Quantitatively analyze the “gene switch” and explore the mechanism of cell fate determination

Cells can constantly adapt to environmental changes through fate-determined processes to achieve and perfect their own functions. Understanding the specific mechanisms by which cell fate determines is important for answering questions about how complex life is born, tissue and organ regeneration, and synthetic artificial life.

On March 24, Beijing time, a study published in Nature Chemical Biology deeply explored the response of the apparent state of the classical synthetic gene circuit “toggle switch” to changes in cell growth rate through quantitative experiments and mathematical models, and revealed the mechanism behind this phenomenon. This research result helps to better understand the global regulation of cell growth rate on cell fate determination, and provides new research ideas for the application of synthetic biotechnology.

Schematic diagram of decoding a new mechanism by which cell fate determines Source: Photo courtesy of the research team

The study was completed by the team of Fu Xiongfei, Institute of Synthetic Biology, Shenzhen Institute of Advanced Technology, Chinese Academy of Sciences, with researcher Fu Xiongfei as the corresponding author of the paper, assistant researcher Zhu Jingwen and doctoral student Chu Pan as the co-first authors of the paper. The major scientific and technological infrastructure of synthetic biology research provides important support for this research.

Synthetic “gene switches” encode cell fate

Cell fate is affected by gene expression, signaling, growth environment and other factors, and the lack of in-depth understanding of its global, quantitative and theoretical level makes understanding and predicting the direction of cell fate a challenge in the field of biological sciences. Quantitative biology and synthetic biology, which have developed in recent years, provide new ideas for understanding this problem.

Back in 2000, synthetic biology pioneer James Collins built the “Toggle switch” gene circuit. This gene circuit mimics the regulatory logic of gene networks that are widely present in the process of biological development, and is reconstructed in a synthetic way. Through the stimulation of different inducers, the switching of the state of the cell can be controlled, and at the same time, after the inducer is removed, the cell can maintain the current state, just like a two-way toggle switch.

In the process of cell growth and differentiation, cell differentiation leads to changes in cell morphology and specialization of function, which is often accompanied by changes in cell growth rate, so it is difficult to evaluate the impact of cell growth rate on the cell fate determination process as a separate variable. Fortunately, the state switching of the synthetic toggle switch gene circuit does not affect the growth rate of cells, so this gene line just becomes a “sharp weapon” to study this problem.

Inspired by the work of bacterial physiology, the research team observed the bistable state of the “gene switch” under different cell growth rate conditions. The experimental results show that the bistable state of the “gene switch” is related to the growth rate of cells. When the cell growth rate is greater than a certain threshold, the “toggle switch” presents bistability; When the cell growth rate falls below a certain threshold, the bistability of the “toggle switch” bifurcates. This suggests that changes in growth rate may cause steady state changes in gene regulatory networks, which may affect the fate of cells.

In order to deeply explore how changes in cell physiological states dominate the fate determination process of cells, the research team quantitatively characterized the expression levels of two repressor proteins in the “switch” in different growth states, and found that although the expression levels of the two generally showed a trend of negative correlation with the cell growth rate, the maximum peak and relative change values of the expression rate were different. Using mathematical models, the research team evaluated the effect of this inconsistent growth rate-dependent gene expression pattern on the steady state of the “toggle switch”, and proved that the dependence of the growth rate of gene expression brought the possibility of bifurcation to the bistable state of the “toggle switch”.

“The fact that cells can take on different phenotypes after division is a fundamental and age-old question in developmental biology. This study is the first to link growth rate with cell fate, which provides a quantitative theoretical basis and implementation plan for future design of direct or indirect control of cell phenotype by growth rate. Hong Kong Baptist University professor Tang Leihan said.

Cell fate determines new mechanisms

The life course of the cell is a non-equilibrium system, so how does the decision on the fate of the cell occur in a changing environment?

The research team explored this problem by disturbing the cell growth rate, dynamically changing the composition of the medium to achieve cell growth rate switching, and tracking the state of the cell population in real time. The team used potential landscape to quantitatively study the influence of gene expression noise on cell fate decisions at different growth rates. The study found that when the cell growth rate is slower or faster, the energy barrier between the two steady states in the potential energy landscape is low, which means that the cell is more likely to switch states due to noise.

Through cell growth perturbation experiments, researchers observed two different fate determination methods: one is the deterministic mechanism, where changes in the steady-state nature of gene regulatory networks cause complete state switching of cell populations; The second is to be noise-driven, and the jump of the state of the cell population near the critical point to control the state switching of the population occurrence.

This study makes full use of quantitative synthetic biology methods, and fully elucidates the relationship between state selection and growth rate in bistable gene circuits through mathematical model analysis and experimental verification, and provides new insights into the mechanism of cell fate determination. At the same time, the study also points to a possible mechanism by which cellular physiological states influence synthetic gene circuits, and can be used to guide the design of more interference-resistant and predictable gene circuits.

Zhao Guoping, an academician and molecular microbiologist of the Chinese Academy of Sciences, commented that the study studies the regulatory mechanism of cell fate from an innovative perspective, which provides new ideas for quantitatively controlling cell fate for medical and industrial purposes through synthetic biological methods, and has an important impact on its related research fields.

Through the research paradigm of combining quantitative analysis and synthetic reconstruction, quantitative synthetic biology is expected to promote the transformation of life science research from qualitative, descriptive and local research to quantitative, theoretical and integrated.

“We are committed to practicing this research paradigm to increase the understanding of living systems through quantitative experiments and mathematical models, and to further apply the techniques of synthetic biology to analyze the mechanisms of living systems.” Researcher Fu Xiongfei said. (Source: China Science News, Diao Wenhui)

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