Project description:Investigation of whole genome gene expression level changes in anaerobic, nitrate-dependent Fe(II) oxidation in the chemolithoautotrophic bacterium Thiobacillus denitrificans Here we report on a study to identify genes associated with nitrate-dependent Fe(II) oxidation by whole-genome transcriptional (microarray) assays including the use of FeCO3, Fe2+, and U(IV) oxides as electron donors under denitrifying conditions. A 25 chip study using total RNA recovered from wild-type T. denitrificans was cultivated at 30oC under strictly anaerobic conditions with growth medium that contained 20 mM thiosulfate, 20 mM nitrate, and 30 mM bicarbonate (pH ~7) and exposed to 8 treatments. Each chip measures the expression level of 2832 ORFs with N 24-mer probe pairs (PM/MM) per gene, with three-fold technical redundancy.
Project description:Beller, H. R., T. E. Letain, A. Chakicherla, S. R. Kane, T. C. Legler, and M. A. Coleman. 2006. Whole-genome transcriptional analysis of chemolithoautotrophic thiosulfate oxidation by Thiobacillus denitrificans under aerobic vs. denitrifying conditions. Journal of Bacteriology 188:7005-7015. Thiobacillus denitrificans is one of the few known obligate chemolithoautotrophic bacteria capable of energetically coupling thiosulfate oxidation to denitrification as well as aerobic respiration. As very little is known about the differential expression of genes associated with key chemolithoautotrophic functions (such as sulfur-compound oxidation and CO2 fixation) under aerobic versus denitrifying conditions, we conducted whole-genome, cDNA microarray studies to explore this topic systematically. The microarrays identified 277 genes (approximately ten percent of the genome) as differentially expressed using Robust Multi-array Average statistical analysis and a 2-fold cutoff. Genes upregulated (ca. 6- to 150-fold) under aerobic conditions included a cluster of genes associated with iron acquisition (e.g., siderophore-related genes), a cluster of cytochrome cbb3 oxidase genes, cbbL and cbbS (encoding the large and small subunits of form I ribulose 1,5-bisphosphate carboxylase/oxygenase, or RubisCO), and multiple molecular chaperone genes. Genes upregulated (ca. 4- to 95-fold) under denitrifying conditions included nar, nir, and nor genes (associated respectively with nitrate reductase, nitrite reductase, and nitric oxide reductase, which catalyze successive steps of denitrification), cbbM (encoding form II RubisCO), and genes involved with sulfur-compound oxidation (including two physically separated but highly similar copies of sulfide:quinone oxidoreductase and of dsrC, associated with dissimilatory sulfite reductase). Among genes associated with denitrification, relative expression levels (i.e., degree of upregulation with nitrate) tended to decrease in the order nar > nir > nor > nos. Reverse transcription, quantitative PCR analysis was used to validate these trends. Keywords: bacterial metabolism
Project description:Quantitative proteomic analysis of Paracoccus denitrificans PD1222 wild type and NtrY defective mutant in denitrifying conditions (anaerobiosis, nitrate as nitrogen source)
