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Funding Information: The research of A.E., A.M., and P.O. was supported by Academy of Finland grant 311639. A.K. and A.K.N. have been supported by Academy of Finland grant 308992. The research of I.K. was supported by Japan Society for the Promotion of Science (JSPS) Early-Career Scientists 18K18144, Fund for the Promotion of Joint International Research (B) 19KK0261, and Young Researcher Dispatch Program (School of Engineering, Tohoku University). The research of A.M. was additionally supported by NSF-DMS (grant numbers 1800443/1764366) and Nokia Foundation (2017). The research of Y.S. has been supported by JSPS Grant-in-Aid for Scientific Research (KAKENHI; grant numbers 18K19831 and 19H04201). The research of L.O. and F.C.S. was supported by European Research Council grant agreement no. 694410, project AEDNA. Publisher Copyright: © 2022 The Authors. Published by American Chemical Society. We address the problem of de novo design and synthesis of nucleic acid nanostructures, a challenge that has been considered in the area of DNA nanotechnology since the 1980s and more recently in the area of RNA nanotechnology. Toward this goal, we introduce a general algorithmic design process and software pipeline for rendering 3D wireframe polyhedral nanostructures in single-stranded RNA. To initiate the pipeline, the user creates a model of the desired polyhedron using standard 3D graphic design software. As its output, the pipeline produces an RNA nucleotide sequence whose corresponding RNA primary structure can be transcribed from a DNA template and folded in the laboratory. As case examples, we design and characterize experimentally three 3D RNA nanostructures: a tetrahedron, a triangular bipyramid, and a triangular prism. The design software is openly available and also provides an export of the targeted 3D structure into the oxDNA molecular dynamics simulator for easy simulation and visualization.

Intaking molecular information from the external environment is essential for the normal functioning of artificial cells/molecular robots. Herein, we report the design and function of a membrane nanopore using a DNA origami square tube with a cross-section of 100 nm2. When the nanopore is added to a giant vesicle that mimics a cell membrane, the permeation of large external hydrophilic fluorescent molecules is observed. Furthermore, the addition of up to four ssDNA strands enables size-based selective transport of molecules. A controllable artificial nanopore should facilitate the communication between the vesicle components and their environment.

Making use of the programmability and structural flexibility of the DNA molecule, a DNA-origami nanoarm capable of undergoing large deformation is constructed. This DNA-origami nanoarm comprised serially repeated tension-adjustable modules, the cumulative actuation of which resulted in a large deformation of the arm structure, which transformed from a linear shape into an arched shape. Combining atomic force microscopy and theoretical analyses based on the mechanics of materials, we demonstrate that the degree of deformation can be systematically controlled by merely replacing a set of strands that is required for the actuation of the module. Moreover, by employing a G-quadruplex-forming sequence for the actuation, we could achieve reversible ion-induced contraction and relaxation of the nanoarm. The adjustability and scalability of this design could enable the production of DNA nanodevices that exhibit large deformation in response to external stimuli.

Reaction-diffusion systems are one of the models of the formation process with various patterns found in nature. Inspired by natural pattern formation, several methods for designing artificial chemical reaction-diffusion systems have been proposed. DNA is a suitable building block to build such artificial systems owing to its programmability. Previously, we reported a line pattern formed due to the reaction and diffusion of synthetic DNA; however, the width of the line was too wide to be used for further applications such as parallel and multi-stage pattern formations. Here, we propose a novel method to programme a reaction-diffusion system in a hydrogel medium to realise a sharp line capable of forming superimposed and cascaded patterns. The mechanism of this system utilises a two-segment polymerisation of DNA caused by hybridisation. To superimpose the system, we designed orthogonal DNA sequences that formed two lines in different locations on the hydrogel. Additionally, we designed a reaction to release DNA and form a cascade pattern, in which the third line appears between the two lines. To explain the mechanism of our system, we modelled the system as partial differential equations, whose simulation results agreed well with the experimental data. Our method to fabricate cascaded patterns may inspire combinations of DNA-based technologies and expand the applications of artificial reaction-diffusion systems.

