Transcription of pyoverdine biosynthetic (pvd) genes in Pseudomonas aeruginosa depends on the ECF sigma factor PvdS. The transmembrane sensor FpvR controls PvdS activity, acting as an antisigma of PvdS in the absence of a signal transmitted from the (ferri)pyoverdine-loaded FpvA receptor. We studied i) the competition between PvdS and RpoD, ii) the effect of FpvR on PvdS stability, and iii) the effect of AlgQ on PvdS-dependent transcription. PvdS attains ca. 60% of RpoD levels in exponentially growing cells, likely reflecting the high PvdS-RNA polymerase (RNAP) demand for recognition of multiple promoters. This explains only in part the changes in transcription pattern, since the major determinant is the ratio between different RNAP holoenzymes. RNAP purification from iron starved cells shows that the PvdS-RNAP complex is poorly represented compared with vegetative holoenzyme; copurification of PvdS-RNAP could only be achieved upon PvdS overexpression. Hence, high levels of PvdS possibly compensate for its weak affinity for RNAP. No significant differences in PvdS levels were observed between wild-type and pyoverdine signalling mutants, suggesting that in the absence of signaling part of the PvdS pool is sequestered by FpvR, but not degraded. Next, we investigated the effect of AlgQ, the Escherichia coli Rsd othologue, on pyoverdine regulation. AlgQ is a global regulatory protein which activates alginate, ppGpp, and polyP synthesis through a cascade involving nucleoside diphosphate kinase (Ndk). AlgQ was also shown to interact with region 4 of RpoD. We demonstrate that AlgQ modulates the transcription of pvd genes by an Ndk- and PvdS-independent mechanisms. Increased intracellular concentration of RpoD mimicked the DalgQ phenotype, whereas PvdS overexpression suppressed the algQ mutation. E. coli rsd could partly substitute for algQ in transcriptional modulation of pvd genes. Thus, AlgQ acts as an antisigma for RpoD, eliciting RNAP recruitment by PvdS and transcription initiation at pvd promoters. Collectively, these data indicate that PvdS activity is finely controlled by two antisigma factors capable of directly antagonizing or indirectly promoting PvdS binding to RNAP
Ambrosi, C., Tiburzi, F., Imperi, F., Putignani, L., Visca, P. (2005). Pyoverdine regulation: a matter of competition between sigma and antisigma factors. In Pseudomonas 2005 - 10th International Congress on Pseudomonas (pp.27). Merseille : Pseudomonas 2005.
Pyoverdine regulation: a matter of competition between sigma and antisigma factors
TIBURZI, FEDERICA;IMPERI, FRANCESCO;VISCA, PAOLO
2005-01-01
Abstract
Transcription of pyoverdine biosynthetic (pvd) genes in Pseudomonas aeruginosa depends on the ECF sigma factor PvdS. The transmembrane sensor FpvR controls PvdS activity, acting as an antisigma of PvdS in the absence of a signal transmitted from the (ferri)pyoverdine-loaded FpvA receptor. We studied i) the competition between PvdS and RpoD, ii) the effect of FpvR on PvdS stability, and iii) the effect of AlgQ on PvdS-dependent transcription. PvdS attains ca. 60% of RpoD levels in exponentially growing cells, likely reflecting the high PvdS-RNA polymerase (RNAP) demand for recognition of multiple promoters. This explains only in part the changes in transcription pattern, since the major determinant is the ratio between different RNAP holoenzymes. RNAP purification from iron starved cells shows that the PvdS-RNAP complex is poorly represented compared with vegetative holoenzyme; copurification of PvdS-RNAP could only be achieved upon PvdS overexpression. Hence, high levels of PvdS possibly compensate for its weak affinity for RNAP. No significant differences in PvdS levels were observed between wild-type and pyoverdine signalling mutants, suggesting that in the absence of signaling part of the PvdS pool is sequestered by FpvR, but not degraded. Next, we investigated the effect of AlgQ, the Escherichia coli Rsd othologue, on pyoverdine regulation. AlgQ is a global regulatory protein which activates alginate, ppGpp, and polyP synthesis through a cascade involving nucleoside diphosphate kinase (Ndk). AlgQ was also shown to interact with region 4 of RpoD. We demonstrate that AlgQ modulates the transcription of pvd genes by an Ndk- and PvdS-independent mechanisms. Increased intracellular concentration of RpoD mimicked the DalgQ phenotype, whereas PvdS overexpression suppressed the algQ mutation. E. coli rsd could partly substitute for algQ in transcriptional modulation of pvd genes. Thus, AlgQ acts as an antisigma for RpoD, eliciting RNAP recruitment by PvdS and transcription initiation at pvd promoters. Collectively, these data indicate that PvdS activity is finely controlled by two antisigma factors capable of directly antagonizing or indirectly promoting PvdS binding to RNAPI documenti in IRIS sono protetti da copyright e tutti i diritti sono riservati, salvo diversa indicazione.
