For more than 400 million years, plants have maintained a mutualistic symbiosis with arbuscular mycorrhizal (AM) fungi. This evolutionary success can be traced to the role of these fungi in providing plants with mineral nutrients, particularly phosphate. In return, photosynthates are given to the fungus, which support its obligate biotrophic lifestyle. Although the mechanisms involved in phosphate transfer have been extensively studied, less is known about the reciprocal transfer of carbon. Here, we present the high-affinity Monosaccharide Transporter2 (MST2) from Glomus sp with a broad substrate spectrum that functions at several symbiotic root locations. Plant cell wall sugars can efficiently outcompete the Glc uptake capacity of MST2, suggesting they can serve as alternative carbon sources. MST2 expression closely correlates with that of the mycorrhiza-specific Phosphate Transporter4 (PT4). Furthermore, reduction of MST2 expression using host-induced gene silencing resulted in impaired mycorrhiza formation, malformed arbuscules, and reduced PT4 expression. These findings highlight the symbiotic role of MST2 and support the hypothesis that the exchange of carbon for phosphate is tightly linked. Unexpectedly, we found that the external mycelium of AM fungi is able to take up sugars in a proton-dependent manner. These results imply that the sugar uptake system operating in this symbiosis is more complex than previously anticipated.

Summary• The arbuscular mycorrhizal symbiosis is arguably the most ecologically important eukaryotic symbiosis, yet it is poorly understood at the molecular level. To provide novel insights into the molecular basis of symbiosis-associated traits, we report the first genome-wide analysis of the transcriptome from Glomus intraradices DAOM 197198.• We generated a set of 25 906 nonredundant virtual transcripts (NRVTs) transcribed in germinated spores, extraradical mycelium and symbiotic roots using Sanger and 454 sequencing. NRVTs were used to construct an oligoarray for investigating gene expression.• We identified transcripts coding for the meiotic recombination machinery, as well as meiosis-specific proteins, suggesting that the lack of a known sexual cycle in G. intraradices is not a result of major deletions of genes essential for sexual reproduction and meiosis. Induced expression of genes encoding membrane transporters and small secreted proteins in intraradical mycelium, together with the lack of expression of hydrolytic enzymes acting on plant cell wall polysaccharides, are all features of G. intraradices that are shared with ectomycorrhizal symbionts and obligate biotrophic pathogens.• Our results illuminate the genetic basis of symbiosis-related traits of the most ancient lineage of plant biotrophs, advancing future research on these agriculturally and ecologically important symbionts.*These authors contributed equally to this work.

Summary• Here, arbuscular mycorrhizal (AM) fungi were monitored in vivo introducing the fluorescent reporters DsRed and GFP (green fluorescent protein) in Glomus intraradices using a biolistic approach and Agrobacterium tumefaciens -mediated transformation.• Both reporter genes were fused to the nuclear localization signal of the Aspergillus nidulans transcription factor StuA to target fluorescence to nuclei. Expression of DsRed was driven by two Glomus mosseae promoters highly expressed during early symbiosis, GmPMA1 and GmFOX2 , while expression of GFP was driven by the A. nidulans gpd promoter.• All promoters worked in G. intraradices as well as in A. nidulans . Red and green fluorescence was localized to nuclei of G. intraradices spores and hyphae 3 d after bombardment. However, expression was transient. The efficiency of the Agrobacterium -mediated transformation was very low.• These results indicate that the biolistic method allows the expression of foreign DNA into G. intraradices with high frequency, but it is insufficient to render stable transformants. DsRed vs GFP is a more appropriate living reporter to be used in G. intraradices because of the lower autofluorescence in the red channel but targeted to the nucleus both marker genes can be visualized. This is the first report of fluorescent marker expression in an AM fungus driven by arbuscular mycorrhizal promoters.

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