We demonstrate a method of pattern formation based on an artificial reaction diffusion system in hydrogel medium. By designing both the reaction term and the diffusion term of the system, we have succeeded in generating a sustainable DNA pattern in the gel. A DNA logic gate anchored in the gel detected the diffused molecules from distant source points to mark the equidistant region from the source points, which produced a Voronoi pattern in the gel. Combined with a diffusion modulation method for DNA molecules, the Voronoi pattern was modified as a weighted Voronoi pattern with different morphologies. The proposed framework will be useful in designing a structured gel system responsive to molecular signals.

An isothermal amplification circuit for specific DNA molecules was implemented in giant unilamellar vesicles. Using this circuit, over 5000-fold amplification of output DNAs was achieved, and the amplification behaviour depended on the concentration of input signal DNAs in a cell-sized compartment. Moreover, initiation of the amplification by photo-stimulation was demonstrated.

Cell motions such as migration and change in cellular morphology are essential activities for multicellular organism in response to environmental stimuli. These activities are a result of coordinated clustering/declustering of integrin molecules at the cell membrane. Here, we prepared DNA origami nanosprings to modulate cell motions by targeting the clustering of integrin molecules. Each nanospring was modified with arginyl-glycyl-aspartic acid (RGD) domains with a spacing such that when the nanospring is coiled, the RGD ligands trigger the clustering of integrin molecules, which changes cell motions. The coiling or uncoiling of the nanospring is controlled, respectively, by the formation or dissolution of an i-motif structure between neighboring piers in the DNA origami nanodevice. At slightly acidic pH (

The invention of DNA origami has expanded the geometric complexity and functionality of DNA nanostructures. Using DNA origami technology, we develop a flexible multi-joint ring motif as a novel self-assembling module. The motif can connect with each other through self-complementary sequences on its segments. The flexible joints can be fixed in a straightened position as desired, thereby allowing the motif to take various shapes. We can adjust the number of flexible joints and the number of connectable segments, thereby enabling programmable self-assembly of the motif. We successfully produced the motif and evaluated several self-assembly patterns. The proposed multi-joint ring motif can provide a novel method for creating functional molecular devices.

Here, we report on the design and function of a membrane nanopore through a DNA origami square tube with a cross-section of 100 nm2 . When the nanopore is added onto the giant vesicle membrane, the permeation of hydrophilic fluorescent molecules was observed. It can be sealed by the existence of the four specific single strand DNAs. A controllable artificial nanopore should help to communicate the vesicle components with their environment

In this paper, we consider the implementation of a cellular automaton by DNA computing. The proposed system is a reaction–diffusion system built on a structured hydrogel matrix that mimics cellular compartments of biological tissues. Since the cellular automaton is materialized by a hydrogel matrix, the system is called gellular automaton which is theoretically capable of pattern formation and computation by chemical reactions. We focus on technical aspects of the implementation of the gellular automata, such as fabrication of the array of cells, the realization of inter-cellular molecular communication, and how to realize state transitions of cells. Along with the evaluation of each technical element, some simple experimental demonstrations of pattern formation are described.

Investigations into the refolding of DNA origami leads to the creation of reconstructable nanostructures and deepens our understanding of the sustainability of life. Here, we report the refolding of the DNA origami structure inside a micron-sized compartment. In our experiments, conventional DNA origami and truss-type DNA origami were annealed and purified to remove the excess staples in a test tube. The DNA origami was then encapsulated inside of a micron-sized compartment of water-in-oil droplets, composed of neutral surfactants. The re-annealing process was then performed to initiate refolding in the compartment. The resulting 100-nm-sized DNA nanostructures were observed using atomic force microscopy (AFM), and the qualities of their structures were evaluated based on their shape. We found that the refolding of the DNA origami structure was favored inside the droplets compared with refolding in bulk solution. The refolded structures were able to fold even under &ldquo, quick&rdquo, one-minute annealing conditions. In addition, the smaller droplets (average diameter: 1.2 &micro, m) appeared to be more advantageous for the refolding of the origamis than larger droplets. These results are expected to contribute to understanding the principles of life phenomena based on multimolecular polymer self-assembly in a micron-sized compartment, and for the production and maintenance of artificially designed molecules.

