Marine Archaeon Methanosarcina acetivorans Enhances Polyphosphate Metabolism Under Persistent Cadmium Stress.

Phosphate metabolism was studied to determine whether polyphosphate (polyP) pools play a role in the enhanced resistance against Cd2+ and metal-removal capacity of Cd2+-preadapted (CdPA) Methanosarcina acetivorans. Polyphosphate kinase (PPK), exopolyphosphatase (PPX) and phosphate transporter transcript levels and their activities increased in CdPA cells compared to control (Cnt) cells. K+ inhibited recombinant Ma-PPK and activated Ma-PPX, whereas divalent cations activated both enzymes. Metal-binding polyP and thiol-containing molecule contents, Cd2+-removal, and biofilm synthesis were significantly higher in CdPA cells >Cnt cells plus a single addition of Cd2+>Cnt cells. Also, CdPA cells showed a higher number of cadmium, sulfur, and phosphorus enriched-acidocalcisomes than control cells. Biochemical and physiological phenotype exhibited by CdPA cells returned to that of Cnt cells when cultured without Cd2+. Furthermore, no differences in the sequenced genomes upstream and downstream of the genes involved in Cd2+ resistance were found between CdPA and Cnt cells, suggesting phenotype loss rather than genome mutations induced by chronic Cd2+-exposure. Instead, a metabolic adaptation induced by Cd2+ stress was apparent. The dynamic ability of M. acetivorans to change its metabolism, depending on the environmental conditions, may be advantageous to remove cadmium in nature and biodigesters.

Exopolyphosphatase of Pseudomonas aeruginosa is essential for the production of virulence factors, and its expression is controlled by NtrC and PhoB acting at two interspaced promoters.

The exopolyphosphatase (Ppx) of Pseudomonas aeruginosa is encoded by the PA5241 gene (ppx). Ppx catalyses the hydrolysis of inorganic polyphosphates to orthophosphate (Pi). In the present work, we identified and characterized the promoter region of ppx and its regulation under environmental stress conditions. The role of Ppx in the production of several virulence factors was demonstrated through studies performed on a ppx null mutant. We found that ppx is under the control of two interspaced promoters, dually regulated by nitrogen and phosphate limitation. Under nitrogen-limiting conditions, its expression was controlled from a σ(54)-dependent promoter activated by the response regulator NtrC. However, under Pi limitation, the expression was controlled from a σ(70) promoter, activated by PhoB. Results obtained from the ppx null mutant demonstrated that Ppx is involved in the production of virulence factors associated with both acute infection (e.g. motility-promoting factors, blue/green pigment production, C6-C12 quorum-sensing homoserine lactones) and chronic infection (e.g. rhamnolipids, biofilm formation). Molecular and physiological approaches used in this study indicated that P. aeruginosa maintains consistently proper levels of Ppx regardless of environmental conditions. The precise control of ppx expression appeared to be essential for the survival of P. aeruginosa and the occurrence of either acute or chronic infection in the host.

A role for the conserved GAFTGA motif of AAA+ transcription activators in sensing promoter DNA conformation.

Transcription from sigma54-dependent bacterial promoters can be regarded as a second paradigm for bacterial gene transcription. The initial sigma54-RNA polymerase (RNAP).promoter complex, the closed complex, is transcriptionally silent. The transcriptionally proficient sigma54-RNAP.promoter complex, the open complex, is formed upon remodeling of the closed complex by actions of a specialized activator protein that belongs to the AAA (ATPases associated with various cellular activities) protein family in an ATP hydrolysis-dependent reaction. The integrity of a highly conserved signature motif in the AAA activator (known as the GAFTGA motif) is important for the remodeling activity of the AAA activator and for open complex formation. We now provide evidence that the invariant threo-nine residue of the GAFTGA motif plays a role in sensing the DNA downstream of the sigma54-RNAP-binding site and in coupling this information to sigma54-RNAP via the conserved regulatory Region I domain of sigma54 during open complex formation.

The genome project of Taenia solium.

We have constituted a consortium of key laboratories at the National Autonomous University of Mexico to carry out a genomic project for Taenia solium. This project will provide powerful resources for the study of taeniasis/cysticercosis, and, in conjunction with the Echinococcus granulosus and Echinococcus multilocularis genome project of expressed sequence tags (ESTs), will mark the advent of genomics for cestode parasites. Our project is planned in two consecutive stages. The first stage is being carried out to determine some basic parameters of the T. solium genome. Afterwards, we will evaluate the best strategy for the second stage, a full blown genome project. We have estimated the T. solium genome size by two different approaches: cytofluorometry on isolated cyton nuclei, as well as a probabilistic calculation based on approximately 2000 sequenced genomic clones, approximately 3000 ESTs, resulting in size estimates of 270 and 251 Mb, respectively. In terms of sequencing, our goal for the first stage is to characterize several thousand EST's (from adult worm and cysticerci cDNA libraries) and genomic clones. Results obtained so far from about 16,000 sequenced ESTs from the adult stage, show that only about 40% of the T. solium coding sequences have a previously sequenced homologue. Many of the best hits are found with mammalian genes, especially with humans. However, 1.5% of the hits lack homologues in humans, making these genes immediate candidates for investigation on pharmaco-therapy, diagnostics and vaccination. Most T. solium ESTs are related to gene regulation, and signal transduction. Other important functions are housekeeping, metabolism, cell division, cytoskeleton, proteases, vacuolar transport, hormone response, and extracellular matrix activities. Preliminary results also suggest that the genome of T. solium is not highly repetitive.