Self-assembled proteins can store cellular “memories” | MIT News

When cells perform their daily functions, they activate a variety of genes and cellular pathways. MIT engineers have now persuaded cells to write the history of these events into a long protein chain that can be imaged using a light microscope.

Cells programmed to produce these chains continually add building blocks that encode particular cellular events. Later, the ordered protein chains can be tagged with fluorescent molecules and read under a microscope, allowing researchers to piece together the timing of events.

This technique could help shed light on the steps that underlie processes such as memory formation, response to drug treatment and gene expression.

“There are a lot of changes that happen at the organ or body level, over hours or weeks, that cannot be tracked over time,” says Edward Boyden, Y. Eva Tan Professor of Neurotechnology. , Professor of Biological and Brain and Cognitive Engineering. science at MIT, a researcher at the Howard Hughes Medical Institute and a member of the McGovern Institute for Brain Research and the Koch Institute for Integrative Cancer Research at MIT.

If the technique could be scaled up to work over longer periods of time, it could also be used to study processes such as aging and disease progression, the researchers say.

Boyden is the lead author of the study, which appears today in Natural biotechnology. Changyang Linghu, a former J. Douglas Tan postdoctoral fellow at the McGovern Institute, who is now an assistant professor at the University of Michigan, is the lead author of the paper.

Cell History

Biological systems such as organs contain many different types of cells, all of which have distinct functions. One way to study these functions is to image proteins, RNA, or other molecules inside cells that provide clues about what the cells are doing. However, most methods for doing this only offer a snapshot of a single moment in time or do not work well with very large populations of cells.

“Biological systems are often composed of a large number of different cell types. For example, the human brain has 86 billion cells,” says Linghu. “To understand these types of biological systems, we need to observe physiological events over time in these large populations of cells.”

To achieve this, the research team came up with the idea of ​​recording cellular events as a series of protein subunits continuously added to a chain. To create their chains, the researchers used modified protein subunits, not normally found in living cells, that can self-assemble into long filaments.

The researchers designed a genetically encoded system in which one of these subunits is continuously produced inside cells, while the other is only generated when a specific event occurs. Each subunit also contains a very short peptide called an epitope tag – in this case the researchers chose tags called HA and V5. Each of these tags can bind to a different fluorescent antibody, making it easier to later visualize the tags and determine the sequence of protein subunits.

For this study, the researchers conditioned the production of the V5-containing subunit on the activation of a gene called c-fos, which is involved in coding new memories. HA-tagged subunits make up the bulk of the chain, but whenever the V5 tag appears in the chain, it means that c-fos has been activated during that time.

“We hope to use this kind of self-assembly of proteins to record activity in each cell,” says Linghu. “It’s not just a snapshot in time, but it also records past history, much like how tree rings can permanently store information over time as wood grows.”

Event registration

In this study, the researchers first used their system to record c-fos activation in growing neurons in a laboratory dish. The c-fos gene was activated by chemically induced activation of neurons, which resulted in the addition of the V5 subunit to the protein chain.

To determine if this approach could work in the animals’ brains, the researchers programmed mouse brain cells to generate protein chains that would reveal when the animals were exposed to a particular drug. Later, the researchers were able to detect this exposure by preserving the tissue and analyzing it under a light microscope.

The researchers designed their system to be modular, so that different epitope tags could be swapped or different types of cellular events could be detected, including, in principle, cell division or cell activation. enzymes called protein kinases, which help control many cellular pathways. .

The researchers also hope to extend the recording period they can achieve. In this study, they recorded events for several days before imaging the tissue. There is a trade-off between recording duration and temporal resolution or event recording frequency, as the length of the protein chain is limited by cell size.

“The total amount of information it could store is fixed, but we could in principle slow down or increase the speed of chain growth,” says Linghu. “If we want to record longer, we could slow down the synthesis to reach cell size in, say, two weeks. This 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, increasing the number of different subunits that can be incorporated.

The research was funded by the Hock E. Tan and K. Lisa Yang Center for Autism Research, John Doerr, National Institutes of Health, National Science Foundation, US Army Research Office, and Howard Hughes Medical Institute.