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FNR is a well-studied global regulator of anaerobiosis, which is widely conserved across bacteria. Despite the importance of FNR and anaerobiosis in microbial lifestyles, the factors that influence its function on a genome-wide scale are poorly understood. Here, we report a functional genomic analysis of FNR action. We find that FNR occupancy at many target sites is strongly influenced by nucleoid-associated proteins (NAPs) that restrict access to many FNR binding sites. At a genome-wide level, only a subset of predicted FNR binding sites were bound under anaerobic fermentative conditions and many appeared to be masked by the NAPs H-NS, IHF and Fis. Similar assays in cells lacking H-NS and its paralog StpA showed increased FNR occupancy at sites bound by H-NS in WT strains, indicating that large regions of the genome are not readily accessible for FNR binding. Genome accessibility may also explain our finding that genome-wide FNR occupancy did not correlate with the match to consensus at binding sites, suggesting that significant variation in ChIP signal was attributable to cross-linking or immunoprecipitation efficiency rather than differences in binding affinities for FNR sites. Correlation of FNR ChIP-seq peaks with transcriptomic data showed that less than half of the FNR-regulated operons could be attributed to direct FNR binding. Conversely, FNR bound some promoters without regulating expression presumably requiring changes in activity of condition-specific transcription factors. Such combinatorial regulation may allow Escherichia coli to respond rapidly to environmental changes and confer an ecological advantage in the anaerobic but nutrient-fluctuating environment of the mammalian gut. [ABSTRACT FROM AUTHOR]

The article presents information on a study in which a systematic and iterative process Reconciling In silico/in vivo mutaNt Growth (RING) was used to analyze discrepancies between in silico prediction and in vivo observations to a genome-scale metabolic model of the fission yeast Schizosaccharomyces pombe, SpoMBEL1693. It says that the RING method helped in improving the overall predictive capability of SpoMBEL1693.

Manipulation of NADH-dependent steps, and particularly disruption of the las-located lactate dehydrogenase (ldh) gene in Lactococcus lactis, is common to engineering strategies envisaging the accumulation of reduced end products other than lactate. Reverse transcription-PCR experiments revealed that three out of the four genes assigned to lactate dehydrogenase in the genome of L. lactis, i.e., the ldh, ldhB, and ldhX genes, were expressed in the parental strain MG1363. Given that genetic redundancy is often a major cause of metabolic instability in engineered strains, we set out to develop a genetically stable lactococcal host tuned for the production of reduced compounds. Therefore, the ldhB and ldhX genes were sequentially deleted in L. lactis FI10089, a strain with a deletion of the ldh gene. The single, double, and triple mutants, FI10089, FI10089ΔldhB, and FI10089ΔldhBΔldhX, showed similar growth profiles and displayed mixed-acid fermentation, ethanol being the main reduced end product. Hence, the alcohol dehydrogenase-encoding gene, the adhE gene, was inactivated in FI10089, but the resulting strain reverted to homolactic fermentation due to induction of the ldhB gene. The three lactate dehydrogenase-deficient mutants were selected as a background for the production of mannitol and 2,3-butanediol. Pathways for the biosynthesis of these compounds were overexpressed under the control of a nisin promoter, and the constructs were analyzed with respect to growth parameters and product yields under anaerobiosis. Glucose was efficiently channeled to mannitol (maximal yield, 42%) or to 2,3-butanediol (maximal yield, 67%). The theoretical yield for 2,3-butanediol was achieved. We show that FI10089ΔldhB is a valuable basis for engineering strategies aiming at the production of reduced compounds [ABSTRACT FROM AUTHOR]

Abstract: Genome-based Flux Balance Analysis (FBA) and steady-state isotopic-labeling-based Metabolic Flux Analysis (MFA) are complimentary approaches to predicting and measuring the operation and regulation of metabolic networks. Here, genome-derived models of Escherichia coli (E. coli) metabolism were used for FBA and 13C-MFA analyses of aerobic and anaerobic growths of wild-type E. coli (K-12 MG1655) cells. Validated MFA flux maps reveal that the fraction of maintenance ATP consumption in total ATP production is about 14% higher under anaerobic (51.1%) than aerobic conditions (37.2%). FBA revealed that an increased ATP utilization is consumed by ATP synthase to secrete protons from fermentation. The TCA cycle is shown to be incomplete in aerobically growing cells and submaximal growth is due to limited oxidative phosphorylation. An FBA was successful in predicting product secretion rates in aerobic culture if both glucose and oxygen uptake measurement were constrained, but the most-frequently predicted values of internal fluxes yielded from sampling the feasible space differ substantially from MFA-derived fluxes. [ABSTRACT FROM AUTHOR]

