Innovative Spy Protein Allows Researchers Record Cellular “Memories” As They develop
An innovative spy protein or in technical words, a genetically encoded system allows researchers record cellular “memories” as they develop or as cells talk to each other and cellular responses to drugs.
MIT engineers have developed a new method that allows cells to record the activity of their genes and pathways in a long protein chain. Using a light microscope, these events can then be easily visualized.
Construction of these chains, which encode specific biological events, is an ongoing process inside cells trained to create them. Later, the fluorescent molecules may be added to the ordered protein chains, which can then be read under a microscope to reconstruct the sequence of the events.
This method could shed light on the steps behind things like how memories are made, how people react to drugs, and how genes are expressed.
“There are a lot of changes that happen at organ or body scale, over hours to weeks, which cannot be tracked over time,” adds professor Edward Boyden.
Researchers believe that if the method could be made to function over longer time periods, it may be used to investigate phenomena like aging and the development of diseases.
Evolution of cells
There are many different types of cells in living systems like organs, and they all perform specialized tasks. Imaging proteins, RNA, or other molecules within the cells, which provide clues as to what the cells are doing, is one technique to explore their processes. The majority of techniques, meanwhile, either only capture a single point in time or are ineffective when dealing with extremely large populations of cells.
“Biological systems are often composed of a large number of different types of cells. For example, the human brain has 86 billion cells,” adds lead author Changyang Linghu. “To understand those kinds of biological systems, we need to observe physiological events over time in these large cell populations.”
To do this, the research team came up with the idea of keeping track of cellular events as a chain of protein subunits that are added to over time. The scientists employed synthetic protein components that are not ordinarily present in live cells but can self-assemble into lengthy filaments to make their chains.
One of these subunits is continually produced within cells in the system the researchers created using genetic coding, but the other is only made when a certain event takes place. Each subunit also includes an epitope tag, a very short peptide; in this instance, the researchers selected the tags HA and V5. These tags can all attach to various fluorescent antibodies, which makes it simple to see the tags afterwards and identify the order of the protein subunits.
For this work, the researchers conditioned the synthesis of the V5-containing subunit on the activation of the c-fos gene, which is involved in the learning of new memories. Most of the chain is made up of HA-tagged subunits, but when the V5 tag shows up, it means that c-fos was active at that time.
“We’re hoping to use this kind of protein self-assembly to record activity in every single cell,” adds Linghu. “It’s not only a snapshot in time, but also records past history, just like how tree rings can permanently store information over time as the wood grows.”
Noting what happens
In this work, the researchers first recorded c-fos activity in developing neurons in a lab dish using their technique. Chemically induced stimulation of the neurons led the V5 subunit to be added to the protein chain, therefore activating the c-fos gene.
To explore whether this approach could work in the brains of animals, the researchers programmed brain cells of mice to generate protein chains that would reveal when the animals were exposed to a particular drug. Later, the researchers were able to detect that exposure by preserving the tissue and analyzing it with a light microscope.
The researchers designed their system to be modular, so that different epitope tags can be swapped in, or different types of cellular events can be detected, including, in principle, cell division or activation of enzymes called protein kinases, which help control many cellular pathways.
The researchers also hope to extend the recording period that they can achieve. In this study, they recorded events for several days before imaging the tissue. There is a tradeoff between the amount of time that can be recorded and the time resolution, or frequency of event recording, because the length of the protein chain is limited by the size of the cell.
“The total amount of information it could store is fixed, but we could in principle slow down or increase the speed of the growth of the chain,” Linghu says. “If we want to record for a longer time, we could slow down the synthesis so that it will reach the size of the cell within, let’s say two weeks. In that way we could record longer, but with less time resolution.”
The researchers are also working on engineering the system so that it can record multiple types of events in the same chain, by increasing the number of different subunits that can be incorporated.
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