Two- and three-input TALE-based AND logic computation in embryonic stem cells. Nucleic Acids Res

Biological computing circuits can enhance our ability to control cellular functions and have poten-tial applications in tissue engineering and medical treatments. Transcriptional activator-like effectors (TALEs) represent attractive components of syn-thetic gene regulatory circuits, as they can be designed de novo to target a given DNA sequence. We here demonstrate that TALEs can perform Boolean logic computation in mammalian cells. Using a split-intein protein-splicing strategy, we show that a functional TALE can be reconstituted from two inactive parts, thus generating two-input AND logic computation. We further demonstrate three-piece intein splicing in mammalian cells and use it to perform three-input AND computation. Using methods for random as well as targeted in-sertion of these relatively large genetic circuits, we show that TALE-based logic circuits are functional when integrated into the genome of mouse embry-onic stem cells. Comparing construct variants in the same genomic context, we modulated the strength of the TALE-responsive promoter to improve the output of these circuits. Our work establishes split TALEs as a tool for building logic computation with the potential of controlling expression of endogenous genes or transgenes in response to a combination of cellular signals