How Does Synthetic Biology Design Gene Circuits?

TL;DR

Synthetic biology designs biological circuits by arranging natural or newly built molecular parts into networks with intended functions. A foundational example is a bistable genetic toggle switch in E. coli, where two mutually inhibitory genes create persistent on and off states that can be flipped by a temporary chemical or environmental stimulus.

Transcript

Hi, and welcome to the A16Z podcast. I'm Hannah, and in this episode, general partner on the Bio Fund, Vijay Pande, and I talk all about the field of synthetic biology with Jim Collins, professor of bioengineering at MIT and one of the pioneers of the discipline of synthetic biology. We talk about what engineering and designing biology really looks... Read More

Key Insights

• Synthetic biology is biology by design, using natural parts in new combinations or newly built parts in intended architectures to produce desired functions.
• Genome annotation identifies coding regions, genes, and promoters, but it does not by itself reveal the interconnected networks through which genes, proteins, and RNA elements generate biology and disease biology.
• Systems biology emerged from recognizing that biological complexity must be embraced through network thinking, moving beyond a narrow focus on individual genes, proteins, and pathways.
• Reverse engineering a biological network works by perturbing the system and observing its responses, much like flipping circuit breakers and monitoring appliances to infer the wiring of an unlabeled house.
• Large-scale network reconstruction was initially unrealistic because microarrays were expensive, only seven public datasets were available, and Collins's group lacked a wet laboratory and sufficient perturbation data.
• The genetic toggle switch is a bistable memory circuit built from two mutually inhibitory genes, allowing either gene one or gene two to remain active while suppressing the other.
• A temporary chemical or environmental stimulus can flip the toggle by disabling the active gene, allowing the suppressed gene to activate, produce its repressor protein, and preserve the new state after the stimulus disappears.
• Engineering models can support biological experimentation even when experts expect failure, as Tim Gardner used mathematical and computational modeling before building a functioning toggle switch within nine months.

Questions & Answers

Q: What is synthetic biology and how does it work?

Synthetic biology is an approach to doing biology by design. Researchers arrange molecular parts into networks with a chosen architecture and intended function. They may combine natural components in new ways or build new parts and place them into new or established structures. The approach borrows ideas from engineering, including circuit design, mathematical modeling, computation, perturbation, and the construction of systems whose behavior can be tested experimentally.

Q: Why did synthetic biology emerge from genome research?

Genome research generated parts lists for organisms by identifying genes, coding regions, and promoters, but sequencing could not fully explain how those parts were interconnected. Researchers still needed to understand the networks linking genes, proteins, and RNA elements to biological functions and disease biology. This gap encouraged genome researchers to involve engineers and physicists, whose methods were suited to analyzing complexity, interconnected systems, and network behavior.

Q: How can researchers reverse engineer a biological network?

Researchers can reverse engineer a network by perturbing it and observing which outputs change. Collins compares the process to mapping an unlabeled electrical system in a house: flipping circuit breakers reveals which lights or appliances they control, while running multiple appliances can expose further connections. In biology, carefully stimulating or disrupting a system provides evidence that can be used to infer its underlying wiring and relationships.

Q: Why was large-scale biological network reconstruction initially impractical?

Large-scale reconstruction was impractical because researchers did not yet have enough data or experimental capacity. In the late 1990s, microarrays could measure the expression states of thousands of genes simultaneously, but they were extremely expensive and only seven public datasets were available. Collins's group also lacked a wet laboratory and had no perturbation data, making reliable inference of a large interconnected network unrealistic.

Q: How does a genetic toggle switch store a biological state?

A genetic toggle switch stores state through two genes that repress one another. Both genes tend toward activation, but each tries to shut the other off. This mutual inhibition creates two stable possibilities: gene one is active while gene two is inactive, or gene two is active while gene one is inactive. Because either configuration can persist, the circuit functions as a simple biological memory element.

Q: How can a genetic toggle switch be flipped between states?

The switch can be flipped using a temporary chemical or environmental stimulus that shuts off the gene currently active. Once that gene is disabled, the previously suppressed gene becomes active and produces enough protein to repress the formerly active gene. The stimulus can then be removed, yet the mutual inhibition maintains the new configuration, allowing the system to remain in its changed state.

Q: Why did molecular biologists doubt the genetic toggle switch would work?

Several experienced molecular biologists believed the proposed toggle switch could not function reliably. They argued that the design required two extremely tight off states, which they considered impossible to achieve. They also warned that introducing a large plasmid would burden the cell and that the cell might not tolerate the foreign element. Despite these concerns, Tim Gardner produced a functioning bistable toggle switch within nine months.

Q: What role did engineering play in creating the first genetic toggle switch?

Engineering provided the central concepts and methods behind the toggle switch. The team borrowed the idea of a flip-flop or S latch from electrical engineering, translated it into a mutually inhibitory gene network, and tested its expected bistability through mathematical and computational modeling. Tim Gardner, who had trained in mechanical engineering and initially knew little microbiology, then implemented the modeled design experimentally in E. coli.

Summary & Key Takeaways

• Synthetic biology emerged as genome projects produced lists of genes, promoters, and coding regions without fully revealing how genes, proteins, and RNA elements formed functional networks. Engineers and physicists entered molecular biology because their experience with complex interconnected systems, mathematical modeling, and circuit analysis offered ways to investigate this missing network structure.

• Large-scale biological network reconstruction initially appeared impractical because late-1990s researchers had few expression datasets, costly microarrays, and limited experimental capabilities. Collins and his collaborators therefore changed direction. Instead of inferring an entire natural network, they adopted a tinkerer’s approach and asked whether molecular components could be intentionally assembled into smaller functional circuits.

• The resulting genetic toggle switch used two genes arranged to repress one another, producing two stable states. Although experienced molecular biologists raised credible concerns about leaky off states, plasmid size, and cellular burden, graduate student Tim Gardner built a functioning bistable switch within nine months, demonstrating that engineering principles could guide biological design.

• Key Insights