Liquid–liquid phase separation, Nucleic-acid condensate, Molecular computing, RNA nanotechnology
See details in Udono, et. al. (2025) ACS Nano.
Hirotake Udono, Minzhi Fan, Yoko Saito, Hirohisa Ohno, Shin-Ichiro M Nomura, Yoshihiro Shimizu, Hirohide Saito, Masahiro Takinoue (2024) Programmable Computational RNA Droplets Assembled via Kissing-Loop Interaction., ACS nano
Published in 2024 Jun 3 (Electronic publication in June 3, 2024, midnight )
(Abstract) DNA droplets, artificial liquid-like condensates of well-engineered DNA sequences, allow the critical aspects of phase-separated biological condensates to be harnessed programmably, such as molecular sensing and phase-state regulation. In contrast, their RNA-based counterparts remain less explored despite more diverse molecular structures and functions ranging from DNA-like to protein-like features. Here, we design and demonstrate computational RNA droplets capable of two-input AND logic operations. We use a multibranched RNA nanostructure as a building block comprising multiple single-stranded RNAs. Its branches engaged in RNA-specific kissing-loop (KL) interaction enables the self-assembly into a network-like microstructure. Upon two inputs of target miRNAs, the nanostructure is programmed to break up into lower-valency structures that are interconnected in a chain-like manner. We optimize KL sequences adapted from viral sequences by numerically and experimentally studying the base-wise adjustability of the interaction strength. Only upon receiving cognate microRNAs, RNA droplets selectively show a drastic phase-state change from liquid to dispersed states due to dismantling of the network-like microstructure. This demonstration strongly suggests that the multistranded motif design offers a flexible means to bottom-up programming of condensate phase behavior. Unlike submicroscopic RNA-based logic operators, the macroscopic phase change provides a naked-eye-distinguishable readout of molecular sensing. Our computational RNA droplets can be applied to in situ programmable assembly of computational biomolecular devices and artificial cells from transcriptionally derived RNA within biological/artificial cells.