Innate Immunity and the Plant Microbiota
I. Structure and functions of the Arabidopsis root microbiota (supported by ERC advanced grant ROOTMICROBIOTA)
Remarkably little is understood about plant-microbe interactions that are, at first glance, ‘symptomless’. These poorly studied plant microbiomes harbor an unknown reservoir of probiotic and plant protective associations. One gram of soil typically contains ~108 to 1010 bacteria. Microbial DNA fingerprints from plant roots or aerial organs uncover microbial communities thriving on the surface of, or within, healthy plant tissue. Rhizosphere microbiomes attached to, and in the first few millimeters away from, the root surface are distinct from bulk soil, suggesting specific colonization events. Organic carbon flux from roots promotes the growth of microbial decomposers that, in turn, recycle plant nutrients for root uptake by transpiration-driven water fluxes. Seedlings exude 30-40%, and adult plants 20%, of photosynthetically fixed carbon into the rhizosphere in the form of poorly characterized rhizodeposits.
We have developed and applied bacterial 16S rRNA gene pyrosequencing to characterize and compare soil and root-inhabiting bacterial communities of Arabidopsis thaliana. We show that roots of Arabidopsis, grown in different natural soils under controlled environmental conditions, are preferentially colonized by Proteobacteria, Bacteroidetes and Actinobacteria, and each bacterial phylum is represented by a dominating class or family. Soil type defines the composition of root-inhabiting bacterial communities and host genotype determines their ribotype profiles to a limited extent. The identification of soil type-specific members within the root-inhabiting assemblies supports our conclusion that these represent soil-derived root endophytes. Surprisingly, plant cell wall features of other tested plant species appear to provide a sufficient cue for the assembly of ~40% of the Arabidopsis bacterial root-inhabiting microbiota, with a bias for Betaproteobacteria. Thus, this root sub-community may not be Arabidopsis-specific but saprophytic bacteria that would naturally be found on any plant root or plant debris in the tested soils. In contrast, colonization of Arabidopsis roots by members of the Actinobacteria depends on additional cues from metabolically active host cells (Bulgarelli et al., Nature 488, 91-95, 2012).
Garrido-Oter, R., Nakano, R. T., Dombrowski, N., Ma, K.-W., The AgBiome Team, McHardy, A. C. and P. Schulze-Lefert: Modular traits of the Rhizobiales root microbiota and their evolutionary relationship with symbiotic rhizobia. Cell Host & Microbe 24, 155–167 (2018). abstract
Hacquard, S., Spaepen, S., Garrido‐Oter R. and Schulze ‐ Lefert, P. Interplay between innate immunity and the plant microbiota. Annu. Rev. Phytopathol. 55, 55, doi: 10.1146/annurev-phyto-080516-035623 (2017). abstract
Dombrowski, N., Schlaeppi, K., Agler, M., Hacquard, S., Kemen, E., Garrido Oter, R., Wunder, J., Coupland, G. and Schulze-Lefert, P., Root microbiota dynamics of perennial Arabis alpina are dependent on soil residence time but independent of flowering time. ISME Journal 11, 43–55 (2017). abstract
Zgadzaj#, R., Garrido-Oter#, R., Jensen, D.B., Schulze-Lefert* P. and Radutoiu* S. Root nodule symbiosis in Lotus japonicus drives the establishment of distinctive rhizosphere, root, and nodule bacterial communities. PNAS 113 (49): E7996–E800, DOI: 10.1073/pnas.1616564113 (2016). #joint first authors, *co-corresponding authors abstract
Hacquard, S., Kracher, B., Hiruma, K, Münch,P. C., Garrido-Oter, R. Thon, M. R., Weimann, A., Damm, U., Dallery,J.-F., Hainaut,M., Henrissat, B., Lespinet, O., Sacristán, S., Ver Loren van Themaat,E., Kemen, E., McHardy, A.C., Schulze-Lefert, P.*, O’Connell, R.* Survival trade-offs in plant roots during colonization by closely related beneficial and pathogenic fungi. Nature Communications 7: 11362, doi: 10.1038/ncomms11362 (2016). *Corresponding authors absctract
Hiruma, K., Gerlach, N., Sacristán, S., Nakano, R. T., Hacquard, S., Kracher, B., Neumann, U., Ramírez, D., Bucher, M, O’Connell, R.* and Schulze-Lefert, P.* Root endophyte Colletotrichum tofieldiae confers plant fitness benefits that are phosphate status-dependent. Cell 165, 464-474 (2016), published online 17 March 2016, doi:10.1016/j.cell.2016.02.028. *Corresponding authors
