Positive and Negative Regulation of Glycerol Utilization by the c-di-GMP Binding Protein PlzA in .

Borrelia burgdorferi, the causative agent of Lyme disease, encounters two disparate host environments during its enzootic life cycle, &odes ticks and mammalian hosts. B. burgdorferi has a small genome that encodes a streamlined cyclic dimeric GMP (c-di-GMP) signaling system comprising a single diguanylate cyclase, Rrpl, and two phosphodiesterases. This system is essential for spirochete survival in ticks, in part because it controls the expression of the glp operon involved in glycerol utilization. In this study, we showed that a B. burgdorferi c-di-GMP receptor, PIzA, functions as both a positive and a negative regulator for glp expression. Deletion of pizA or mutation in pizA that impaired c-di-GMP binding abolished glp expression. On the other hand, overexpression of pizA resulted in gip repression, which could be rescued by simultaneous overexpression of rip!. pizA overexpression in the rrpl mutant, which is devoid of c-di-GMP, or overexpression of a pizA mutant incapable of c-di-GMP binding further enhanced gip repression. Combined results suggest that c-di-GMP-bound PIzA functions as a positive regulator, whereas ligand-free PIzA acts as a negative regulator for gip expression. Thus, PIzA of B. burgdorferi with a streamlined c-di-GMP signaling system not only controls multiple targets, as previously envisioned, but has also evolved different modes of action. IMPORTANCE The Lyme disease pathogen, Borrelia burgdorferi, has a simple cyclic dimeric GMP (c-di-GMP) signaling system essential for adaptation of the pathogen to the complicated tick environment. The c-di-GMP effector of B. burgdorferi, PIzA, has been shown to regulate multiple cellular processes, including motility, osmolality sensing, and nutrient utilization. The findings of this study demonstrate that PIzA not only controls multiple targets but also has different functional modalities, allowing it to act as both positive and negative regulator of the glp operon expression. This work highlights how bacteria with a small genome can compensate for the limited regulatory repertoire by increasing the complexity of targets and modes of action in their regulatory proteins.