Probing flagellar promoter occupancy in wild-type and mutant Caulobacter crescentus by chromatin immunoprecipitation

In the asymmetric predivisional cell of Caulobacter crescentus, TipF and TipN mark the cellular pole for future flagellar development. TipF is essential for motility and contains a cyclic-di-GMP phosphodiesterase-like (EAL) domain that is necessary for proper function. TipN is localized to the flagellar pole before TipF and is essential for the proper placement of the flagellum in C. crescentus. Using β-galactosidase promoter-probe assays and quantitative chromatin immunoprecipitation, we investigated the influence of the C. crescentus flagellar assembly regulator TipF on flagellar gene transcription. We compared the transcriptional activity of class II-fliF-lacZ, class III-flgE-lacZ, and class IV-fljL-lacZ fusions in a ΔtipF mutant with that of other flagellar mutants and the wild-type strain. We subsequently verified the in vivo occupancy of the fliF, flgE, and fljL flagellar promoters by the flagellar regulators CtrA, FlbD, and FliX in addition to RNA polymerase. We deduce that TipF contributes to proper expression of flagellar genes in C. crescentus by acting both within and outside of the canonical flagellar gene expression hierarch

Decoding Caulobacter development

Caulobacter crescentus uses a multi-layered system of oscillating regulators to program different developmental fates into each daughter cell at division. This is achieved by superimposing gene expression, subcellular localization, phosphorylation, and regulated proteolysis to form a complex regulatory network that integrates chromosome replication, segregation, polar differentiation, and cytokinesis. In this review, we outline the current state of research in the field of Caulobacter development, emphasizing new findings that elaborate how the developmental program is modulated by factors such as the environment or the metabolic state of the cell. New findings detailing how the developmental program is modulated by factors such as the environment or the metabolic state of the cell are discusse

Versatility of global transcriptional regulators in alpha-Proteobacteria: from essential cell cycle control to ancillary functions

Recent data indicate that cell cycle transcription in many alpha-Proteobacteria is executed by at least three conserved functional modules in which pairs of antagonistic regulators act jointly, rather than in isolation, to control transcription in S-, G2- or G1-phase. Inactivation of module components often results in pleiotropic defects, ranging from cell death and impaired cell division to fairly benign deficiencies in motility. Expression of module components can follow systemic (cell cycle) or external (nutritional/cell density) cues and may be implemented by auto-regulation, ancillary regulators or other (unknown) mechanisms. Here, we highlight the recent progress in understanding the molecular events and the genetic relationships of the module components in environmental, pathogenic and/or symbiotic alpha-proteobacterial genera. Additionally, we take advantage of the recent genome-wide transcriptional analyses performed in the model alpha-Proteobacterium Caulobacter crescentus to illustrate the complexity of the interactions of the global regulators at selected cell cycle-regulated promoters and we detail the consequences of (mis-)expression when the regulators are absent. This review thus provides the first detailed mechanistic framework for understanding orthologous operational principles acting on cell cycle-regulated promoters in other alpha-Proteobacteri

The role of peptidoglycan in chlamydial cell division: towards resolving the chlamydial anomaly

Chlamydiales are obligate intracellular bacteria including some important pathogens causing trachoma, genital tract infections and pneumonia, among others. They share an atypical division mechanism, which is independent of an FtsZ homologue. However, they divide by binary fission, in a process inhibited by penicillin derivatives, causing the formation of an aberrant form of the bacteria, which is able to survive in the presence of the antibiotic. The paradox of penicillin sensitivity of chlamydial cells in the absence of detectable peptidoglycan (PG) was dubbed the chlamydial anomaly, since no PG modified by enzymes (Pbps) that are the usual target of penicillin could be detected in Chlamydiales. We review here the recent advances in this field with the first direct and indirect evidences of PG-like material in both Chlamydiaceae and Chlamydia-related bacteria. Moreover, PG biosynthesis is required for proper localization of the newly described septal proteins RodZ and NlpD. Taken together, these new results set the stage for a better understanding of the role of PG and septal proteins in the division mechanism of Chlamydiales and illuminate the long-standing chlamydial anomaly. Moreover, understanding the chlamydial division mechanism is critical for the development of new antibiotics for the treatment of chlamydial chronic infection

Dynamic of dissolved organic matter and trace elements in a steady state lake bottom layer : molecular size fractionation.

International audienceDissolved organic matter (DOM) in aquatic systems interacts with most processes including redox reactions, trace elements complexation, sorption and sedimentation. The complex nature of DOM in natural waters suggests that all the fractions constituting it do not contribute in the same way to its reactivity. The total stock of DOM can be split by various means like the difference in size of the molecules or the difference in affinity for adsorbent phases. The deep and stable layers of meromictic lakes are of a particular interest for the study of the mechanisms controlling the behaviour of chemical elements in natural systems. Quasi stationary conditions unroll and stratify the processes through a certain thickness of the water column. In the deep layer of the lake Pavin (Massif-Central, France), solutes dispersing from the water-sediment interface (92 meter depth) cross a succession of levels characterized by different physicochemical and microbiological conditions (redox conditions, interactions with neoformed or settling particles, bacterial metabolic types) before to reach the oxic layer around 60 meter depth (Viollier et al., 1995, 1997, Lehours et al., 2003)

