A Big Step In Synthetic Life Control

The basis of modern computers is the logic gate, a device that makes simple comparisons between the bits, the 1s and 0s, in which computers encode information. Each logic gate has multiple inputs and one output. The output of the gate depends on the inputs and the operation the gate performs.

An AND gate, for example, turns on only if all of its inputs are on. An OR gate turns on if any of its inputs are on.

Suggestively, genes are turned on or off when a transcription factor binds to a region of DNA adjacent to the gene called a promotor.

To make an AND gate out of genes, however, Moon had to find a gene whose activation is controlled by at least two molecules, not one. So only if both molecule 1 AND molecule 2 are present will the gene be turned on and translated into protein.

Such a genetic circuit had been identified in Salmonella typhimurium, the bacterium that causes food poisoning. In this circuit, the transcription factor can bind to the promotor of a gene only if a molecule called a chaperone is present. This meant the genetic circuit could form the basis of a two-input AND gate.

The circuit Moon eventually built consisted of four sensors for four different molecules that fed into three two-input AND gates. If all four molecules were present, all three AND gates turned on and the last one produced a reporter protein that fluoresced red, so that the operation of the circuit could be easily monitored.

In the future, Moon says, a synthetic bacterium with this circuit might sense four different cancer indicators and, in the presence of all four, release a tumor-killing factor.

Crosstalk and timing faults 

There are huge differences, of course, between the floppy molecules that embody biological logic gates and the diodes and transistors that embody electronic ones.

Engineers designing biological circuits worry a great deal about crosstalk, or interference. If a circuit is to work properly, the molecules that make up one gate cannot bind to molecules that are part of another gate.

This is much more of a problem in a biological circuit than in an electronic circuit because the interior of a cell is a kind of soup where molecules mingle freely.

To ensure that there wouldn’t be crosstalk among his AND gates, Moon mined parts for his gates from three different strains of bacteria: Shigella flexneri and Pseudomonas aeruginosa, as well asSalmonella.

Although the parts from the three different strains were already quite dissimilar, he made them even more so by subjecting them to error-prone copying cycles and screening the copies for ones that were even less prone to crosstalk (but still functional).

Another problem Moon faced is that biological circuits, unlike electronic ones, don’t have internal clocks that keep the bits moving through the logic gates in lockstep. If signals progress through layers of gates at different speeds, the output of the entire circuit may be wrong, a problem called a timing fault.

Experiments designed to detect such faults in the synthetic circuit showed that they didn’t occur, probably because the chaperones for one layer of logic gates degrades before the transcription factors for the next layer are generated, and this forces a kind of rhythm on the circuit.

Hijacking a bacterium’s controller