A tunable zinc finger-based framework for Boolean logic computation in mammalian cells

• Published in: Nucleic acids research Vol. 40; no. 11; pp. 5180 - 5187
• Main Authors: Lohmueller, Jason J., Armel, Thomas Z., Silver, Pamela A.
• Format: Journal Article
• Language: English
• Published: England Oxford University Press 01.06.2012
• Subjects: Cell Line, Tumor; Circuits; Construction; Gene Regulatory Networks; Humans; Hybrids; Inteins; Leucine zipper proteins; Mammalian cells; Mathematical Concepts; Nucleotide sequence; Promoters; Protein Engineering - methods; Protein Splicing; Protein Structure, Tertiary; Repressor Proteins - chemistry; Repressor Proteins - metabolism; Repressors; Splicing; Synthetic Biology and Chemistry; Trans-Activators - chemistry; Trans-Activators - metabolism; Transcription factors; Waves; Zinc; Zinc finger proteins; Zinc Fingers
• ISSN: 0305-1048; 1362-4962; 1362-4962
• DOI: 10.1093/nar/gks142

The ability to perform molecular-level computation in mammalian cells has the potential to enable a new wave of sophisticated cell-based therapies and diagnostics. To this end, we developed a Boolean logic framework utilizing artificial Cys(2)-His(2) zinc finger transcription factors (ZF-TFs) as computing elements. Artificial ZFs can be designed to specifically bind different DNA sequences and thus comprise a diverse set of components ideal for the construction of scalable networks. We generate ZF-TF activators and repressors and demonstrate a novel, general method to tune ZF-TF response by fusing ZF-TFs to leucine zipper homodimerization domains. We describe 15 transcriptional activators that display 2- to 463-fold induction and 15 transcriptional repressors that show 1.3- to 16-fold repression. Using these ZF-TFs, we compute OR, NOR, AND and NAND logic, employing hybrid promoters and split intein-mediated protein splicing to integrate signals. The split intein strategy is able to fully reconstitute the ZF-TFs, maintaining them as a uniform set of computing elements. Together, these components comprise a robust platform for building mammalian synthetic gene circuits capable of precisely modulating cellular behavior.