middleMemories can live as a surprisingly compelling substance in the brain. That is the definition of the word engram, which describes the physical content of how memories are stored. But memories extend beyond the organismal level. Cells also retain memories of the various genes they express over time. Because cis-regulatory elements (CREs) regulate gene expression, measuring the activity of these CREs can provide insight into how genes are expressed in cells. However, Existing technology Cellular gene regulation can only be recorded at a single time point.1
We were motivated to find a way to document the activities of these CREs over time. Jay SendourA geneticist at the University of Washington, he and his team developed a technique called enhancer-driven genomic recording of transcriptional activity in multiplex, or ENGRAM.2 By leveraging prime editing, which allows short, unique DNA sequences to be inserted into specific locations, Shendure and his team were able to trace and record gene regulatory information into genomic locations. Their approach nature, Over time, this could potentially provide insight into how specific genes shape a cell’s identity.2
A CRE is a segment of DNA that controls the expression of nearby genes. When a gene is about to be turned on, a transcription factor binds to a portion of the CRE, starting the process of gene expression. For the ENGRAM system, the team used the CRE to drive a prime editing guide RNA (pegRNA) that inserts a small DNA sequence (a barcode) into a designated location in the genome. Once the CRE is turned on, prime editing follows, creating the corresponding barcode. By assigning a unique barcode to each CRE, the scientists were able to track the activity of different CREs.
The scientists used the same spacer sequence in each pegRNA to ensure that all the barcodes were written to the same location in the genome, creating a barcode “list.” “That’s the key to multiplicity,” he said. Way (Will) ChenCo-author of the paper and a former graduate student in Shendure’s lab, Chen is currently a postdoctoral researcher at the Institute for Protein Design.
Scientists could simply sequence the cellular DNA, examine which barcodes were written, and unravel the cell’s regulatory history. When the scientists tested the system in living cells to see if the number of barcodes written matched the amount of RNA produced by gene expression, they found that the two correlated, confirming that the CRE quantitatively captured the extent of expression.
The scientists then tested whether the ENGRAM recorder system could correctly capture the effects of multiple signals by integrating three CREs that were activated in response to various stimuli, such as drug administration. They found that the recorded barcodes reflected the amount of external signal applied. For example, the more signals, the more barcodes were recorded.
Finally, when the team tested ENGRAM in mouse embryonic cells, they were able to measure the activity of 98 synthetic CREs as the embryonic cells differentiated over time. “Finally, we can map the dynamics of different signals in a time series,” Chen said.
“[The study] “A major limitation in this field is that it requires the ability to record expression levels directly from genes,” he said. Harris KingA Columbia University biologist who was not involved in the study.
Some may worry that the very process of inserting synthetic DNA into the genome to record the activity of regulatory elements could disrupt the normal functioning of the cell. However, Reza KalhorIt’s an unavoidable risk, says a bioengineer at Johns Hopkins University who was not involved in the study. “The Heisenberg principle says we can’t measure something without changing it,” he says.
For Shendure, the key is to uncover the cell’s journey, not just see its final destination. “The vision is that we can get to a world where we make regular measurements of every cell in a system, and those measurements reflect not only the final dynamics of the system, but everything that happened along the way,” he said.
