New papers from Gutjahr lab.
The Lotus japonicus acyl-acyl carrier protein thioesterase FatM is required for mycorrhiza formation and lipid accumulation of Rhizophagus irregularis.
Mathias Brands, Vera Wewer, Andreas Keymer, Caroline Gutjahr, Peter Dörmann
Arbuscular mycorrhiza (AM) fungi establish symbiotic interactions with plants, providing the host plant with minerals, i.e. phosphate, in exchange with organic carbon. AM fungi of the Glominea produce vesicles, which store lipids as energy and carbon source. Acyl-acyl carrier protein (ACP) thioesterases (Fat) are essential components of the plant plastid?localized fatty acid synthase and determine the chain length of de novosynthesized fatty acids. In addition to the ubiquitous FatA and FatB thioesterases, AM?competent plants contain an additional, AM-specific FatM gene. Here, we characterize FatM from Lotus japonicus by phenotypically analyzing fatm mutant lines and by studying the biochemical function of the recombinant FatM protein. Reduced shoot phosphate content in fatm indicates compromised symbiotic phosphate uptake due to reduced arbuscule branching, and the fungus shows reduced lipid accumulation accompanied with the occurrence of smaller and less frequent vesicles. Lipid profiling reveals a decrease in mycorrhiza-specific phospholipid forms, AM fungal signature fatty acids (e.g. 16:1w5, 18:1w7 and 20:3) and storage lipids. Recombinant FatM shows preference for palmitoyl (16:0)-ACP, indicating that large amounts of 16:0 fatty acid are exported from the plastids of arbuscule containing cells. Stable isotope labeling with [13C2]acetate showed reduced incorporation into mycorrhiza?specific fatty acids in the fatm mutant. Therefore, colonized cells reprogram plastidial fatty acid de novo synthesis towards the production of extra amounts of 16:0, which is in agreement with previous results that fatty acid-containing lipids are transported from the plant to the fungus.
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Tracking lipid transfer by fatty acid isotopolog profiling from host plants to arbuscular mycorrhiza fungi.
Andreas Keymer, Claudia Huber, Wolfgang Eisenreich, Caroline Gutjahr
Bio-protocol
Lipid transfer from host plants to arbuscular mycorrhiza fungi was hypothesized for several years because sequenced arbuscular mycorrhiza fungal genomes lack genes encoding cytosolic fatty acid synthase (Wewer et al., 2014; Rich et al., 2017). It was finally shown by two independent experimental approaches (Jiang et al., 2017; Keymer et al., 2017; Luginbuehl et al., 2017). One approach used a technique called isotopolog profiling (Keymer et al., 2017). Isotopologs are molecules, which differ only in their isotopic composition. For isotopolog profiling an organism is fed with a heavy isotope labelled precursor metabolite. Subsequently, the labelled isotopolog composition of metabolic products is analysed via mass spectrometry. The detected isotopolog pattern of the metabolite(s) of interest yields information about metabolic pathways and fluxes (Ahmed et al., 2014). The following protocol describes an experimental setup, which enables separate isotopolog profiling of fatty acids in plant roots colonized by arbuscular mycorrhiza fungi and their associated fungal extraradical mycelium, to elucidate fluxes between both symbiotic organisms. We predict that this strategy can also be used to study metabolite fluxes between other organisms if the two interacting organisms can be physically separated.
