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FRETting Over Protein Synthesis in Single Cells

11/01/2013
Jesse Jenkins

Researchers have developed a method for monitoring active synthesis of a specific protein in single cells using FRET. Learn more...


Because changes in protein synthesis or degradation can signal a variety of illnesses, researchers have explored various methods to measure or control translation. But until recently, researchers lacked a general method for measuring the rates of synthesis of specific proteins in single cells.

DiCoMPS reveals synthesis levels of a viral protein in single cells

In a new study published in Nucleic Acids Research (1), a team of scientists from Tel-Aviv University, University of Pennsylvania, and Anima Cell Metrology describe an approach that allows them to measure levels of synthesis for a specific viral protein in individual infected cells. The authors claim that this technique could play a key role in understanding the complex nature of viral-host interactions and may have future applications in live-cell diagnostics.

The team developed the approach, which they call DiCodon Monitoring of Protein Synthesis (DiCoMPS), to observe the translation of non-structural protein 3 (NS3) within cells infected with the Epizootic Hemorrhagic Disease Virus 2-Ibaraki. Marcelo Ehrlich, Associate Professor of the Department of Cell Research and Immunology at Tel-Aviv University, who led the team, said that because viruses shut down host protein synthesis, infected cells present an optimal environment for monitoring viral protein synthesis.

“Here, we have a biological situation in which the virus not only pushes the translation of its messages, but it does so with the background of host protein synthesis shutdown,” explained Ehrlich.

The research team first identified pairs of transfer RNAs (tRNAs) whose usage is enriched in the translation of viral messages relative to cellular messages. Those tRNAs were then fluorescently labeled so the group could directly monitor protein synthesis levels using Förster Resonance Energy Transfer (FRET) signals. The FRET signals generated by the labeled tRNAs were specifically enhanced in infected cells and increased during the course of the infection.

To confirm that they were able to accurately record viral protein synthesis, Ehrlich and colleagues knocked down expression of the NS3 protein with siRNA and observed a subsequent decrease in the FRET signal in infected cells.

Ehrlich said the biggest advantage of this method is that tRNA is a natural component of the translation process. Unlike other approaches currently being tested, Ehrlich believes that relying on tRNA may allow for protein synthesis measurement in live cells and could one day contribute to therapeutic applications.

“That will be the next big step,” Ehrlich said. “I also think that this could be used for diagnostics to address questions related to levels of synthesis in general or possibly the synthesis of a specific protein.”

But for now, Ehrlich’s lab will experiment more generally to see if this method can be modified to work in uninfected cells.

“With improvement of the technique, it could have very practical applications, and with time these things may become possible,” said Ehrlich.

Reference

Barhoom S, Farrell I, Shai B, Dahary D, Cooperman BS, Smilansky Z, Elroy-Stein O, Ehrlich M. Dicodon monitoring of protein synthesis (DiCoMPS) reveals levels of synthesis of a viral protein in single cells. Nucleic Acids Res. 2013 Oct 1;41(18):e177.