A Better Way to See How Brain Cells Falter in Disease

To gain better insight into what’s happening in the brain, researchers examine the molecules produced by brain cells, including RNA and proteins. But existing methods for molecular profiling don’t always capture the cells’ full complexity.

Now, a new technique developed by the Rangaraju Lab at Yale School of Medicine (YSM) and the Sloan Lab at Emory University School of Medicine aims to improve molecular profiling by simultaneously comparing protein and RNA levels across individual cell types in living tissues.

According to a study published March 31 in Nature Communications, this method uses an enzyme called TurboID to chemically label protein and capture interacting RNA. This technique could be used to better understand the inner workings of cells affected by neurodegenerative diseases, such as Alzheimer’s, as well as to conduct research beyond neuroscience, says Srikant Rangaraju, MD, a physician-scientist and associate professor of neurology at YSM.

“The brain is one of the most complex organs in the body,” he says. With this new method, “we can measure both RNA and protein levels simultaneously from the same animal and the same tissue without needing to run duplicate experiments.”

All cells in the body have the same set of genetic instructions. How cells end up differentiating—becoming things like neurons, skin cells, or hair—depends on how those instructions are executed.

RNA and proteins are the two main ways that cells carry out DNA’s orders. Different cell types have distinct combinations of RNA and proteins, allowing researchers to use their profiles to understand how cells function and what happens to them in disease. However, not all techniques are reliable for measuring these molecules.

In 2021, Christina Ramelow, PhD, now a postdoctoral researcher at Emory University, started working in Rangaraju’s lab. His research group was broadly interested in studying stroke and Alzheimer’s disease. One way they were doing this was by looking at the molecular profiles of brain cells.

But while the team had good techniques for collecting RNA data from brain cells, they struggled to obtain accurate protein measurements, partly because isolating brain cells for study often altered or damaged them and their proteins. This was a problem, because “one of the best surrogates of cell function is protein level,” says Rangaraju.

The team wanted to know whether they could develop a new technique to better understand the proteome—the complete set of proteins expressed by a cell—in brain cells. To do this, the researchers turned to an enzyme called TurboID, first identified by Stanford researchers.

TurboID chemically “tags” proteins, “basically like adding a zip code,” says Ramelow. This chemical signature allows researchers to isolate proteins. But the researchers also noticed that TurboID tagged proteins that interact with RNA, opening the possibility of also getting information on RNA in the process—something that, if it worked, could make molecular profiling more efficient.

To test TurboID, the researchers used the enzyme to generate proteomes and transcriptomes (the complete sets of RNA in a cell) of a well-studied cell line. They then compared the list generated by their method to the existing map of proteins and RNA found in the cell line. The list matched, suggesting that TurboID could work.

Next, the team needed to see how TurboID would perform in a living brain. To do this, they genetically altered mice to express TurboID in either neurons or another type of brain cell called astrocytes. The technique captured RNA and protein profiles in both cell types.

These results suggest that the technique—dubbed Simultaneous Protein and RNA-Omics, or SPARO—can be used to look at the molecular profiles of brain cells, as well as cells in other parts of the body, says Rangaraju.

There’s also reason to think that the proteome produced by SPARO may be more accurate than other profiling techniques. In a living brain, cells are constantly communicating. Their protein levels should reflect these interactions, but extraction can damage or alter their communications. This technique should provide a better snapshot of brain cells as they are in their natural environment, says Ramelow.

The team expects to use SPARO in their own research. But the technique can also be used in neuroscience more broadly, and even beyond. The molecular profile of cells is one of biology’s “fundamental questions,” says Ramelow. “There’s so much to learn by applying this method very broadly in the field.”