Researchers from MIT developed a technique to induce cells to record the history of cellular events in a long protein chain that can be imaged using a light microscope. The technique could help understand the critical steps involved in the processes, such as memory formation, response to drug treatment, and gene expression.
Studying the molecular processes within cells can provide important insights into their function and how they contribute to the overall functioning of an organ. However, most techniques for imaging cells only allow researchers to obtain a snapshot of a single moment in time, which can be limited in understanding the dynamic processes occurring within cells.
“Biological systems are often composed of a large number of different types of cells. To understand those kinds of biological systems, we need to observe physiological events over time in these large cell populations,” said Changyang Linghu, Assistant Professor at the Michigan Neuroscience Institute and author of the study.
Cellular history
A research team from MIT came up with the idea of recording cellular events as a series of protein subunits that are continuously added to a chain. To create their chains, they used engineered protein subunits that can self-assemble into long filaments. These protein subunits are absent in living cells.
Using encoding, researchers designed a system to continuously produce one of the protein subunits within cells while the other subunit is only produced in response to a specific event.
The subunits are labeled with short peptides called epitope tags (HA and V5 in this case) that can bind to fluorescent antibodies, allowing researchers to easily visualize and determine the sequence of the protein subunits.
In this study, the V5-tagged subunit was only produced when a gene called c-fos was activated, which encodes new memories. The majority of the protein chain consists of HA-tagged subunits, but whenever the V5 tag showed up, it indicated that the c-fos gene was activated at that specific time.
“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,” said Linghu.
Recording cellular events
Researchers first used their system to record the activation of c-fos in neurons growing in a lab dish. They did this by using a chemical to activate the neurons, which caused the V5 subunit to be added to a protein chain.
To test whether this approach could be used in the brains of animals, researchers modified brain cells in mice to produce protein chains that would reveal when the animals were exposed to a particular drug. They detected this exposure by analyzing preserved tissue with a light microscope.
A flexible design
The system is designed to be flexible. Different epitope tags can be used to detect various cellular events, such as cell division or the activation of protein kinases.
The researchers also hope to extend the recording period. This study recorded events for several days before imaging the tissue. However, there is a trade-off. Increasing the recording time will decrease the time resolution or frequency of event recording.
“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,” Linghu says.
The researchers are working on engineering the system to record multiple events in the same chain by increasing the number of different subunits that can be incorporated.
The study is published in Nature Biotechnology.
Study abstract:
Observing cellular physiological histories is key to understanding normal and disease-related processes. Here we describe expression recording islands—a fully genetically encoded approach that enables both continual digital recording of biological information within cells and subsequent high-throughput readout in fixed cells. The information is stored in growing intracellular protein chains made of self-assembling subunits, human-designed filament-forming proteins bearing different epitope tags that each correspond to a different cellular state or function (for example, gene expression downstream of neural activity or pharmacological exposure), allowing the physiological history to be read out along the ordered subunits of protein chains with conventional optical microscopy. We use expression recording islands to record gene expression timecourse downstream of specific pharmacological and physiological stimuli in cultured neurons and in living mouse brain, with a time resolution of a fraction of a day, over periods of days to weeks.
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