Horizontal gene transfer (HGT) is known to have contributed to the content of eukaryotic genomes, but the direct effects of HGT on eukaryotic evolution are more obscure because many of the best supported cases involve a new gene replacing a functionally similar homologue. Here, several cases of HGT conferring a plausible adaptive advantage are reviewed to examine emerging trends in such transfer events. In particular, HGT seems to play an important role in adaptation to parasitism and pathogenesis, as well as to other specific environmental conditions such as anaerobiosis or nitrogen and iron limitation in marine environments. Most, but not all, of the functionally significant HGT to eukaryotes comes from bacteria, in part due to chance, but probably also because bacteria have greater metabolic diversity to offer. [Copyright &y& Elsevier]

Background: The Escherichia coli heterodimeric HU protein is a small DNA-bending protein associated with the bacterial nucleoid. It can introduce negative supercoils into closed circular DNA in the presence of topoisomerase I. Cells lacking HU grow very poorly and display many phenotypes. Methodology/Principal Findings: We analyzed the transcription profile of every Escherichia coli gene in the absence of one or both HU subunits. This genome-wide in silico transcriptomic approach, performed in parallel with in vivo genetic experimentation, defined the HU regulon. This large regulon, which comprises 8% of the genome, is composed of four biologically relevant gene classes whose regulation responds to anaerobiosis, acid stress, high osmolarity, and SOS induction. Conclusions/Significance: The regulation a large number of genes encoding enzymes involved in energy metabolism and catabolism pathways by HU explains the highly pleiotropic phenotype of HU-deficient cells. The uniform chromosomal distribution of the many operons regulated by HU strongly suggests that the transcriptional and nucleoid architectural functions of HU constitute two aspects of a unique protein-DNA interaction mechanism. [ABSTRACT FROM AUTHOR]

Background: The capacity of respiring cultures of Saccharomyces cerevisiae to immediately switch to fast alcoholic fermentation upon a transfer to anaerobic sugar-excess conditions is a key characteristic of Saccharomyces cerevisiae in many of its industrial applications. This transition was studied by exposing aerobic glucose-limited chemostat cultures grown at a low specific growth rate to two simultaneous perturbations: oxygen depletion and relief of glucose limitation. Results: The shift towards fully fermentative conditions caused a massive transcriptional reprogramming, where one third of all genes within the genome were transcribed differentially. The changes in transcript levels were mostly driven by relief from glucose-limitation. After an initial strong response to the addition of glucose, the expression profile of most transcriptionally regulated genes displayed a clear switch at 30 minutes. In this respect, a striking difference was observed between the transcript profiles of genes encoding ribosomal proteins and those encoding ribosomal biogenesis components. Not all regulated genes responded with this binary profile. A group of 87 genes showed a delayed and steady increase in expression that specifically responded to anaerobiosis. Conclusion: Our study demonstrated that, despite the complexity of this multiple-input perturbation, the transcriptional responses could be categorized and biologically interpreted. By comparing this study with public datasets representing dynamic and steady conditions, 14 up-regulated and 11 down-regulated genes were determined to be anaerobic specific. Therefore, these can be seen as true "signature" transcripts for anaerobicity under dynamic as well as under steady state conditions. [ABSTRACT FROM AUTHOR]

The abundance of 4Fe–4S motifs of the form CX2CX2CX3C was analyzed in the open reading frames (ORFs) of 120 prokaryotic genomes. The abundance of ORFs containing the CX2CX2CX3C motif or isORFs correlated (r=0.82) with methanogenesis (p=0.0001), archaea (p=0.0173), anaerobiosis (p<0.0001) and genome size (p<0.0001). Optimal growth temperature (hyperthermophily) did not correlate with the number of isORFs (p=0.6283). Large numbers of CX2CX2CX3C motifs may be associated with unique physiologies: methanogenic archaea contained the greatest number of CX2CX2CX3C motifs found among the prokaryotic groups; however, only about 15% of the motifs were in genes directly involved in methanogenesis. Large numbers of CX2CX2CX3C motifs may also be associated with generalists such as Desulfitobacterium hafniense, which is an anaerobic bacterium containing multiple reductases. [Copyright &y& Elsevier]