Project description:Until recently, rare-earth elements (REEs) had been thought to be biologically inactive. This view changed with the discovery of the methanol dehydrogenase (Mdh) XoxF that strictly relies on REEs for its activity. Some methylotrophs only contain xoxF, while others, including the model phyllosphere colonizer Methylobacterium extorquens PA1, harbor this gene in addition to mxaFI encoding a Ca2+-dependent enzyme. Here we found that REEs induce the expression of xoxF in M. extorquens PA1, while repressing mxaFI, suggesting that XoxF is the preferred Mdh. Using reporter assays and a suppressor screen, we found that La3+ is sensed both in a XoxF-dependent and independent manner. Furthermore, we investigated the role of REEs during Arabidopsis thaliana colonization. Element analysis of the phyllosphere revealed the presence of several REEs at concentrations up to 10 μg per g dry weight. Complementary proteome analyses of M. extorquens PA1 revealed XoxF as a top induced protein in planta and resulted in the identification of a core set of La3+-regulated proteins under defined artificial media conditions. Among these, we identified a potential REE-binding protein that is encoded next to a gene for a TonB-dependent transporter. The latter was essential for REE-dependent growth on methanol indicating chelator-assisted uptake of REEs.
Project description:The goal of this study was to use microarrays to identify genes differentially regulated under conditions of formaldehyde stress relative to two other stress conditions (oxidative, osmotic) in an effort to identify genes that might be involved in a formaldehyde-specific stress response, rather than a general stress response, in the model methylotroph Methylobacterium extorquens AM1.
Project description:Efforts towards microbial conversion of lignin to value-added products face many challenges because lignin’s methoxylated aromatic monomers release toxic C1 byproducts such as formaldehyde. The ability to grow on methoxylated aromatic acids (e.g., vanillic acid) has recently been identified in certain clades of methylotrophs, bacteria characterized by their unique ability to tolerate and metabolize high concentrations of formaldehyde. Here, we use a phyllosphere methylotroph isolate, Methylobacterium extorquens SLI 505, as a model to identify the fate of formaldehyde during methylotrophic growth on vanillic acids. M. extorquens SLI 505 displays concentration-dependent growth phenotypes on vanillic acid without concomitant formaldehyde accumulation. We conclude that M. extorquens SLI 505 overcomes potential metabolic bottlenecks from simultaneous assimilation of multicarbon and C1 intermediates by allocating formaldehyde towards dissimilation and assimilating the ring carbons of vanillic acid heterotrophically. We correlate this strategy with maximization of bioenergetic yields and demonstrate that formaldehyde dissimilation for energy generation rather than formaldehyde detoxification is advantageous for growth on aromatic acids. M. extorquens SLI 505 also exhibits catabolite repression during growth on methanol and low concentrations of vanillic acid, but no diauxie during growth on methanol and high concentrations of vanillic acid. Results from this study outline metabolic strategies employed by M. extorquens SLI 505 for growth on a complex single substrate that generates both C1 and multicarbon intermediates and emphasizes the robustness of M. extorquens for biotechnological applications for lignin valorization.