A team of University of California San Diego (UCSD) scientists has genetically rewired the circuit that controls cell aging in yeast. From its normal role functioning like a toggle switch, they engineered a negative feedback loop to stall the cellular aging process. The rewired circuit operates as a clock-like device, called a gene oscillator, that drives the cell to periodically switch between two detrimental “aged” states, avoiding prolonged commitment to either and thereby slowing the cell’s degeneration. Their approach dramatically extended cellular lifespan, setting a new record for life extension through genetic and chemical interventions.

The findings represent a proof-of-concept example of using synthetic biology to reprogram the cellular aging process. Given that the underlying aging pathways are conserved, the findings may one day enable the design of synthetic gene circuits that promote longevity in more complex organisms.

“This is the first time computationally guided synthetic biology and engineering principles were used to rationally redesign gene circuits and reprogram the aging process to effectively promote longevity,” said Nan Hao, PhD, of the School of Biological Sciences’ Department of Molecular Biology, and co-director of UC San Diego’s Synthetic Biology Institute.

Hao is senior author of the group’s published study in Science, titled “Engineering longevity—design of a synthetic gene oscillator to slow cellular aging,” in which they  concluded “Our results establish a connection between gene network architecture and cellular longevity that could lead to rationally designed gene circuits that slow aging.”

Human lifespan is related to the aging of our individual cells, and cellular aging is a fundamental and complex biological process and is an underlying driver for many diseases. Cells, including those of yeast, plants, animals and humans, all contain gene regulatory circuits that are responsible for many physiological functions, including aging. “These gene circuits can operate like our home electric circuits that control devices like appliances and automobiles,” said Hao.

However, the UC San Diego group had previously uncovered that, under the control of a central gene regulatory circuit, cells don’t necessarily age the same way. Several years ago the research team began studying the mechanisms behind cell aging. Using Saccharomyces cerevisiae yeast as a model for human cell aging, they discovered that cells follow a cascade of molecular changes through their entire lifespan until they eventually degenerate and die. But the scientists noticed that cells of the same genetic material and within the same environment can travel along distinct aging routes. About half of the cells age through a gradual decline in the stability of DNA, where genetic information is stored. The other half ages along a path tied to the decline of mitochondria, the energy production units of cells.

After identifying two distinct directions that cells follow during aging, the researchers genetically manipulated these processes to extend the lifespan of cells. For their newly reported work the team extended their research using synthetic biology to engineer a synthetic gene oscillator that keeps cells from reaching their normal levels of deterioration associated with aging.

Engineered cells show oscillating abundance of a master aging regulator. [Hao Lab, UC San Diego]

Imagine a car that ages either as the engine deteriorates or as the transmission wears out, but not both at the same time. The UC San Diego team envisioned a “smart aging process” that extends cellular longevity by cycling deterioration from one aging mechanism to another.

As electrical engineers often do, the researchers first used computer simulations of how the core aging circuit operates. This helped them design and test ideas before building or modifying the circuit in the cell. This approach has advantages in saving time and resources to identify effective pro-longevity strategies, compared to more traditional genetic strategies.

For their research and tests using Saccharomyces cerevisiae yeast cells the team developed and employed microfluidics and time-lapse microscopy to track the aging processes across the cell’s lifespan. To control aging in the yeast cells the team manipulated the expression of two conserved transcriptional regulators: silent information regulator 2 (Sir2), which drives nucleolar decline, and heme activator protein 4 (Hap4), which is associated with mitochondrial biogenesis.

The expression of Sir2 and Hap4 are linked in that expression of one cross-represses the other. The result is a naturally occurring and widely conserved transcriptional toggle-switch that drives cellular fate decisions. The authors newly reported work describes how they engineered a synthetic gene oscillator within yeast cells that re-wires this transcriptional toggle switch to generate sustained oscillations between the two states of cellular degeneration in individual cells. By creating a negative feedback loop in the Sir2-HAP circuit, the synthetic oscillator delays the commitment of yeast to one of the two cellular deterioration states. “These oscillations increased cellular life span through the delay of the commitment to aging that resulted from either the loss of chromatin silencing or the depletion of heme.”

They found that cells containing the synthetic gene oscillator lived considerably longer than wild-type cells, exhibiting an 82% increase in lifespan.  “This is the most pronounced life-span extension in yeast that we have observed with genetic perturbations,” the team noted.

During the process of circuit engineering the investigators also constructed and characterized versions of the synthetic circuit with broken or weakened feedback interactions. “None of these circuits enabled sustained oscillations in a major fraction of cells, which demonstrated the importance of connectivity and strength of feedback interactions in generating oscillations,” they pointed out.

The new synthetic biology achievement has the potential to reconfigure scientific approaches to age delay. Distinct from numerous chemical and genetic attempts to force cells into artificial states of “youth,” the new research provides evidence that slowing the ticks of the aging clock is possible by actively preventing cells from committing to a pre-destined path of decline and death, and the clock-like gene oscillators could be a universal system to achieve that.

“This is the first time computationally guided synthetic biology and engineering principles were used to rationally redesign gene circuits and reprogram the aging process to effectively promote longevity,” said Hao. “Our oscillator cells live longer than any of the longest-lived strains previously identified by unbiased genetic screens.”

The researchers added, “Our results establish a connection between gene network architecture and cellular longevity that could lead to rationally designed gene circuits that slow aging … Our work represents a proof-of-concept example, demonstrating the successful application of synthetic biology to reprogram the cellular aging process, and may lay the foundation for designing synthetic gene circuits to effectively promote longevity in more complex organisms.”

The team is currently expanding their research to the aging of diverse human cell types, including stem cells and neurons.

In a related Perspective, Howard Salis, PhD, at Pennsylvania State University, discusses the study in greater detail. He noted that, as Hao et al noted, one road to understanding and controlling cellular aging is to measure the dynamics of pathways that control cellular maintenance and aging, develop system-wide models, and apply mathematical analysis to pinpoint what he calls “the tunable knobs and swappable wires” that can be manipulated to redirect a cell’s natural dynamics away from aging and toward the maintenance of healthy cell states. “By com­bining system-wide models with engineered genetic systems, candidate thera­peutics could be developed—for example, a small-molecule inhibitor that pushes cell dynamics away from dysfunctional states or a combination strategy that removes senes­cent cells and replaces them with improved cells through ex vivo therapy.” And reflecting on how the results may inform development of human therapeutics, Salis wrote, “If the collective objective of these interven­tions is to maintain healthier cell states, then the risk and morbidity of age-associated dis­eases will be reduced.”

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