Project description:Aerobic methanotrophic bacteria use methane as their sole source of carbon and energy and serve as a major sink for the potent greenhouse gas methane in freshwater ecosystems. Despite this important environmental role, little is known about the molecular details of how these organisms interact in the environment. Many bacterial species use quorum sensing systems to regulate gene expression in a density-dependent manner. We have identified a quorum sensing system in the genome of Methylobacter tundripaludum, a dominant methane-oxidizer in methane enrichments of sediment from Lake Washington (Seattle, WA, USA). We determined that M. tundripaludum primarily produces N-3-hydroxydecanoyl-L-homoserine lactone (3-OH-C10-HSL) and that production is governed by a positive feedback loop. We then further characterized this system by determining which genes are regulated by quorum sensing in this methane-oxidizer using RNA-seq, and discovered this system regulates the expression of a novel nonribosomal peptide synthetase biosynthetic gene cluster. These results identify and characterize a mode of cellular communication in an aerobic methane-oxidizing bacterium.
Project description:Aerobic methanotrophic bacteria use methane as their sole source of carbon and energy and serve as a major sink for the potent greenhouse gas methane in freshwater ecosystems. Despite this important environmental role, little is known about the molecular details of how these organisms interact in the environment. Many bacterial species use quorum sensing systems to regulate gene expression in a density-dependent manner. We have identified a quorum sensing system in the genome of Methylobacter tundripaludum, a dominant methane-oxidizer in methane enrichments of sediment from Lake Washington (Seattle, WA, USA). We determined that M. tundripaludum primarily produces N-3-hydroxydecanoyl-L-homoserine lactone (3-OH-CÂ10-HSL) and that production is governed by a positive feedback loop. We then further characterized this system by determining which genes are regulated by quorum sensing in this methane-oxidizer using RNA-seq, and discovered this system regulates the expression of a novel nonribosomal peptide synthetase biosynthetic gene cluster. These results identify and characterize a mode of cellular communication in an aerobic methane-oxidizing bacterium. Samples are 2 sets of biological replicates of a Methylobacter tundripaludum strain 21/22 mutant where the acyl-homoserine lactone (AHL) synthase gene mbaI (T451DRAFT_0796) has been deleted. The mutant strain was grown to log (48 hours) or stationary (68 hours) phase in the absence or presence of the AHL 3-OH-C10-HSL.
Project description:Tris(4-chlorophenyl)methane (TCPM) and tris(4-chlorophenyl)methanol (TCPMOH) are two marine pollutants of emerging concern, found in marine mammals and coastal bird populations. Here, we assess the response to these compounds due to acute exposures in zebrafish embryos (from 96-100 hours post fertilization).
Project description:Tris(4-chlorophenyl)methane (TCPM) and tris(4-chlorophenyl)methanol (TCPMOH) are two marine pollutants of emerging concern, found in marine mammals and coastal bird populations. Here, we assess the response to these compounds due to developmental (24-100 hourrs post fertilization) exposures in zebrafish embryos
Project description:RNA-Seq profiling of Methylomicrobium alcaliphilum strain 20Z grown in batch on methane. The RNA-Seq work is one part of a systems approach to characterizing metabolism of 20Z during growth on methane. We demonstrate that methane assimilation is coupled with a highly efficient pyrophosphate-mediated glycolytic pathway, which under O2 limitation participates in a novel form of fermentation-based methanotrophy. This surprising discovery suggests a novel mode of methane utilization in oxygen-limited environments, and opens new opportunities for a modular approach towards producing a variety of excreted chemical products using methane as a feedstock. Four replicates of batch growth
Project description:RNA-Seq profiling of Methylomicrobium alcaliphilum strain 20Z grown in batch on methane. The RNA-Seq work is one part of a systems approach to characterizing metabolism of 20Z during growth on methane. We demonstrate that methane assimilation is coupled with a highly efficient pyrophosphate-mediated glycolytic pathway, which under O2 limitation participates in a novel form of fermentation-based methanotrophy. This surprising discovery suggests a novel mode of methane utilization in oxygen-limited environments, and opens new opportunities for a modular approach towards producing a variety of excreted chemical products using methane as a feedstock.
Project description:Gas hydrates, also known as clathrates, are cages of ice-like water crystals encasing gas molecules such as methane (CH4). Despite the global importance of gas hydrates, their microbiomes remain mysterious. Microbial cells are physically associated with hydrates, and the taxonomy of these hydrate-associated microbiomes is distinct from non-hydrate-bearing sites. Global 16S rRNA gene surveys show that members of sub-clade JS-1 of the uncultivated bacterial candidate phylum Atribacteria are the dominant taxa in gas hydrates. The Atribacteria phylogeny is highly diverse, suggesting the potential for wide functional variation and niche specialization. Here, we examined the distribution, phylogeny, and metabolic potential of uncultivated Atribacteria in cold, salty, and high-pressure sediments beneath Hydrate Ridge, off the coast of Oregon, USA, using a combination of 16S rRNA gene amplicon, metagenomic, and metaproteomic analysis. Methods were developed to extract bacterial cellular protein from these sediments, as outlined below. Sample Description Three sediments samples were collected from beneath Hydrate Ridge, off the coast of Oregon, USA. Sediments were cored at ODP site 1244 (44°35.1784´N; 125°7.1902´W; 895 m water depth) on the eastern flank of Hydrate Ridge ~3 km northeast of the southern summit on ODP Leg 204 in 2002 and stored at -80°C at the IODP Gulf Coast Repository. E10H5 sediment is from 68.5 meters below sediment surface interface C1H2 sediment is from 2 meters below sediment surface interface. C3H4 sediment is from 21 meters below sediment surface interface.
Project description:Methanotrophs, which help regulate atmospheric levels of methane, are active in diverse natural and man-made environments. This range of habitats and the feast-famine cycles seen by many environmental methanotrophs suggest that methanotrophs dynamically mediate rates of methane oxidation. Global methane budgets require ways to account for this variability in time and space. Functional gene biomarker transcripts are increasingly being studied to inform the dynamics of diverse biogeochemical cycles. Previously, per-cell transcript levels of the methane oxidation biomarker, pmoA, were found to vary quantitatively with respect to methane oxidation rates in model aerobic methanotroph, Methylosinus trichosporium OB3b. In the present study, these trends were explored for two additional aerobic methanotroph pure cultures, Methylocystis parvus OBBP and Methylomicrobium album BG8. At steady-state conditions, per cell pmoA mRNA transcript levels strongly correlated with per cell methane oxidation across the three methanotrophs across many orders of magnitude of activity (R2 = 0.91). Additionally, genome-wide expression data (RNA-seq) were used to explore transcriptomic responses of steady state M. album BG8 cultures to short-term CH4 and O2 limitation. These limitations induced regulation of genes involved in central carbon metabolism (including carbon storage), cell motility, and stress response.
Project description:We established simple synthetic microbial communities in a microcosm model system to determine the mechanisms that underlay cross-feeding in microbial methane-consuming communities. Co-occurring strains from Lake Washington sediment were used that are involved in methane consumption, a methanotroph and two non-methanotrophic methylotrophs.