Alignment by feedback and comparator-adjuster. (A) Control theory diagram, where “amplifier” is a device to be controlled and “comparator-adjuster” compares input to output and adjusts signal in proportion to the difference. (B) Biochemical reaction network where nodes have active and nominally inactive states. Feedback alone cannot produce alignment in such a network, but can if network contains a comparator-adjuster, shown with an unspecified mechanism. (C) A human-built cell system that aligned output with variant input using feedback and a comparator-adjuster (Nevozhay et al. (2009). An inhibitor, anhydrotetracycline (ATc) diffuses into (and out of) yeast cells slowly. ATc binds and inactivates tetracycline repressor, TetR. If ATc level rises so that intracellular ATc exceeds TetR, then all TetR is bound, while some ATc is free. Because all TetR is inactivated, it does not repress yEGFP expression, so system output increases. Meanwhile, TetR synthesis driven by an identical promoter is derepressed. Once total TetR concentration exceeds that of ATc, some TetR remains free and active. Free TetR represses yEGFP expression, capping yEGFP synthesis and TetR synthesis at a new, higher level. The comparator uses binding between ATc and TetR to compute their concentration difference, and the adjuster (free TetR) aligns system output (yEGFP and total TetR) with the input, ATc.