Implementation Outline: Goal 1 embedded with a signalling molecule Bacteria start off with receptors for this Once first bacteria reach goal, they secrete chemoattractant and an Autoinducer (AI) Once enough bacteria are attracted to the goal, the AI triggers a switch to chemorepellant release. Also, receptors replaced with those for compound at goal 2. Bacteria move off to goal 2 START Goal 1 Goal 2
Toggle Switch B0034 R0011: Promotor, repressed by LacI. C0012: LacI R0051: Promotor, lambda cI repressed C0051: cI B0010 B0012 Initial state: LacI is produced, cI is repressed.
Checkpoint 1: AgrC AIP AgrAAgrA - P R0051 cI repressed AttractantLuxIRepellant R0011 lacI repressed Checkpoint embedded with AIP Arrival triggers AgrA - P production This activates attractant production ( cI is low) and production of the LuxI autoinducer When enough bacteria arrive, LuxI triggers switching of the toggle (next slide) lacI falls and repellant is produced C0061
Switching: R0062: Activated by LuxR and HSL cI AgrA ComA R0051: cI Repressed R0011: LacI Repressed Activation of cI expression should cause switching of toggle. LacI goes low, therefore ComA (part of signalling pathway for molecule at next checkpoint) can be expressed cI stops production of AgrA.
Checkpoint 2: ComP ComX ComAComA - P Attractant R0011: LacI Repressed LuxM R0051 cI repressed Repellant On arrival: LacI = Low, cI = high Promotor activated by LuxO - P (present when LuxM is low) Q00400: Tet inverter LacR