A composite feed-forward loop I4-FFL involving IHF and Crc stabilizes expression of the XylR regulator of Pseudomonas putida mt-2 from growth phase perturbations

Genetic networks are typically composed of a series of connected motifs that confer specific logic and dynamic properties to the resulting circuits. While some feed forward loop (FFL) variants abound in such networks, others (e.g. the type-4 incoherent FFL or I4-FFL) are virtually absent from the known regulatory devices. We report here that the key node that rules the expression of the m-xylene biodegradation pathway of the soil bacterium Pseudomonas putida mt-2 merges opposite physiological effects of the growth phase by means of a regulatory device based on the rarely found I4-FFL motif. Specifically, the FFL includes the integration host factor (IHF), which both co-activates the master P(u) promoter and represses transcription of its cognate regulatory gene xylR at the onset of the stationary phase. On the other hand, the catabolite repression control (Crc) protein inhibits translation of XylR during exponential growth. By computing these two conflicting regulatory actions within a composite I4-FFL gate, cells shield the expression of XylR from perturbations caused by the growth phase, thereby ensuring a steady supply of the regulator regardless of physiological conditions. This device thus endows xylR expression with a degree of robustness in respect to the growth phase that could hardly be achieved with e.g. a simple constitutive promoter.