Stanford University researchers have devised a new technique to control protein production

Thanks to new RNA vaccines, we humans have been able to protect ourselves incredibly quickly from new viruses like SARS-CoV-2, the virus that causes COVID-19. These vaccines insert a piece of ephemeral genetic material into the body’s cells, which then read its code and produce a specific protein — in this case, telltale “spikes” that stud the outside of the coronavirus — causing the immune system to fight future invaders.

The technique is effective, and it’s promising for all kinds of treatments, says Eric Cassiniet, a bioengineering doctoral student at Stanford University. Currently, these types of RNA therapies cannot focus on specific cells. Once injected into the body, they make the protein encoded randomly in every cell they enter. If you want to use them to treat only one type of cell — such as those inside a cancerous tumor — you’ll need something more precise.

Cassiniet and his advisor, associate professor of chemical engineering Xiao Jing Zhao, may have found a way to make it happen. They have created a new tool called an RNA “sensor” – a strand of RNA manufactured in a laboratory that reveals its contents only when it enters certain tissues inside the body. The method is so precise that it can settle in both cells Species and the cell States, It only activates when its target cell is creating a specific RNA, says Zhao. The pair published their findings on October 5 in the journal nature biotechnology.

For the first time, you can have only the cells in question directly produce a protein under very specific conditions. This kind of accuracy wasn’t previously possible.”

Xiaojing Gao, Assistant Professor of Chemical Engineering

The protein produced could be an antigen — a foreign substance that elicits an immune response — as in the case of vaccines, an enzyme that restores function to a broken cell, a fluorescent protein that can be used to track specific cells in a research study, or a protein that causes cell death to remove pathogenic or non-pathogenic cells. Desirable, among other possibilities.

Raising the level of the immune system

The pair’s new system, called RADAR, essentially consists of two parts: a “sensor” region that sticks to specific RNA molecules inside the body, and a “payload” region that the cell will read and convert into a protein. The two sections are separated by a stop codon, which is the part of the RNA sequence that renders part of the RADAR genetic code inaccessible.

If the RADAR sensor area succeeds in reaching its target, the stop symbol will disappear, suddenly making the remaining area – its “payload” – readable. In theory, this payload could carry instructions for making any protein, in any cell type, at any time.

This process happens thanks to an existing set of enzymes called ADAR (Adenosine Deaminases Acting on RNA) — a byproduct of the ongoing viral arms race that has raged within the human body for thousands of years, says Gao.

Some viruses, such as SARS-CoV-2, influenza, and norovirus, are just a protein shell with RNA inside. In the process of reproduction, these viruses create very long stretches of double-stranded RNA. Because viruses can have devastating effects on the body, our immune system has gradually learned to see those double-stranded RNAs as a threat and will quickly shut them down.

“It’s kind of a danger signal — if a cell saw the double-stranded RNA, it would immediately panic about it,” Cassiniet says.

But in a strange twist of evolution, our bodies also Make double-stranded RNA. As viruses have attacked us over thousands of years, digging into our cells and playing with our genetic machinery, some of their genes have been absorbed and incorporated into our DNA. (This is no coincidence: it has happened so many times in the past that today the human genome is roughly 8% a virus.)

To solve this problem, ADAR is being developed as a kind of “testing” system – a way for the body to tell if a piece of double-stranded RNA is friend or foe. If it finds one generated by our genome, ADAR edits it slightly to make it look less menacing, causing holes or gaps between the two threads to open, like removing a few stitches in the middle of a fabric seam. The immune system, which has a larger fish for frying, immediately discards this coarse RNA and continues to fight the real enemy.

RADAR takes advantage of this mechanism. When its “sensor” unit attaches to a specific target molecule (another piece of RNA), ADAR sees the resulting double-stranded pair as a friendly, harmless species and faithfully edits it for the immune system to ignore. In the process, the precise molecular “stop” tag that the researchers created in the middle of the RNA strand erases it. Once removed, the payload section of RADAR becomes visible to the cell, and the code it contains turns into a protein.

Potential for new programmable therapies

Currently, Kaseniit, Gao, and their collaborators are still testing RADAR in a variety of settings, but the results look promising. With co-authors, Associate Professor of Chemical Engineering Elizabeth Satelli, and postdocs Diego Winger and Will Cody, they tried it even in plants, which don’t naturally have ADAR systems — but after adding ADAR enzymes to the mix, they were able to get the same Results. And they say the flexibility and accuracy of RADAR in the future could provide a valuable tool in both research and medicine, giving scientists a way to get rid of specific cells in the lab or deliver treatments inside the body.

“This is the hope and dream of RNA as a platform, because you can just encode any protein you want on a piece of RNA and the cells will make it. Now with these controls, we can determine which target cell it will be activated in. This is very powerful,” Cassiniet says.


Journal reference:

Kaseniit, K.E., et al. (2022) Programmable RNA sensor modules using ADAR editing in living cells. Nature Biotechnology.

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