Dataset Information

Copper-responsive gene expression in the methanotroph Methylosinus trichosporium OB3b.

ABSTRACT: Methanotrophic bacteria convert methane to methanol using methane monooxygenase (MMO) enzymes. In many strains, either an iron-containing soluble (sMMO) or a copper-containing particulate (pMMO) enzyme can be produced depending on copper availability; the mechanism of this copper switch has not been elucidated. A key player in methanotroph copper homeostasis is methanobactin (Mbn), a ribosomally produced, post-translationally modified natural product with a high affinity for copper. The Mbn precursor peptide is encoded within an operon that contains a range of putative transporters, regulators, and biosynthetic proteins, but the involvement of these genes in Mbn-related processes remains unclear. Extensive time-dependent qRT-PCR studies of Methylosinus trichosporium OB3b and the constitutive sMMO-producing mutant M. trichosporium OB3b PP358 show that the Mbn operon is indeed copper-regulated, providing experimental support for its bioinformatics-based identification. Moreover, the Mbn operon is co-regulated with the sMMO operon and reciprocally regulated with the pMMO operon. Within the Mbn and sMMO operons, a subset of regulatory genes exhibits a distinct and shared pattern of expression, consistent with their proposed functions as internal regulators. In addition, genome sequencing of the M. trichosporium OB3b PP358 mutant provides new evidence for the involvement of genes adjacent to the pMMO operon in methanotroph copper homeostasis.

SUBMITTER: Kenney GE

PROVIDER: S-EPMC6195801 | biostudies-literature | 2016 Sep

REPOSITORIES: biostudies-literature

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Copper-responsive gene expression in the methanotroph Methylosinus trichosporium OB3b.

Kenney Grace E GE Sadek Monica M Rosenzweig Amy C AC

Metallomics : integrated biometal science 20160418 9

Methanotrophic bacteria convert methane to methanol using methane monooxygenase (MMO) enzymes. In many strains, either an iron-containing soluble (sMMO) or a copper-containing particulate (pMMO) enzyme can be produced depending on copper availability; the mechanism of this copper switch has not been elucidated. A key player in methanotroph copper homeostasis is methanobactin (Mbn), a ribosomally produced, post-translationally modified natural product with a high affinity for copper. The Mbn prec ...[more]

PMID: 27087171

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Project description:Methane is a potent greenhouse gas in the atmosphere, and its concentration has continued to increase in recent decades. Aerobic methanotrophs, bacteria that use methane as the sole carbon source, are an important biological sink for methane, and they are widely distributed in the natural environment. However, relatively little is known on how methanotroph activity is regulated by nutrients, particularly phosphorus (P). P is the principal nutrient constraining plant and microbial productivity in many ecosystems, ranging from agricultural land to the open ocean. Using a model methanotrophic bacterium, Methylosinus trichosporium OB3b, we demonstrate here that this bacterium can produce P-free glycolipids to replace membrane phospholipids in response to P limitation. The formation of the glycolipid monoglucuronic acid diacylglycerol requires plcP-agt genes since the plcP-agt mutant is unable to produce this glycolipid. This plcP-agt-mediated lipid remodeling pathway appears to be important for M. trichosporium OB3b to cope with P stress, and the mutant grew significantly slower under P limitation. Interestingly, comparative genomics analysis shows that the ability to perform lipid remodeling appears to be a conserved trait in proteobacterial methanotrophs; indeed, plcP is found in all proteobacterial methanotroph genomes, and plcP transcripts from methanotrophs are readily detectable in metatranscriptomics data sets. Together, our study provides new insights into the adaptation to P limitation in this ecologically important group of bacteria. IMPORTANCE Methane is a potent greenhouse gas in the atmosphere, and its concentration has continued to increase steadily in recent decades. In the natural environment, bacteria known as methanotrophs help mitigate methane emissions at no cost to human beings. However, relatively little is known regarding how methane oxidation activity in methanotrophs is regulated by soil nutrients, particularly phosphorus. Here, we show that methanotrophs can modify their membrane in response to phosphorus limitation and that the ability to change membrane lipids is important for methanotroph activity. Genome and metatranscriptome analyses suggest that such an adaptation strategy appears to be strictly conserved in all proteobacterial methanotrophs and is used by these bacteria in the natural environment. Together, our study provides a plausible molecular mechanism for better understanding the role of phosphorus on methane oxidation in the natural environment.

Project description:Methanotrophs require copper for their activity as it plays a critical role in the oxidation of methane to methanol. To sequester copper, some methanotrophs secrete a copper-binding compound termed methanobactin (MB). MB, after binding copper, is reinternalized via a specific outer membrane TonB-dependent transporter (TBDT). Methylosinus trichosporium OB3b has two such TBDTs (MbnT1 and MbnT2) that enable M. trichosporium OB3b to take up not only its own MB (MB-OB3b) but also heterologous MB produced from other methanotrophs, e.g., MB of Methylocystis sp. strain SB2 (MB-SB2). Here, we show that uptake of copper in the presence of heterologous MB-SB2 can either be achieved by initiating transcription of mbnT2 or by using its own MB-OB3b to extract copper from MB-SB2. Transcription of mbnT2 is mediated by the N-terminal signaling domain of MbnT2 together with an extracytoplasmic function sigma factor and an anti-sigma factor encoded by mbnI2 and mbnR2, respectively. Deletion of mbnI2R2 or excision of the N-terminal region of MbnT2 abolished induction of mbnT2. However, copper uptake from MB-SB2 was still observed in M. trichosporium OB3b mutants that were defective in MbnT2 induction/function, suggesting another mechanism for uptake copper-loaded MB-SB2. Additional deletion of MB-OB3b synthesis genes in the M. trichosporium OB3b mutants defective in MbnT2 induction/function disrupted their ability to take up copper in the presence of MB-SB2, indicating a role of MB-OB3b in copper extraction from MB-SB2. IMPORTANCE Methanotrophs play a critical role in the global carbon cycle, as well as in future strategies for mitigating climate change through their consumption of methane, a trace atmospheric gas much more potent than carbon dioxide in global warming potential. Copper uptake is critical for methanotrophic activity, and here, we show different approaches for copper uptake. This study expands our knowledge and understanding of how methanotrophs collect and compete for copper, and such information may be useful in future manipulation of methanotrophs for a variety of environmental and industrial applications.

Dataset Information

Copper-responsive gene expression in the methanotroph Methylosinus trichosporium OB3b.

Publications

Copper-responsive gene expression in the methanotroph Methylosinus trichosporium OB3b.

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