Interactions between trace elements and dissolved organic matter in the stagnant anoxic deep layer of a meromictic lake

About 80% and more than 90% of the dissolved organic carbon (DOC) in the bottom water of Lake Pavin were isolated, respectively, on Amberlite XAD and Spherodex DEAE resins. Trace element concentrations in the fractions isolated were analyzed by using ICP-MS. Uranium, molybdenum, and antimony were found to be about 40% associated with fulvic and hydrophilic acids adsorbed on XAD resins at pH 2, the metal-organic association being not dissociated. Much higher percentages of the same elements, plus vanadium, were adsorbed on the DEAE resin at the pH of natural water, possibly because of the better preservation of acid labile organic complexes or of the supplementary adsorption of inorganic compounds. Ultrafiltration and dialysis made it possible to exclude colloidal fractions together with much U (78%), Mo (80%), V (55%), and DOC (65%). These elements and many others (including barium) were previously found to be quite reactive at the bottom of the lake, (apparently being scavenged by settling particles, which are mostly diatoms, then dissolved at the sediment-water interface). Conversely, trace elements with a conservative behavior in the bottom layer (such as lithium and cesium) were not found associated with dissolved organic mater (DOM). Barium was not strongly associated with the extractable DOM, which may argue for a direct interaction with inorganic particles or the existence of very labile complexes. Our work suggests the existence of relatively stable (nonacid labile) U and Mo-DOM colloidal associations in the anoxic bottom waters of the lake and their importance in the scavenging of those metals

Biogeochemical modelling of anaerobic vs. aerobic methane oxidation in a meromictic crater lake (Lake Pavin, France)

International audienceMethane is a powerful greenhouse gas and its concentration in the atmosphere has increased over the past decades. Methane produced by methanogenic Archae can be consumed through aerobic and anaerobic oxidation pathways. In anoxic conditions found in freshwater environments such as meromictic lakes, CH4 oxidation pathways involving different terminal electron acceptors such as NO 3 , SO2 4 , and oxides of Fe and Mn are thermodynamically possible. In this study, a reactive transport model was developed to assess the relative significance of the different pathways of CH4 consumption in the water column of Lake Pavin. In most cases, the model reproduced experimental data collected from the field from June 2006 to June 2007. Although the model and the field measurements suggest that anaerobic CH4 oxidation may contribute to CH4 consumption in the water column of Lake Pavin, aerobic oxidation remains the major sink of CH4 in this lake

Genetic and Computational Identification of a Conserved Bacterial Metabolic Module

We have experimentally and computationally defined a set of genes that form a conserved metabolic module in the α-proteobacterium Caulobacter crescentus and used this module to illustrate a schema for the propagation of pathway-level annotation across bacterial genera. Applying comprehensive forward and reverse genetic methods and genome-wide transcriptional analysis, we (1) confirmed the presence of genes involved in catabolism of the abundant environmental sugar myo-inositol, (2) defined an operon encoding an ABC-family myo-inositol transmembrane transporter, and (3) identified a novel myo-inositol regulator protein and cis-acting regulatory motif that control expression of genes in this metabolic module. Despite being encoded from non-contiguous loci on the C. crescentus chromosome, these myo-inositol catabolic enzymes and transporter proteins form a tightly linked functional group in a computationally inferred network of protein associations. Primary sequence comparison was not sufficient to confidently extend annotation of all components of this novel metabolic module to related bacterial genera. Consequently, we implemented the Graemlin multiple-network alignment algorithm to generate cross-species predictions of genes involved in myo-inositol transport and catabolism in other α-proteobacteria. Although the chromosomal organization of genes in this functional module varied between species, the upstream regions of genes in this aligned network were enriched for the same palindromic cis-regulatory motif identified experimentally in C. crescentus. Transposon disruption of the operon encoding the computationally predicted ABC myo-inositol transporter of Sinorhizobium meliloti abolished growth on myo-inositol as the sole carbon source, confirming our cross-genera functional prediction. Thus, we have defined regulatory, transport, and catabolic genes and a cis-acting regulatory sequence that form a conserved module required for myo-inositol metabolism in select α-proteobacteria. Moreover, this study describes a forward validation of gene-network alignment, and illustrates a strategy for reliably transferring pathway-level annotation across bacterial species.</p

Biogeochemical Dynamics of Molybdenum in a Crater Lake: Seasonal Impact and Long-Term Removal

International audienceDespite a large variety of processes that can control Mo and its potential to become an environmental tracer of euxinic environment, this element is not often studied in lakes. The aim of this paper is to identify main seasonal biogeochemical processes that involve Mo in a well constrained freshwater system (Lake Pavin water-column) in order to evaluate their respective importance. In Lake Pavin, 4 main processes have been identified: 1) the transitional process represented by Mo assimilation of by phytoplankton in the epilimnion (nitrogen biological fixation and nitrate assimilation); 2) transient process represented by dissolved Mo adsorption onto Fe and Mn metal oxides at oxic/anoxic interface (depth 50 - 60 m); 3) Mo precipitation where apparent sulfide production rate is maximum, and from 80 m depths; 4) release of dissolved Mo due to Mo benthic flux or input from a deep source