Project description:Small RNAs were isolated from the hyperthermophilic archaeon Thermoproteus tenax and subjected to Illumina Hiseq2000 sequencing. The RNA reads revealed significant abundance of constitutively transcribed CRISPR RNAs and a plethora of C/D box sRNAs. The presence of a rare RNase P RNA variant was verified. Circular RNA molecules could be identified.
Project description:Hydrogen served as a competitive inorganic energy source, impacting the CuFeS2 bioleaching efficiency of the extremely thermoacidophilic archaeon Metallosphaera sedula. Open reading frames encoding key terminal oxidase and electron transport chain components were triggered by CuFeS2. Evidence of heterotrophic metabolism was noted after extended periods of bioleaching, presumably related to cell lysis.
Project description:(from abstract): Iron oxidation is a desirable trait of biomining microorganisms, although the mechanism is not well-understood in extreme thermoacidophiles. The complete genome sequence of the extremely thermoacidophilic archaeon Metallosphaera sedula DSM 5348 (2.2 Mb, ~2300 ORFs) provides insights into biologically catalyzed metal sulfide oxidation. Comparative genomics was used to identify pathways and proteins (in)directly involved with bioleaching. As expected, the M. sedula genome encodes genes related to autotrophic carbon fixation, metal tolerance, and adhesion. Also, terminal oxidase cluster organization indicates the presence of hybrid quinol-cytochrome oxidase complexes. Comparisons with the mesophilic biomining bacterium Acidithiobacillus ferrooxidans ATCC 23270 indicate that the M. sedula genome encodes at least one putative rusticyanin, involved in iron oxidation. The fox gene cluster, involved in iron oxidation in the thermoacidophilic archaeon Sulfolobus metallicus, could also be identified. These iron-oxidizing components are missing from genomes of non-leaching Sulfolobales like Sulfolobus solfataricus P2 and Sulfolobus acidocaldarius DSM 639. Whole genome transcriptional response analysis showed that 88 ORFs were up-regulated 2-fold or more in M. sedula upon addition of ferrous sulfate to yeast extract-based medium; these included components of terminal oxidase clusters predicted to be involved with iron oxidation, as well as genes predicted to be involved with sulfur metabolism. Many hypothetical proteins were also differentially transcribed, indicating that aspects of the iron and sulfur metabolism of M. sedula remain to be identified and characterized. Keywords: substrate response
Project description:(from abstract): Iron oxidation is a desirable trait of biomining microorganisms, although the mechanism is not well-understood in extreme thermoacidophiles. The complete genome sequence of the extremely thermoacidophilic archaeon Metallosphaera sedula DSM 5348 (2.2 Mb, ~2300 ORFs) provides insights into biologically catalyzed metal sulfide oxidation. Comparative genomics was used to identify pathways and proteins (in)directly involved with bioleaching. As expected, the M. sedula genome encodes genes related to autotrophic carbon fixation, metal tolerance, and adhesion. Also, terminal oxidase cluster organization indicates the presence of hybrid quinol-cytochrome oxidase complexes. Comparisons with the mesophilic biomining bacterium Acidithiobacillus ferrooxidans ATCC 23270 indicate that the M. sedula genome encodes at least one putative rusticyanin, involved in iron oxidation. The fox gene cluster, involved in iron oxidation in the thermoacidophilic archaeon Sulfolobus metallicus, could also be identified. These iron-oxidizing components are missing from genomes of non-leaching Sulfolobales like Sulfolobus solfataricus P2 and Sulfolobus acidocaldarius DSM 639. Whole genome transcriptional response analysis showed that 88 ORFs were up-regulated 2-fold or more in M. sedula upon addition of ferrous sulfate to yeast extract-based medium; these included components of terminal oxidase clusters predicted to be involved with iron oxidation, as well as genes predicted to be involved with sulfur metabolism. Many hypothetical proteins were also differentially transcribed, indicating that aspects of the iron and sulfur metabolism of M. sedula remain to be identified and characterized. Dye flip of Mse cells includes two samples, yeast exact (YE) and yeast extract + ferrous sulfate (YEF). The first slide, Y3F5, has YE RNA labeled with cy3 and YEF RNA labeled with cy5, while the second slide, F3Y5, has YEF RNA labeled with cy3 and YE RNA labeled with cy5. YE serves as the reference condition, with the expectation that ORFs involved with Fe2+ oxidation (and SO4 metabolism) will be upregulated on YEF (log2 fold change of YE-YEF < -1).
Project description:Metallosphaera sedula is an extremely thermoacidophilic archaeon that grows heterotrophically on peptides, and chemolithoautotrophically on hydrogen, sulfur, or reduced metals as energy sources. During autotrophic growth, carbon dioxide is incorporated into cellular carbon via the 3-hydroxypropionate /4-hydroxybutyrate cycle (3HP/4HB). To date, all of the steps in the pathway have been connected to enzymes encoded in specific ORFs, except for the one responsible for ligation of coenzyme A (CoA) to 4-hydroxybutyrate (4HB). While several candidates for this step have been identified through bioinformatic analysis of the M. sedula genome, none have been shown to catalyze this biotransformation. Transcriptomic analysis of cells grown under strict H2-CO2 autotrophy was used elucidate additional candidate genes involved in carbon fixation and identify the genes which encode for 4HB-CoA synthetase.
Project description:Hydrogen served as a competitive inorganic energy source, impacting the CuFeS2 bioleaching efficiency of the extremely thermoacidophilic archaeon Metallosphaera sedula. Open reading frames encoding key terminal oxidase and electron transport chain components were triggered by CuFeS2. Evidence of heterotrophic metabolism was noted after extended periods of bioleaching, presumably related to cell lysis. One 3 slide loop for Mse cells includes 3 conditions: M. sedula inoculum prior to introduction to chalcopyrite (d0~day 0), M. sedula after 3 days on chalcopyrite (d3~day 3), and M. sedula after 9 days on chalcopyrite (d9~day 9). Half of an RNA sample for one condition (consisting of pools of RNA from multiple cultures) was labeled with Cy3 while the other half was labeled with Cy5. The two differently labeled samples were run on different slides. Each probe is spotted on each slide 5 times (5 spots/slide x 2 slides = 10 technical replicates per condition; spot intensities for all replicates on slide provided in associated raw data file).