A synthetic genetic tool has been developed to quantitatively sense, measure, and manipulate protein aggregation in live cells. A paper published last week in Cell, describes yTRAP (yeast Transcriptional Reporting of Aggregating Proteins) and its use to study a variety of protein aggregates, including disease-relevant proteins, RNA-binding proteins and prions.
Even though protein aggregation is prevalent in biology, many of the causes and consequences are unknown. This is largely because no simple, standardized research tool had previously existed to study this phenomenon in live cells, according to Boston University assistant professor Ahmad S. Khalil along with colleagues from MIT and the Whitehead Institute for Biomedical Research.
Using yTRAP, Khalil and the team created sensors to track aggregation of prions and other proteins, manipulated prions to engineer synthetic memories in cells, identified genes that can cure cells of prions, and enabled high-throughput studies to learn what can influence protein aggregation and its consequences.
The tool is composed of two parts: one piece couples to the protein of interest and the other produces a fluorescent signal to measure the amount of aggregation in a cell. Each piece can be customized to study different proteins or express different genes and signals. For example, the team was able to measure how one prion influenced another by developing a dual sensor that produced either a red or green fluorescent signal depending on how abundant each prion was.
Khalil and his team also demonstrated how the tool can be used to study other proteins, including RNA-binding proteins. Many of these proteins in yeast and humans have similarities to prions, and mutations of those similarities have been linked to neurodegenerative diseases like ALS. With the help of the tool, they uncovered aggregation prone RNA-binding proteins, monitored the consequences of their aggregation, and performed high-throughput screens to see how the aggregation of one protein influences another.
"Protein aggregates can cause a cell to gain or lose a function," says Khalil. "It could be beneficial or harmful. For example, it could allow a cell to survive stressful conditions or change its metabolic function to digest a different type of sugar. And the discovery of these beneficial functions has often been serendipitous." With yTRAP, Khalil hopes to change that.
Image: yTRAP can be used to create cellular sensors for protein aggregation. The sensor produces fluorescent signals that depend on the protein aggregation state in the cell. In this experiment, three cell types were tested that produce varying signal strength depending on the aggregation of a prion protein. The brightest green for no prion aggregation (bottom), a slightly dimmer green to show a weak presence of prion aggregation (top right) and an even dimmer green signal for a strong prion presence (top left). Image courtesy of Ahmad S. Khalil.