Information on what a cell did a few hours or days ago is generally unavailable to researchers, who can only measure the cell's current status. This is a problem if the aim is to understand how immune responses arise or why individual cancer cells survive treatment.
It is particularly revealing to know which genes in a cell are active, as this gives scientists key insights into how cells and the body work. To this end, they measure the RNA molecules – which are transcripts of the active genes but only ever provide a snapshot. However, this means that researchers can only record gene activity at the time of measurement. Moreover, each cell can only be measured once, as the cell is destroyed when the RNA is collected and measured.
Now, researchers at the Department of Biosystems Science and Engineering of ETH Zurich in Basel have developed a method that can be used to archive RNA molecules in cells. For the first time, this allows them to examine not only the current state of a cell, but also to reconstruct which genes were active in the past.
"You no longer see that the response has even taken place"
"Among other reasons, this glimpse into the past is of interest because it tells you about a cell's short-lived responses – of which there are many. Immune and stress responses are just a few examples," explains Christopher Reinkemeier, a postdoc in the group led by ETH professor Randall Platt. In these short-lived responses, an external stimulus activates specific genes. As soon as the stimulus has passed, the cell shuts these genes down again.
As part of a study, the researchers in Basel have shown how human cells in a cell culture respond to a signalling molecule of the immune system. "If you measure too late with the conventional method, you no longer see that the response has even taken place," says Marcos Manero Carranza, a doctoral student in Platt's group and first author of the study alongside Reinkemeier.
How the RNA archive works
For the RNA archive inside the cells, the researchers need to capture RNA molecules and protect them against natural breakdown. This is because molecules of this kind are normally not made to last for long.
Among other things, the scientists achieve this using a protein that occurs naturally in stress granules – accumulations of RNA that form in cells under stress, such as during an infection. In such situations, the cell temporarily shuts down parts of its metabolism and saves RNA molecules for later.
The researchers took this stress granule protein and two other proteins and optimised them using biotechnology. By doing so, they created a system that concentrates a cell's RNA molecules in a film around the cell's mitochondria, where it shields the molecules from naturally occurring enzymes in the cells that break down RNA. The researchers introduce the blueprints for the three optimised proteins into the cells via plasmids, and the cells then assemble the archiving system inside themselves. Per cell, this allows several thousand RNA molecules to be archived over several days.
Researching how cancer recurs
The new method could be used to detect rare events in cells, which is particularly useful if researchers only find out which cells are of interest to them in retrospect. In such cases, they can select these cells and then use the RNA archive to investigate which genes were previously active within them.
The researchers have applied for a patent on the new technology and want to develop it further with a view to future applications in animals and cancer research.
One potential area of application are tumour cells that survive a course of chemotherapy. Cells of this kind can subsequently contribute to a cancer's recurrence. At present, it is very difficult to investigate which cells will survive treatment and how they previously developed.
"Which cells will develop into these persister cells is something that is not known in advance and is poorly understood," says Reinkemeier. "If we can put our system to use in cancer research, we'll be able to isolate persister cells and look into their past. This will allow us to research how these cells form and develop and could pave the way for decisive advances in cancer research."
