Engineered DNA counts it out

Graceful waltzers can count to three, and now stretches of man-made DNA can do it too. Researchers have built a series of genes and put them into bacterial cells, enabling the cells to tally events. The new counters may endow engineered cells with previously impossible functions, the team reports in the May 29 Science.

The engineered counters may be used to monitor toxins in the environment or keep track of the number of times a cell divides. The system can even be programmed to destroy the cell that holds it after a certain number of events.

“This is the first example of a synthetic counter in the field,” says Christina Smolke, a bioengineer at Stanford University and the author of a commentary published in the same issue of Science. Although these new counters are simple, “the first step is building the framework. The next step is, how do we start tailoring these to respond to something relevant? There are a lot of places to take this.”

The new research adds a tool to the burgeoning field of synthetic biology, in which scientists engineer biological systems such as DNA to create new capabilities. DNA molecules are designed to direct certain activities in a cell, and so can respond to specific signals and start and terminate protein production. Since the field emerged in the late 1970s, scientists have been creating artificial cellular “parts” that could be used to modify a living organism or even build a synthetic simple one from scratch. Assembling the right parts in the right order could, for example, allow engineered bacteria to produce biofuels or eat toxins in polluted areas in the environment.

A strong motivator for developing a system that can count, says study coauthor James Collins, was worry over the presence of genetically modified organisms in the environment.

“This came from growing concern that programmed cells could pose a danger to the environment or human bodies. You’d be worried about how long these things were going to stick around,” says Collins, of Boston University. Organisms endowed with counting abilities could be programmed to commit suicide after a certain number of cell divisions or day-and-night cycles, he says. This built-in kill switch may offer a greater level of control over the spread of introduced genes into wild organisms.

These counters rely on the novel assembly of simpler genetic tools. Collins and his team created “multiple numbers of switches cascaded behind one another to create more complex circuits,” says Kaustubh Bhalerao, a biological engineer at the University of Illinois at Urbana-Champaign.

Collins and his colleagues built two systems that count in different ways but are both based on the same basic idea. “Each of the counters is what you call daisy chain cascades: You have to do the first event before you do the next event,” Collins says. This is what endows the systems with the counting ability.

One of the team’s systems counts by starting and stopping the production of certain proteins. In the experiments, the first bit of a strip of modified DNA acts as a detector. When it detects a pulse of the sugar arabinose, it responds by triggering the production of a specific protein. When the DNA detects a second pulse of the sugar, the first protein helps produce a second protein. After a final pulse of the sugar, the second protein helps make green fluorescent protein as an output. When the cells glow green under ultraviolet light, the researchers know that the cells have counted exactly three pulses of sugar. The team could easily make the counting region of the modified DNA longer, allowing for higher counting.

The second counting system relies on enzymes that chop out and invert specific pieces of DNA. When the DNA strip detects the first signal, it causes one of these enzymes to be made. The enzyme then chops its own DNA sequence out of the modified strand of DNA, flips it and reinserts it backward, rendering it unreadable and useless. A second signal leads to the production of another enzyme, which chops another bit of DNA further along on the strand. At the end of the process, an output protein is produced.

The second system can be programmed to respond to different signals at each step of the process. By putting outputs between counts, researchers could track exactly when each step in a series happens.

The first system is better for counting relatively quick events, those that happen every 30 minutes or so. The second system is more useful for counting longer events that unfold over days, because the enzymes need more time to do their cutting and flipping.

Tinkering with the detector and the output, and leaving the basic process intact, may make for innumerable functions, Bhalerao says. Already, some bacteria have DNA that respond to light, arsenic, temperature, nutrients and some metals. In the new counting system, swapping out the signal, such as sugar, to be detected is trivial, says Bhalerao. “It’s like switching brands of mouse on your computer” but leaving the processor alone.

At the other end of the process, the proteins produced after counting can accomplish a wide variety of functions, Collins says. Proteins could “explode the cell, make the cell long, short, fat.” Researchers could even tailor the artificial network to produce different signals — like fluorescent proteins — at different counts. Cells could glow yellow after the first event, red after the second, green after the third and so on. This would allow researchers to monitor every step of complex processes, such as the development and growth of a cell.

The mix-and-match capabilities offer many possibilities, Bhalerao says, but “there is still a long way to go. These things don’t work all the time, and that’s because you’re making the cells do things they don’t want to do.”