Mildew resistance Locus O (MLO) proteins are polytopic integral membrane proteins that have long been considered as plant-specific and being primarily involved in plant–powdery mildew interactions. However, research in the past decade has revealed that MLO proteins diverged into a family with several clades whose members are associated with different physiological processes. We provide a largely increased dataset of MLO amino acid sequences, comprising nearly all major land plant lineages. Based on this comprehensive dataset, we defined seven phylogenetic clades and reconstructed the likely evolution of the MLO family in embryophytes. We further identified several MLO peptide motifs that are either conserved in all MLO proteins or confined to one or several clades, supporting the notion that clade-specific diversification of MLO functions is associated with particular sequence motifs. In baker’s yeast, some of these motifs are functionally linked to transmembrane (TM) transport of organic molecules and ions. In addition, we attempted to define the evolutionary origin of the MLO family and found that MLO-like proteins with highly diverse membrane topologies are present in green algae, but also in the distinctly related red algae (Rhodophyta), Amoebozoa, and Chromalveolata. Finally, we discovered several instances of putative fusion events between MLO proteins and different kinds of proteins. Such Rosetta stone-type hybrid proteins might be instructive for future analysis of potential MLO functions. Our findings suggest that MLO is an ancient protein that possibly evolved in unicellular photosynthetic eukaryotes, and consolidated in land plants with a conserved topology, comprising seven TM domains and an intrinsically unstructured C-terminus.

Plants encounter beneficial and detrimental microorganisms both above- and belowground and the health status of the plant depends on the composition of this pan-microbiome. Beneficial microorganisms contribute to plant nutrition or systemically or locally protect plants against pathogens, thus facilitating adaptation to a variety of environments. Induced systemic resistance, caused by root-associated microbes, manifests as aboveground resistance against necrotrophic pathogens and is mediated by jasmonic acid/ethylene-dependent signaling. By contrast, systemic acquired resistance relies on salicylic acid (SA) signaling and confers resistance against secondary infection by (hemi)biotrophic pathogens. To investigate whether symbiotic rhizobia that are ubiquitously found in natural ecosystems are able to modulate resistance against biotrophs, we tested the impact of preestablished nodulation of Medicago truncatula and pea (Pisum sativum) plants against infection by the powdery mildew fungus Erysiphe pisi. We found that root symbiosis interfered with fungal penetration of M. truncatula and reduced asexual spore formation on pea leaves independently of symbiotic nitrogen fixation. Improved resistance of nodulated plants correlated with elevated levels of free SA and SA-dependent marker gene expression upon powdery mildew infection. Our results suggest that nodulation primes the plants systemically for E. pisi-triggered SA accumulation and defense gene expression, resulting in increased resistance.

Maize, a main crop worldwide, establishes a mutualistic symbiosis with arbuscular mycorrhizal (AM) fungi providing nutrients to the roots from soil volumes which are normally not in reach of the non-colonized root. The mycorrhizal phosphate uptake pathway (MPU) spans from extraradical hyphae to root cortex cells housing fungal arbuscules and promotes the supply of phosphate to the mycorrhizal host in exchange for photosynthetic carbon. This symbiotic association with the mycobiont has been shown to affect plant host nutritional status and growth performance. However, whether and how the MPU affects the root microbial community associated with mycorrhizal hosts in association with neighboring plants, remains to be demonstrated. Here the maize germinal Mu transposon insertion mutant pht1;6, defective in mycorrhiza-specific Pi transporter PHT1;6 gene, and wild type B73 (wt) plants were grown in mono-and mixed culture and examined under greenhouse and field conditions. Disruption of the MPU in pht1;6 resulted in strongly diminished growth performance, in reduced P allocation to photosynthetic source leaves, and in imbalances in leaf elemental composition beyond P. At the microbial community level a loss of MPU activity had a minor effect on the root-associated fungal microbiome which was almost fully restricted to AM fungi of the Glomeromycotina. Moreover, while wt grew better in presence of pht1;6, pht1;6 accumulated little biomass irrespective of whether it was grown in mono-or mixed culture and despite of an enhanced fungal colonization of its roots in co-culture with wt. This suggested that a functional MPU is prerequisite to maintain maize growth and that neighboring plants competed for AM fungal Pi in low P soil. Thus future strategies towards improving yield in maize populations on soils with low inputs of P fertilizer could be realized by enhancing MPU at the individual plant level while leaving the root-associated fungal community largely unaffected.

Hosts and pathogens typically engage in an evolutionary arms race. This also applies to phytopathogenic powdery mildew fungi, which can rapidly overcome plant resistance and perform host jumps. Using experimental evolution, we show that the powdery mildew pathogen Blumeria graminis f.sp. hordei is capable of breaking the agriculturally important broad-spectrum resistance conditioned by barley loss-of-function mlo mutants. Partial mlo virulence is associated with a distinctive pattern of adaptive mutations, including small-sized (8-40 kb) deletions, one of which likely affects spore morphology. The detected mutational spectrum comprises the same loci in at least two independent mlo-virulent isolates, indicating convergent multigenic evolution. This work highlights the dynamic genome evolution of an obligate biotrophic plant pathogen with a transposon-enriched genome.