra consist of triangular faces, but others do not. According to tensegrity principle, triangular faces will lead to rigid s...