The most common way to fabricate DNA nanostructures is to mix individually synthesized DNA oligomers in one pot. However, if DNA nanostructures could be produced through enzymatic reactions, they could be applied in various environments, including in vivo. Herein, an enzymatic method developed to construct a DNA nanostructure from a simple motif called a T-motif is reported. A long, repeated structure was replicated from a circular template by rolling circle amplification and then cleaved into T-motif segments by restriction enzymes. These motifs have been successfully assembled into a ladder-like nanostructure without purification or controlled annealing. This approach is widely applicable to constructing a variety of DNA nanostructures through enzymatic reactions.

How to construct a reliable deoxyribonucleic acid (DNA) circuit is one of the most important issues in the field of molecular programming and computing. Such a circuit frequently suffers from various kinds of unintended binding reactions on account of a lower specificity of the molecular interaction emerging from base complementarity between DNA strands, which results in low reliability of the molecular circuit. In this study, we propose a method to quantitatively evaluate the robustness of a DNA circuit created by DNA strand displacement reactions. The highlight of our method is representing the circuit system contaminated by unintended reactions by a standard form of a disturbed nonlinear system and evaluating the robustness with the L2 gain by solving the Hamilton–Jacobi inequality.

Cell-sized liposomes and droplets coated with lipid layers have been used as platforms for understanding live cells, constructing artificial cells, and implementing functional biomedical tools such as biosensing platforms and drug delivery systems. However, these systems are very fragile, which results from the absence of cytoskeletons in these systems. Here, we construct an artificial cytoskeleton using DNA nanostructures. The designed DNA oligomers form a Y-shaped nanostructure and connect to each other with their complementary sticky ends to form networks. To undercoat lipid membranes with this DNA network, we used cationic lipids that attract negatively charged DNA. By encapsulating the DNA into the droplets, we successfully created a DNA shell underneath the membrane. The DNA shells increased interfacial tension, elastic modulus, and shear modulus of the droplet surface, consequently stabilizing the lipid droplets. Such drastic changes in stability were detected only when the DNA shell was in the gel phase. Furthermore, we demonstrate that liposomes with the DNA gel shell are substantially tolerant against outer osmotic shock. These results clearly show the DNA gel shell is a stabilizer of the lipid membrane akin to the cytoskeleton in live cells.

We investigated spatio-temporal pattern formation in a reaction-diffusion system assuming a periodic cellular space. The reaction space is an array of cells, where diffusion is assumed to be fast inside the cells and relatively slow in the walls separating them. The simulation results showed that the spatio-temporal development of the concentration pattern on the array depends on some specific parameters such as the diffusion coefficients in the cell walls and the size of cells. In a certain parameter region, the concentration inside each cell takes either a high or low value, and moreover, these locally discretized cells generate an alternating pattern spreading into the entire space. Whole of this process can be regarded as discretization of state, time, and space of a chemical reaction system which is intrinsically continuous. This kind of discretization mechanism is expected to provide a new way of the implementation of cellular automata based on molecular reactions.

We propose a method to control the diffusion speed of DNA molecules with a target sequence in a polymer solution. The interaction between solute DNA and diffusion-suppressing DNA that has been anchored to a polymer matrix is modulated by the concentration of the third DNA molecule called the competitor by a mechanism called toehold exchange. Experimental results show that the sequence-specific modulation of the diffusion coefficient is successfully achieved. The diffusion coefficient can be modulated up to sixfold by changing the concentration of the competitor. The specificity of the modulation is also verified under the coexistence of a set of DNA with noninteracting base sequences. With this mechanism, we are able to control the diffusion coefficient of individual DNA species by the concentration of another DNA species. This methodology introduces a programmability to a DNA-based reaction-diffusion system.

We constructed a rotary DNA origami device and tested its stepping operation on a mica substrate by sequential strand displacement with four different sets of signal DNA strands. This work paves the way for building a variety of dynamic rotary DNA nanodevices which respond to multiple signals.

We show electric control of unzipping and shearing dehybridization of a DNA duplex anchored to a hydrogel. Tensile force is applied by electrophoresing (25 V cm−1) gold nanoparticles pulling the DNA duplex. The pulled DNA strand is gradually released from the hydrogel. The unzipping release rate is faster than shearing; for example, 3-fold for a 15 base pair duplex, which helps to design electrically driven DNA devices.

