Genes at work: see how protein synthesis is going
Biologists have improved the method of recording gene expressionDmitry Safin, Compulenta
Previously, specialists studying protein synthesis had to work with data averaged over time and over a huge volume of cells. At the end of the last century, a technique was proposed for labeling genes that acquired the ability to provide a fluorescent signal at the time of transcription (the formation of an informational RNA matrix to produce proteins). This made it possible to analyze the functioning of individual genes in bacteria and unicellular organisms, but not in mammalian cells.
The main problem was that copies of the labeled gene delivered to the cell were inserted into the genome randomly: some areas of the latter demonstrate a naturally high rate of transcription, while others, on the contrary, are "silent", and all this prevents the study of the work of the genes themselves. "The genomes of our cell lines contain a sequence that serves as a kind of target for the introduced sequences," says Yaron Shav–Tal, one of the authors of the work under consideration. "Now you don't have to worry about where the gene will end up."
One of the illustrations to the article:
the intensity of the glow of the green fluorescent protein
corresponds to the activity of gene expression at the moment – VM.
The Israelis created two slightly different lines of embryonic kidney cells with an "artificial" version of the cyclin D1 gene that regulates the cell cycle. Both had a DNA sequence that allows the fluorescent protein expressed in cells to bind to cyclin D1 RNA at the time of transcription. At the same time, the first variant of the line contained a natural gene promoter – a landing site of an enzyme (polymerase) that synthesizes informational RNA on a DNA matrix, and the second got a viral promoter that overexpresses genes.
Having prepared the material for the study, biologists traced the influence of the promoter on the dynamics of transcription. As it turned out, in "normal" cells, the process stopped for about 20 minutes every 200 minutes, while cells with a viral promoter remained active for 10 hours. "This is a whole new level of detail," concludes Mr. Shav–Tal. "Even if the cells are considered identical, each of them will still, as we now understand, have its own unique set of expression parameters."
The full version of the Single-allele analysis of transcription kinetics in living mammalian cells report is published in the journal Nature Methods.
Prepared based on Nature News: Watching a gene at work.
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