Article

Microarray and bioinformatic analyses suggest models for carbon metabolism in the autotroph Acidithiobacillus ferrooxidans

Laboratoire de Chimie Bactérienne, IBSM, CNRS, Marseille, France; Andrés Bello University and Millennium Institute for Fundamental and Applied Biology, Santiago, Chile; University of Illinois, Chicago, USA; Idaho National Laboratory, Idaho Falls, USA; University of Santiago, Santiago, Chile; ICBM, Faculty of Medicine, University of Chile, Santiago, Chile
Hydrometallurgy (Impact Factor: 2.17). 09/2006; DOI: 10.1016/j.hydromet.2006.03.029

ABSTRACT Acidithiobacillus ferrooxidans is a chemolithoautotrophic bacterium that uses iron or sulfur as an energy and electron source. Bioinformatic analysis of the A. ferrooxidans draft genome sequence was used to identify putative genes and potential metabolic pathways involved in CO2 fixation, 2P-glycolate detoxification, carboxysome formation and glycogen utilization. Microarray transcript profiling was carried out to compare the relative expression of the predicted genes of these pathways when the microorganism was grown in the presence of iron versus sulfur. Several gene expression patterns were confirmed by real-time PCR. Genes for each of the above-predicted pathways were found to be organized into discrete clusters. Clusters exhibited differential gene expression depending on the presence of iron or sulfur in the medium. Concordance of gene expression within each cluster suggested that they are operons. Most notably, clusters of genes predicted to be involved in CO2 fixation, carboxysome formation, 2P-glycolate detoxification and glycogen biosynthesis were upregulated in sulfur medium, whereas genes involved in glycogen utilization were preferentially expressed in iron medium. These results can be explained in terms of models of gene regulation that suggest how A. ferrooxidans can adjust its central carbon management to respond to changes in its environment.

1 Bookmark
 · 
83 Views
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: The extremely acidophilic, chemolithoautotrophic Acidithiobacillus ferrooxidans is an important bioleaching bacterium of great value in the metallurgical industry and environmental protection. In this report, a mutagenesis system based on the homing endonuclease I-SceI was developed to produce targeted, unmarked gene deletions in the strain A. ferrooxidans ATCC 23270. A targeted phosphofructokinase (PFK) gene (pfkB) mutant of A. ferrooxidans ATCC 23270 was constructed by homologous recombination and identified by PCR with specific primers as well as Southern blot analysis. This potential pfkB gene (AFE_1807) was also characterized by expression in PFK-deficient Escherichia coli cells, and heteroexpression of the PFKB protein demonstrated that it had functional PFK activity, though it was significantly lower (about 800-fold) than that of phosphofructokinase-2 (PFK-B) expressed by the pfkB gene from E. coli K-12. The function of the potential PFKB protein in A. ferrooxidans was demonstrated by comparing the properties of the pfkB mutant with those of the wild type. The pfkB mutant strain displayed a relatively reduced growth capacity in S(0) medium (0.5% [wt/vol] elemental sulfur in 9K basal salts solution adjusted to pH 3.0 with H(2)SO(4)), but the mutation did not completely prevent A. ferrooxidans from assimilating exogenous glucose. The transcriptional analysis of some related genes in central carbohydrate metabolism in the wild-type and mutant strains with or without supplementation of glucose was carried out by quantitative reverse transcription-PCR. This report suggests that the markerless mutagenesis strategy could serve as a model for functional studies of other genes of interest from A. ferrooxidans and multiple mutations could be made in a single A. ferrooxidans strain.
    Applied and Environmental Microbiology 12/2011; 78(6):1826-35. · 3.95 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: To define the molecular response of Acidithiobacillus ferrooxidans under pH up-shift, temporal gene expression profiles were examined by using whole-genome DNA microarrays for A. ferrooxidans. Approximately 30% of the 3,132 genes represented on the microarray were significantly upregulated over a 160-min period, while about 14% were significantly downregulated. Our results revealed that A. ferrooxidans showed potential self-protection and self-regulation performance in response to pH up-shift stress. Many genes involved in regulation of membrane components were differentially expressed under the pH up-shift stress. Likewise, most of genes involved in phosphate metabolism, sulfur assimilation, and CO(2) fixation were obviously induced. Conversely, the transcription of a polyphosphate kinase gene (AFE1210) associated with phosphate storage was significantly repressed, which probably stemmed from the depletion of polyphosphate. Besides, most of the genes involved in hydrogen uptake were significantly induced, whereas many genes involved in nitrogen fixation were obviously repressed, which suggested that hydrogen uptake and nitrogen fixation could contribute to cytoplasmic pH homeostasis.
    Folia Microbiologica 09/2011; 56(5):439-51. · 0.79 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: In contrast to iron-oxidizing Acidithiobacillus ferrooxidans, A. ferrooxidans from a stationary phase elemental sulfur-oxidizing culture exhibited a lag phase in pyrite oxidation, which is similar to its behaviour during ferrous iron oxidation. The ability of elemental sulfur-oxidizing A. ferrooxidans to immediately oxidize ferrous iron or pyrite without a lag phase was only observed in bacteria obtained from growing cultures with elemental sulfur. However, these cultures that shifted to ferrous iron oxidation showed a low rate of ferrous iron oxidation while no growth was observed. Two-dimensional gel electrophoresis was used for a quantitative proteomic analysis of the adaptation process when bacteria were switched from elemental sulfur to ferrous iron. A comparison of total cell lysates revealed 39 proteins whose increase or decrease in abundance was related to this phenotypic switching. However, only a few proteins were closely related to iron and sulfur metabolism. Reverse-transcription quantitative PCR was used to further characterize the bacterial adaptation process. The expression profiles of selected genes primarily involved in the ferrous iron oxidation indicated that phenotypic switching is a complex process that includes the activation of genes encoding a membrane protein, maturation proteins, electron transport proteins and their regulators.
    Antonie van Leeuwenhoek 01/2013; · 2.07 Impact Factor

Full-text (4 Sources)

View
51 Downloads
Available from
Jun 2, 2014