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New Method Controls Synthetic DNA Droplet Division Timing
Context:
Researchers at the Tokyo Institute of Technology have developed a method to precisely control when synthetic DNA droplets divide.
More on News:
- These droplets mimic biological liquid-liquid phase separation (LLPS), opening new possibilities for studying cellular processes.
- They achieved this by creating a time-delay circuit that uses a combination of inhibitor RNAs and the enzyme Ribonuclease H (RNase H) to regulate the droplet division process.
Liquid-Liquid Phase Separation (LLPS) Overview:
- LLPS droplets are non-membrane-bound structures formed by soft biological materials within living cells.
- These droplets play crucial roles in various cellular processes by providing a flexible environment.
- LLPS droplets can rapidly adapt to the cell’s needs, allowing them to move, divide, and alter their contents.
- Transcription of ribosomal RNA (rRNA): Particularly in the nucleolus.
- Sol-Gel Transitions: Enabling materials to shift between fluid-like and gel-like states.
- Chemical Reactions: Facilitating control over biochemical processes within the cell.
- The ability of LLPS droplets to regulate cellular functions underscores their significance in maintaining cellular homeostasis and responding to environmental changes.
Methodology
- DNA Structure: The DNA droplets are formed using Y-shaped DNA nanostructures connected by six-branched DNA linkers. These linkers can be cleaved by specific DNA sequences that act as division triggers.
- Division Trigger Mechanism: Initially, division triggers are bound to single-stranded RNA (ssRNA) inhibitors. The enzyme RNase H is introduced to degrade these inhibitors, freeing the triggers to cleave the DNA linkers and initiate droplet division.
- Timing Control: This mechanism introduces a time delay in the cleavage of the DNA linker, allowing precise control over when division occurs.
Key Findings:
- The researchers achieved pathway-controlled division in a ternary-mixed C·A·B-droplet system, which consists of three Y-shaped DNA nanostructures.
- They established two distinct division pathways:
- Pathway 1: C·A·B droplets divide into C droplets, followed by A·B droplets.
- Pathway 2: C·A·B droplets divide into B droplets, followed by C·A droplets.
Applications:
- The pathway control was applied to a molecular computing element known as a comparator, which compares concentrations of microRNA (miRNA) used as RNA inhibitors.
- This allows for quantitative comparisons of RNA levels, opening potential applications in diagnostics.
Challenges and Future Directions:
- Although the study demonstrated promising chemical reactions, these reactions did not create a stable non-equilibrium state akin to natural cellular systems.
- To achieve sustainable non-equilibrium systems, the researchers stress the importance of chemical reactions that maintain a continuous energy supply.