UK researchers have developed a method to turn gene expression on or off using multiple wavelengths of light. Their technique could help other scientists study complex cellular processes and advance synthetic cell research and technologies.

In cells, DNA is like a cookbook. It contains all the instructions necessary for the cell to function properly. These instructions are read and processed by the cells during the process of gene expression, to produce RNA molecules and proteins. RNA is primarily involved in the production of proteins, but it has other functions and proteins are the doers of the cell. Proteins are behind almost all cellular activities: cell division, the cell’s ability to sense its environment and adapt, communication with other cells, CO2 capture, etc.

Scientists working in the field of synthetic cell research seek to reproduce the mechanisms that occur in the cells. What if, once they have assembled their cell-like structure, that is, once they have encapsulated all the components in a droplet or vesicle, they could control from the outside what happens on the inside, turning on or off certain processes. This could help them study complex cellular processes and advance research in this area. The team of Michael Booth, a Royal Society University Research fellow at University College London, UK, has developed a method to turn gene expression on or off using two different wavelengths of light.

Remote activation of gene expression by light

Michael Booth explained: “We had previously shown that we could control DNA with light: we attached photocages, small light-sensitive chemical molecules, to the DNA, which completely blocked expression. However, when we shone UV light on the DNA we succeeded in activating the process of gene expression.”

But UV light is not ideal for applications such as drug delivery: it can be damaging and does not penetrate tissue. Denis Hartmann, a PhD student in the Booth group, then synthesized a new blue-light photocage that can activate expression using a less toxic visible wavelength.

“We built two activation buttons that are orthogonal, so they don’t interact with each other. With UV and blue photocages, we were able to activate two different genes, depending on the wavelength used. And we went further: we combined the two light-activated DNA templates to create an AND logic gate.”

The AND gate device allowed an output only when the two wavelengths of light were applied: “We used the AND gate in synthetic cells to activate gene expression only in a specific region where the two wavelengths of light intersected, allowing us to have a precise spatiotemporal control over this process.”

Remote shutdown of protein production by light

Can we imagine a drug inspired by synthetic cell research that, once in the patient’s body, would start producing proteins when the doctor gave the order and at a specific location? Once enough proteins have been created, the doctor could send a different signal to stop the process.

In another set of experiments, led by Giacomo Mazzotti and Denis Hartman, and published in the Journal of the American Chemical Society, Michael’s team created an “off button” and combined it with the “on button”.

“We used ASOs [antisense oligonucleotides], which are short sequences of DNA that, when they bind to RNA, initiate the degradation of the RNA, in the presence of an RNase protein, and thus stops protein production,” said Michael.

“We made our ASOs and then attached our photocages to them so that we could orthogonally control the degradation of different RNA using UV or blue light. With these blue and UV light photocages, we had two orthogonal stop buttons and, from our previous experiments, two orthogonal activation buttons. So we then combined an activating button with a stop button and were able to start the production of a specific protein using the blue light and stop its production using the UV light.”

Next steps: near-infrared light, temperature, and magnetism

The team is currently conducting experiments with near-infrared photocages, which are more tissue-penetrating. They are also exploring other stimuli to activate gene expression, such as temperature and magnetic stimuli, both of which can penetrate tissues and are therefore interesting for future in vivo applications.

More information

• Orthogonal Light-Activated DNA for Patterned Biocomputing within Synthetic Cells
  Denis Hartmann, Razia Chowdhry, Jefferson M. Smith, and Michael J. Booth
  Journal of the American Chemical Society 2023 145 (17), 9471-9480
  DOI: 10.1021/jacs.3c02350
• Precise, Orthogonal Remote-Control of Cell-Free Systems Using Photocaged Nucleic Acids
  Giacomo Mazzotti, Denis Hartmann, and Michael J. Booth
  Journal of the American Chemical Society 2023 145 (17), 9481-9487, DOI: 10.1021/jacs.3c01238
• Light-activated communication in synthetic tissues
  Michael J. Booth et al., Science Advances, 2016, Vol 2, Issue 4, e1600056(2016), DOI:10.1126/sciadv.1600056

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