Spudcell: the first synthetic cell with a complete cell cycle
overview
Professor Kate Adamala and her team at the University of Minnesota have created Spudcell, a cell-like system built entirely from known chemical components that can execute the complete cell cycle.
The system consists of 36 purified enzymes, a 90,000 base pair genome spread across nine different DNA molecules, and a lipid membrane. The spudcell is able to grow, replicate its genome, divide, and undergo selection and competition over many generations.
Unlike earlier work on minimalist cells that modeled living cells, the Spudcell is built from the bottom up from completely individually purified, nonliving components. This is the first time that such a system has demonstrated a complete cell cycle.
What does the spudcell display?
- Genetically controlled food and growth. Spudcells grow by fusion with smaller “feeder liposomes” that provide lipids and nutrients including ribosomes, enzymes, and small molecules for membrane growth. Fusion occurs when a protein that the spudcell makes from its own DNA locks onto the membrane of the feeder, with the cell’s DNA directly controlling whether it can feed, how fast it grows, and how big it becomes. Natural cells make their own nutrients through metabolism, which requires hundreds of genes encoding metabolic enzymes. By feeding externally instead, the spudcell can complete a full cell cycle with a much smaller genome.
- Division without cytoskeleton. Natural cells divide using an internal scaffold called the cytoskeleton. Building a functional cytoskeleton from scratch has been a major hurdle in synthetic cell research because it requires dozens of proteins working in coordination. The spudcell bypasses this completely, causing the proteins to aggregate together on the membrane surface until mechanical stress splits the membrane. Cells that make this surface protein in greater quantities divide more efficiently, directly linking the genome to reproductive success.
- Selection and competition. When researchers introduced a genetic change that increased production of the fusion protein, cells with that change grew faster and produced more offspring. After five generations, the faster-growing version had outperformed the original. Under nutrient deficiency, profits increased. This perfectly reflects the selection and competition going on in synthetic chemical systems.
technical architecture breakdown
Genome Organization: The 90 kbp genome is divided into seven separate DNA plasmids instead of one chromosome. Each plasmid encodes specific functions. This modular architecture allows individual functions to be modified independently. Previous analyzes had estimated that the minimum genome for a living cell could be as small as 113 kbp. The 90 kbp genome of Spudcella is smaller than this theoretical minimum.
Protein Expression System: SpudCell uses the PURE (Protein Synthesis Using Recombinant Elements) system for protein expression. Pure is a defined blend of 36 purified enzymes derived from E. coli bacteria, including the ribosome, which reads DNA and makes proteins. Unlike earlier methods, which used crude bacterial cell extracts, in PURE every ingredient and its concentrations are known, meaning researchers can track exactly what is happening inside the cell.
Chemical Composition: Synthetic cells have a defined chemical composition with known concentrations of all components at the time of manufacture. The cells are liposomes – hollow spheres made of lipid molecules (the same fatty molecules that make up natural cell membranes). Inside each liposome is a DNA genome and a purified protein expression system. All the proteins the spudcell needs to function are made inside the cell from its own genome.
Food Arrangement: The Spudcell uses a protein called α-hemolysin to fuse with the feeder liposome. When the spudcell makes this protein from its DNA, the protein itself inserts into the membrane and spreads out completely. A chemical tag attached to the protein sticks to the surface of the membrane, which binds to matching hooks on the feeder liposomes and triggers fusion.
Excellent questions and next steps
Spudcell demonstrates that many of life’s core processes can be reconstituted from completely specified, individually purified components. Although much remains to be done, the nanovesicle-based feeding approach provides a foundation on which we can build. Some of the challenges remaining to solve include:
- Formation of ribosomes from genetic instructions. Spudcell currently uses ribosomes from E. coli bacteria. Without the ability to remake ribosomes, the spudcell lasts 5–10 generations before the machinery breaks down. Building ribosomes from scratch means synthesizing dozens of protein and RNA molecules, then assembling them in the right order.
- Improving genome distribution. After five generations, about 30% of the daughter cells contain the complete set of seven DNA plasmids. Natural cells solve this with cytoskeletal machinery that pulls chromosomes apart during division. Spudcells do not have this yet, and better genome inheritance would require a more sophisticated splicing mechanism.
- To reduce dependence on external food. Nutrient-carrying liposomes have to be added regularly, and require streptavidin and molecular linker proteins from the outside for partitioning. Making the system more autonomous will require the creation of metabolic pathways that can synthesize components from simple starting materials.
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