Life on the microscopic scale is in continual flux. It may take thousands of years for new species of mammals to form, whereas individual cells are capable of evolving much faster. Whether it is the emergence of antibiotic-resistant bacteria or a cancerous tumor, cellular evolution shapes life at every level. Thanks to one Stony Brook group, scientists are now able to track thousands of cell lineages—complete with evolutions and developments—simultaneously and in high detail.
Published last month in the journal Nature, Sasha Levy and his colleagues developed a DNA barcoding system that enables quantitative monitoring of entire populations of cells. Levy is an assistant professor of physical and quantitative biology at the Louis and Beatrice Laufer Center at Stony Brook University.
“Essentially, barcoding is a way to get a last name into the genome. And that last name is just a random piece of DNA,” Levy said in an interview. “It is sort of a nonsensical name, but what it signifies is a lineage that gets passed from generation to generation. Fission yeast grows asexually so there’s never a mixing of last names.”
Levy and his colleagues developed the barcoding technique in Saccharomyces cerevisiae, commonly known as baker’s yeast—the gold-standard model organism for studying eukaryotic cell growth. The yeast grows while floating in nutrient broth, forming suspended cell cultures.
“With yeast, history happens much faster, because their generation time is 90 minutes,” Levy said. “In the lab we can look at 200 generations and try to understand how those populations evolved and competed—who expanded and who got wiped out.”
Each DNA barcode consists of about 20 random base pairs inserted specifically into each cell’s genome.
For each generation the fission yeast replicates, the barcode is copied faithfully into two identical daughter cells along with the rest of their genome.
“Occasionally there are errors in sequencing, so we wanted to make the names different enough such that if the yeast make a typographic error you won’t confuse the names,” Levy said.
Because yeast has a fixed replication time of 90 minutes, the researchers could allow the yeast colonies to grow a set number of generations. Afterward, genomic DNA was extracted from cells, sequenced with next-generation techniques and quantified using a computer software.
“By looking at the trajectories of the fastest growing colonies we can try to figure out what happened to those colonies, when it happened, and how good was the mutation that made its growth better,” Levy said. “These trajectories increase at a rate that tells us exactly how much better a particular mutation is, since the ones that increase fitness do not follow a typical pattern.”
One of the next steps will be trying to use the yeast barcode system to study protein-protein interactions in living cells.
“With yeast you essentially have the whole protein interactome in one tube,” Levy said. “We are working on testing millions of protein interactions simultaneously and asking what is happening in a particular environment.”
Levy said DNA barcoding is not restricted to yeast, but can also be applied to other cell types.
“We think this technology will be useful in seeing how microbes become resistant to antibiotic drugs or how cancers develop into tumors,” he said.
Because cellular evolution is fundamental to life, the ability to trace individual cell lineages with high precision and sensitivity could help shed insight on the mechanisms behind abnormal growth and perhaps offer a way to stop that process in its tracks.
