Researchers at Stanford University and the University of Pennsylvania reported in Science online (early content) on March 28 that they had engineered a genetic circuit that performs like a transistor in bacterial cells. The findings pave the way for “cellular computers” that could operate by using gene functions as circuit components.
The authors of the current work say that most previous approaches to engineering cell-based logic use two-terminal device architectures upon which gate-gate layering, similar to conventional electronics, is used to realize all logic functions. In the Science paper, they described development of device architecture similar to a transistor but that leverages unique properties of genetic regulation to implement all gates without requiring that multiple instances of simpler gates be connected in series.
The design and construction of six basic logic gates consisted of three genes: two encoding the inputs and an output gene containing different configurations of transcriptional control elements (promoters, terminators) flanked by recombinase recognition sites.
The three-terminal device architecture, termed the transcriptor, used bacteriophage serine integrases, enzymes that control the flow of RNA polymerase along DNA. These enzymes modulate inversion or deletion of DNA encoding transcription terminators or a promoter.
For their specific transcriptor design, the investigators constructed low-copy plasmids encoding AND, OR, XOR, NAND, NOR, and XNOR logic elements between a standard strong prokaryotic promoter (input signal source) and green fluorescent protein (GFP) expression cassette (indirect output signal reporter). Measurement of bulk GFP levels from bacterial cultures expressing varying amounts of the integrases that controlled the six Boolean gates showed that exact output levels varied among and within some gates.
Their three-terminal gate architecture, the authors say, decouples logic gate operation from both input and output signals, enabling simple tuning via changes to the transcription input signal and ready reuse—that is, reprogramming of natural transcription systems by seamless integration of transcriptor logic elements within natural operons. Also, by separating gate inputs from gate control signals, and by using a strong input signal modulated by an efficient asymmetric terminator, the authors could demonstrate and quantify signal amplification for all gates.
The authors concluded that with further refinements the single-layer digital logic architecture could enable engineering of amplifying logic gates to control transcription rates within and across diverse organisms.
“We can write and erase DNA in a living cell,” said Jerome Bonnet, a bioengineer at Stanford University. “Now we can bring logic and computation inside a cell itself.” The authors also commented that studies of the reprogramming of aging, cancer, or development would benefit from genetically encoded “counters” capable of recording up to several hundred cell division or differentiation events.