Project description:Supernatant from a stationary phase culture of Methylobacter tundripaludum was run over a C18 solid phase extraction column. The column was washed with 10% methanol in water and then eluted with 90% methanol in water. The eluate was dried and run over a standard LC-MS/MS gradient (ACN in H2O with 0.1% formic acid) focusing on ions with a parent m/z of 150-600. The excreted metabolome of the methane-oxidizing bacterium Methylobacter tundripaludum 21/22 and mutants related to the production of the natural product tundrenone. Supernatant from a stationary phase culture of Methylobacter tundripaludum 21/22 was extracted twice with an equal volume of ethyl acetate containing 0.01% acetic acid. The organic extract was then dried and analyzed over a standard LC-MS/MS gradient (ACN in H2O with 0.1% formic acid). mbaI: Quorum sensing deficient mutant. mbaIplusSig: Quorum sensing deficient mutant grown in the presence of 1 uM exogenous quorum sensing signal 3-OH-C10-HSL. tunJ: Tundrenone deficient mutant lacking the annotated acyl-CoA ligase TunJ.

2017-10-20 | MSV000081637 | GNPS

A complex interplay between nitric oxide, quorum sensing and a unique secondary metabolite tundrenone constitutes hypoxia response in Methylobacter

Project description:Methylobacter species, members of the Methylococcales, have recently emerged as some of the globally widespread, cosmopolitan species, appearing to play a key role in consumption of methane in a variety of environments and across gradients of dioxygen tensions. We approached the question of how Methylobacter copes with low dioxygen partial pressures that it encounters in its natural habitats, via laboratory manipulation. Through comparative transcriptomics of cultures grown under high dioxygen partial pressure and cultures starved for dioxygen, we identified a gene cluster encoding a hybrid cluster protein along with sensing and regulatory functions. Through mutant analysis we demonstrated that this gene cluster is involved in both oxidative and nitrosative stress responses and in NO reduction, while being responsive to NO, likely through direct sensing. Through additional transcriptomic analyses, we further uncovered a complex interconnection between the NO-mediated stress response and quorum sensing in turn controlling synthesis of the secondary metabolite tundrenone, these two systems being inversely regulated. Some of the key methylotrophy functions, such as the two alternative methanol dehydrogenases seem to be parts of the stress response regulatory cascade. One intriguing scenario that emerged from our analyses is that the main role of the respiratory denitrification pathway may be in producing the signaling molecule, NO, in response to hypoxia, rather than in serving as an energy-generating pathway. This novel and complex hypoxia stress response system is unique to Methylobacter tundripaludum, and it may play a role in the environmental fitness of these organisms and in their cosmopolitan environmental distribution.

2019-10-01 | GSE138276 | GEO

QUORUM SENSING IN A 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.

2016-08-16 | E-GEOD-85617 | biostudies-arrayexpress

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