Supplementary Figure 4: ANDNOT-logic gate module optimization using hammerhead ribozymes.
From: Programmable full-adder computations in communicating three-dimensional cell cultures
a) Logic description of trigger-inducible ANDNOT-logic gate modules responsive to ID, IP and IV with the corresponding truth table. The modules were optimized using relevant inducer combinations as shown using grey boxes (only in the presence of the trigger-inducible transcription factor). b) Performance of three unmodified input-programmable ANDNOT-logic gate modules (tTA/PD, VanR-VP16/PV-OFF, TtgR-VP16/PP-OFF). HEK293T cells were engineered with respective ANDNOT-logic gate components and induced with corresponding inducer molecules. After 48 h, secreted reporter protein SEAP was quantified. OFF-state activities of the reporter SEAP are displayed. c) Performance and schematic symbol of ribozyme-optimized ANDNOT-logic gate modules in which hammerhead ribozymes (HHRs) env140 or sTRSV were installed into the 3’-UTR of indicated reporter plasmids. As illustrated, the SEAP levels in the OFF-state are reduced in the presence of HHRs compared to unmodified modules. Partial data are reproduced from Supplementary Fig. 2b. d) Performance of minimal promoter-optimized ANDNOT-logic gate modules with replaced minimal promoter (human minimal CMV promoter replaced with a minimal TATA-box) in indicated reporter plasmids. As illustrated, the SEAP levels in the OFF-state are reduced when using a minimal TATA-box compared to unmodified modules. e) Performance and schematic symbol of optimized ANDNOT-logic gate modules with installed HHRs and replaced minimal promoters in indicated reporter plasmids. Supplementary Table S2 describes transfection details. Error bars represent the means ± s.d. of n=3 independent experiments performed in triplicate.