Part 2: Regular Papers; International audience; Asynchronous Boolean totalistic cellular automata have recently attracted attention as promising models for the implementation of reaction-diffusion systems. It is unknown, however, to what extent they are able to conduct computation. In this paper, we introduce the so-called non-camouflage property, which means that a cell’s update is insensitive to neighboring states that equal its own state. This property is stronger than the Boolean totalistic property, which signifies the existence of states in a cell’s neighborhood, but is not concerned with how many cells are in those states. We argue that the non-camouflage property is extremely useful for the implementation of reaction-diffusion systems, and we construct an asynchronous cellular automaton with this property that is Turing-complete. This indicates the feasibility of computation by reaction-diffusion systems.

Rapid progress in nanoscale bioengineering has allowed for the design of biomolecular devices that act as sensors, actuators, and even logic circuits. Realization of micrometer-sized robots assembled from these components is one of the ultimate goals of bioinspired robotics. We constructed an amoeba-like molecular robot that can express continuous shape change in response to specific signal molecules. The robot is composed of a body, an actuator, and an actuator-controlling device (clutch). The body is a vesicle made from a lipid bilayer, and the actuator consists of proteins, kinesin, and microtubules. We made the clutch using designed DNA molecules. It transmits the force generated by the motor to the membrane, in response to a signal molecule composed of another sequence-designed DNA with chemical modifications. When the clutch was engaged, the robot exhibited continuous shape change. After the robot was illuminated with light to trigger the release of the signal molecule, the clutch was disengaged, and consequently, the shape-changing behavior was successfully terminated. In addition, the reverse process-that is, initiation of shape change by input of a signal-was also demonstrated. These results show that the components of the robot were consistently integrated into a functional system. We expect that this study can provide a platform to build increasingly complex and functional molecular systems with controllable motility.

We introduce two autonomous chemical reaction-diffusion models that can emulate the behavior of specific cellular automata. One model conducts formation of a 3-color checker-board pattern using an abstract chemical reaction network. The other model is based on a DNA reaction-diffusion system that is capable of emulating a Turing-complete one-dimensional cellular automaton. These frameworks can be used to systematically program spatiotemporal pattern formation, and thus has a potential for an effective macro-scale control of molecular systems.

Stimuli-responsive DNA gels that can undergo a sol-gel transition in response to photo-irradiation provide a way to engineer functional gel material with fully designed DNA base sequences. We propose an X-shaped DNA motif that turns into a gel by hybridization of self-complementary sticky ends. By embedding a photo-crosslinking artificial base in the sticky-end sequence, repetitive gel-sol transitions are achieved through UV irradiation at different wavelengths. The concentration of the DNA motif necessary for gelation is as low as 40 μm after modification of the geometrical properties of the motif. The physical properties, such as swelling degree and diffusion coefficient, were assessed experimentally.

This paper presents a Cellular Automaton CA model designed for possible implementation by the reaction and diffusion of DNA strands. The proposed CA works asynchronously, whereby each cell undergoes its transitions independently from other cells and at random times. The state of a cell changes in a cyclic manner, rather than according to an any-to-any mapping. The transition rules are designed as totalistic, i.e., the next state of a cell is determined only by the number of states in the neighborhood of the cell, not by their relative positions. Universal circuit elements are designed for the CA as well as wires and crossings to connect them, which implies that the CA is Turing-complete.

We introduce a theoretical model of DNA chemical reaction-diffusion network capable of performing a simple cellular automaton. The model is based on well-characterized enzymatic bistable switch that was reported to work in vitro. Our main purpose is to propose an autonomous, feasible, and macro DNA system for experimental implementation. As a demonstration, we choose a maze-solving cellular automaton. The key idea to emulate the automaton by chemical reactions is assuming a space discretized by hydrogel capsules which can be regarded as cells. The capsule is used both to keep the state uniform and control the communication between neighboring capsules. Simulations under continuous and discrete space are successfully performed. The simulation results indicate that our model evolves as expected both in space and time from initial conditions. Further investigation also suggests that the ability of the model can be extended by changing parameters. Possible applications of this research include pattern formation and a simple computation. By overcoming some experimental difficulties, we expect that our framework can be a good candidate to program and implement a spatio-temporal chemical reaction system.