Abstract:
A research group led by Professor SUZUKI Hiroaki from Faculty of Science and Engineering at Chuo University, graduate students YONEYAMA Ryotaro (at the time), MORIKAWA Naoya, and USHIYAMA Ryota (at the time), Research Fellow TSUGANE Mamiko, Technical Assistant SATO Reiko (at the time), and Special Appointed Assistant Professor MARUYAMA Tomoya from Research Center for Autonomous Systems Materialogy (ASMat), Institute of Integrated Research (IIR), Institute of Science Tokyo, along with Professor TAKINOUE Masahiro from Department of Computer Science, Institute of Science Tokyo, has developed a technology for mass-producing uniform artificial cells (lipid bilayer vesicles) with artificial model nuclei using microfluidic devices with high reproducibility. They also demonstrated that protein synthesis from this model nuclei was possible.
Research on bottom-up construction of the cell model by combining molecules such as lipids, DNA, and proteins is advancing globally, and in recent years, research on reproducing the complex hierarchical structures found in eukaryotic cells has accelerated. Among the various organelles within cells, nucleus is the largest and the most important organelle where genomic DNA is condensed. If this structure could be mimicked within artificial cells, they would become closer to actual living cells, leading to the development of future cell-replacement biotechnology.
This research group has previously developed microfluidic devices as uniform artificial cell production lines*1,2). In this study, they utilized this microfluidic device to encapsulate ingredients necessary to form DNA condensates (DNA nanostars and salt) into cell-sized uniform lipid membrane vesicles (liposomes). Followingly, they concentrated ingredients by inducing volume changes of vesicles using osmotic action to generates uniform model nuclei. Conventionally, thermal annealing was necessary to prepare DNA condensates*3), but it could induce devastating effects on other components comprising artificial cells. The present approach achieves the construction of model nuclei while preserving other enzymes and reaction systems within artificial cells by controlling the concentration changes of materials. As a result, they succeeded in synthesizing green fluorescent protein (GFP) from genes incorporated into this model nuclei.
These artificial cells with model nuclei are expected to provide additional functions such as molecular recognition and environmental response, and we expect this to be utilized for the creation of artificial cells that can replace natural cells in the future.
This research achievement was published in the American Chemical Society's international online academic journal JACS Au on June 16, 2025 (Eastern Time).
【Glossary】
*1) Artificial cell production lines (Assembly line)
Cells of the same type are approximately uniform in size, but conventional liposome preparation methods used in artificial cell research have fundamentally only been able to produce those with varying sizes. Additionally, the ability to encapsulate biomacromolecules at high concentrations is important for constructing artificial cells. Production methods using microfluidic channels can satisfy both of these requirements, but versatile methods have not yet become widespread. Since we belong to the Department of Precision Mechanics, which specializes in the design and manufacturing of precision machinery, we aim to develop technology that can efficiently produce homogeneous artificial cells, which could be called an assembly line for artificial cell production.
*2) Microfluidic Device for Artificial Cell Production (Previously Published)
R. Ushiyama, K. Koiwai, H. Suzuki, "Plug-and-play microfluidic production of monodisperse giant unilamellar vesicles using droplet transfer across water-oil interface", Sensors and Actuators B: Chemical, 355, 131281, 2021.
https://doi.org/10.1016/j.snb.2021.131281
R. Ushiyama, S. Nanjo, M. Tsugane, R. Sato, T. Matsuura, H. Suzuki, "Identifying condition for protein synthesis inside giant vesicles using microfluidics toward standardized artificial cell production," ACS Synthetic Biology, 13(1), 68-76, 2024.
https://doi.org/10.1021/acssynbio.3c00629
*3) DNA Condensates
Chromosomes within cells are highly condensed DNA, and DNA condensates are also being researched and applied in the field of DNA nanotechnology. Of particular interest are condensates of DNA nanostructures called DNA nanostars, consisted of several short single-stranded DNA molecules to form structures with multiple arms such as Y-shaped or X-shaped configurations. By providing sticky ends of 4-10 bases at the tips of the arms, the binding between nanostars can be precisely controlled. This allows for the control of condensation and dissociation while incorporating functions such as molecular recognition.
