Breakthrough for ‘smart cell’ design

“Imagine that microprocessors made of proteins within cells can ‘decide’ how to respond to specific signals such as inflammation, tumor growth markers, or blood sugar levels,” said Xiaoyu Yang, a doctoral candidate in systems, synthetic and physical biology. project at Rice University, where he is the lead author of the study. “This work brings us one step closer to building ‘smart cells’ that can detect signs of disease and immediately issue customizable treatments.”

A new approach to the design of artificial cellular circuits relies on phosphorylation—a natural process that cells use to respond to their environment, characterized by the addition of phosphate groups to proteins. Phosphorylation is involved in a wide range of cellular functions, including the conversion of extracellular signals into intracellular responses such as movement, secretion of substances, response to pathogens, or expression of genes.

In multicellular organisms, phosphorylation-based signaling often involves a multi-stage cascade of effects, like a falling domino. Previous attempts to exploit this mechanism therapeutically in human cells have focused on redesigning natural, existing signaling pathways. However, the complexity of these pathways makes them difficult to use and thus their applications remain rather limited.

However, phosphorylation-based innovations in “smart cell” engineering may grow significantly in the coming years, thanks to new discoveries by Rice University researchers. What enabled this breakthrough was a change in perspective:

Phosphorylation is a continuous process that unfolds in a series of interconnected cycles, from cellular input (i.e., what the cell encounters or senses in its environment) to output (what the cell responds to). What the team realized and set out to prove was that each loop in the cascade could be considered a basic unit, and these units could be connected together in new ways to create entirely new pathways connecting the cell’s inputs and outputs.

“This greatly opens up the design space for signaling circuits,” said Caleb Bashor, assistant professor of bioengineering and biological sciences and corresponding author of the study. “It turns out that the phosphorylation cycles are not only interconnected, but interconnected – something we weren’t previously sure could be done with this level of complexity.

“Our design strategy allows us to engineer synthetic phosphorylation circuits that are not only highly tunable but also operate in parallel with the cell’s own processes without affecting its viability or growth rate.”

While this may sound simple, figuring out the rules for how to build, connect, and tune these units—including the design of intracellular and extracellular outputs—is anything but. Furthermore, the fact that synthetic circuits can be constructed and implemented in living cells is not a given.

“We didn’t necessarily expect our synthetic signaling circuits, composed entirely of engineered protein parts, to perform at a similar speed and efficiency to natural signaling pathways found in human cells,” Yang said. “Needless to say, we were surprised to find that this was indeed the case. So. It takes a lot of effort and collaboration to achieve this.”

It turns out that a do-it-yourself modular cellular circuit design approach can reproduce the important system-level ability of natural phosphorylation cascades to amplify weak input signals into macroscopic outputs. Experimental observation of this effect validates the team’s quantitative model predictions and reinforces the value of the new framework as a fundamental tool in synthetic biology.

Another clear advantage of new approaches to the design of sensing and responding cellular circuits is that phosphorylation occurs rapidly within seconds or minutes, so new synthetic phospho-signaling circuits may be programmed to respond to physiological events occurring on similar time scales. In comparison, many previous synthetic circuit designs are based on different molecular processes, such as transcription, which can take hours to activate.

The researchers also tested the circuit’s sensitivity and ability to respond to external signals such as inflammatory factors. To demonstrate its translational potential, the team used the framework to design a cellular circuit that could detect these factors and could be used to control autoimmune episodes and reduce immunotherapy-related toxicities.

“Our study demonstrates that programmable circuits that respond quickly and accurately to signals can be created in human cells, and is the first report of a building kit for engineering synthetic phosphorylation circuits,” Bashor said. Established earlier this year to leverage Rice’s deep expertise in the field and promote collaborative research.

Institute Director Caroline Ajo-Franklin said the study’s results are an example of the transformative work Rice researchers are doing in the field of synthetic biology.

“If over the past 20 years synthetic biologists have learned how to manipulate the way bacteria gradually respond to environmental cues, the work in Bashor’s lab brings us to a new frontier,” Ajo-Franklin said. —Control the immediate response of mammalian cells to changes.