A variety of soil-dwelling bacteria produce polyhydroxybutyrate (PHB), which serves as a source of energy and carbon under nutrient deprivation. Bacteria belonging to the genus Pseudomonas do not generally produce PHB but are capable of using the PHB degradation product (R)-3-hydroxybutyrate [(R)-3-HB] as a growth substrate. Essential to this utilization is the NAD+-dependent dehydrogenase BdhA that converts (R)-3-HB into acetoacetate, a molecule that readily enters central metabolism. Apart from the numerous studies that had focused on the biochemical characterization of BdhA, there was nothing known about the assimilation of (R)-3-HB in Pseudomonas, including the genetic regulation of bdhA expression. This study aimed to define the regulatory factors that govern or dictate the expression of the bdhA gene and (R)-3-HB assimilation in Pseudomonas aeruginosa PAO1. Importantly, expression of the bdhA gene was found to be specifically induced by (R)-3-HB in a manner dependent on the alternative sigma factor RpoN and the enhancer-binding protein PA2005.This mode of regulation was essential for the utilization of (R)-3-HB as a sole source of energy and carbon. However, non-induced levels of bdhA expression were sufficient for P. aeruginosa PAO1 to grow on ( ± )-1,3-butanediol, which is catabolized through an (R)-3-HB intermediate. Because this is, we believe, the first report of an enhancer-binding protein that responds to (R)-3-HB, PA2005 was named HbcR for (R)-3-hydroxybutyrate catabolism regulator.
Borrelia burgdorferi is the causative agent of Lyme disease. In B. burgdorferi, RpoS controls the expression of virulence genes needed for mammalian infection. The Fur homologue BosR regulates the transcription of rpoS and therefore BosR determines, albeit indirectly, the infection status of the spirochaete. Transcription of rpoS in B. burgdorferi is complex: rpoS can be transcribed either from an RpoD-dependent promoter to yield a long transcript or from an RpoN-dependent promoter to yield a short transcript. This study shows that BosR repressed synthesis of the long transcript while at the same time activating synthesis of the short transcript. How BosR does this is unclear. To address this, spirochaetes were engineered to express either BosR or the naturally occurring variant BosRR39K. Mice became infected by the spirochaetes expressing BosR but not by the spirochaetes expressing BosRR39K. Furthermore, the spirochaetes expressing BosR activated rpoS transcription during growth in culture whereas the spirochaetes expressing BosRR39K did not. Thus, BosR's activation of rpoS transcription somehow involves Arg39. This arginine is highly conserved in other FUR proteins and therefore other FUR proteins may also require this arginine to function.
Rhodospirillum centenum utilizes 3′,5′-cyclic guanosine monophosphate (cGMP) as a messenger to regulate development of desiccation-resistant cysts. In this study, we demonstrated that gcyA, gcyB and gcyC, coding for putative subunits of a guanylyl cyclase, increase expression from 8- to 500-fold when cells transition from vegetative to cyst phases of growth. This induction did not occur in a strain that is defective in cGMP synthesis or in a strain that contains a deletion of cgrA that codes for a cGMP-binding homologue of Escherichia coli catabolite repressor protein (CRP). We also demonstrated that cgrA auto-induces its own expression in the presence of cGMP, indicating that a feed-forward loop is used to ramp up cGMP production as cells undergo encystment. Inspection of an intragenic region upstream of gcyB revealed a sequence that is identical to the CRP consensus sequence from E. coli. DNase I and fluorescence anisotropy analyses demonstrated that CgrA bound to this target sequence at a protein : cGMP ratio of 1 : 2 with K d ∼61 nM. This was in contrast to CgrA in the presence of cAMP, which exhibited K d ∼1795 nM. CgrA thus constitutes a novel variant of CRP that utilizes cGMP to regulate production of cGMP synthase for the control of cyst development.