- This work was featured in: Cell 165, 269-271 (2016) by Louisa M. Liberman and Philip N. Benfey: A Friend in Need (of Nutrients) Is a …
- This article has been recommended by Faculty 1000:
Bai, Y., Müller, D. B., Srinivas, G., Garrido-Oter, R., Potthoff, E., Rott, M., Dombrowski, N., Münch, P.C., Spaepen, S., Remus-Emsermann, M., Hüttel, B., McHardy, A., Vorholt, J.A., and Schulze-Lefert, P. Functional overlap of the Arabidopsis leaf and root microbiota. Nature 528, 364-369 (2015), doi:10.1038/nature16192.abstract
Hacquard, S., Garrido-Oter, R., González, A., Spaepen, S., Ackermann, G., Lebeis, S., McHardy, A. C., Dangl, J. L., Knight, R., Ley, R., Schulze-Lefert, P. Microbiota and Host Nutrition across Plant and Animal Kingdoms.Cell Host & Microbe 17 603-616 (2015). abstract
Bulgarelli, D., Garrido-Oter, R., Münch, P.C., Weiman, A., Dröge, J., Pan, Y., McHardy, A.C. and Schulze-Lefert, P. (2015) Structure and functions of the bacterial root microbiota in wild and domesticated barley. Cell Host & Microbe 17, 392-404 (2015). abstract
Schlaeppi, K., Dombrowski, N., Garrido Oter, R., ver Loren van Themaat, E. and Schulze-Lefert, P. (2014) Quantitative divergence of the bacterial root microbiota in Arabidopsis thaliana relatives, PNAS, 111(2), 585-592. abstract
Bulgarelli, D., Schlaeppi, K., Spaepen, S., Ver Loren van Themaat, E. and Schulze-Lefert, P. Structure and Functions of the Bacterial Microbiota of Plants. Annual Review of Plant Biology Vol. 64 (2013). abstract
Bulgarelli, D., Rott, M., Schlaeppi, K., Ver Loren van Themaat, E., Ahmadinejad, N, Assenza, F., Rauf, P., Huettel, B., Reinhardt R., Schmelzer, E., Peplies, J., Gloeckner, F., Amann, R., Eickhorst, T., and Schulze-Lefert P. Revealing structure and assembly cues for Arabidopsis root-inhabiting bacterial microbiota. Nature 488, 91-95 (2012). abstract
Lebeis, S.L., Rott, M., Dangl, J.L.* and Schulze-Lefert, P.*: Culturing a plant microbiome at the cross-Rhodes, New Phytologist 196, 341–344 (2012). abstract
Bisseling, T., Dangl, J. and Schulze-Lefert, P. (2009), Next Generation Communication, Science Editorial, Science 324, p.691, 8 May (2009). abstract
Current and future work
We have begun to systematically purify Arabidopsis bacterial endophytes on the basis of the aforementioned culture-independent 16S rRNA gene survey. We also examine presumed probiotic functions of the root microbiota for plant growth using defined bacterial communities under laboratory conditions.
Stijn Spaepen (post-doc), Yang Bai (post-doc), Stéphane Hacquard (post-doc), Rafal Zgadzaj (post-doc), Ryohei Thomas Nakano (post-doc), Masayoshi Hashimoto (post-doc), Ruben Garrido Oter (PhD), Paloma Durán Ballesteros (PhD), Amine Hassani (PhD), Thorsten Thiergart (bioinformatician), Petra Köchner (technical assistant), Simone Remmel (technical assistant), Brigitte Pickel (technical assistant), Wolfgang Schmalenbach (technical assistant).
II. Non-self perception and signaling by intracellular immune sensors
One class of immune receptors, designated NB-LRR proteins, detects isolate-specific pathogen effector molecules inside plant cells. The Mla locus in barley encodes coiled-coil (CC) domain, nucleotide-binding (NB) site, and leucine-rich repeat (LRR)-containing receptor proteins and mediate isolate-specific immunity to the grass powdery mildew fungus (Blumeria graminis f. sp. hordei). Genetic studies identified a large number of allelic variants at the Mla locus each recognizing a cognate race-specific B. graminis effector (AVRA). We have shown that barley MLA partitions between the cytoplasm and nucleus. We have also shown that nucleocytoplasmic partitioning of the receptor results in a bifurcation of MLA‐triggered host cell death and disease resistance signaling in a cell compartment‐dependent manner. MLA-mediated host cell death signaling is initiated in the cytoplasm whereas disease resistance signaling takes place in the nucleus. Recognition of B. graminis AVRA effectors by MLA induces nuclear associations between the receptor and WRKY transcription factors. These WRKY proteins act as repressors of MAMP (also called PAMP-)-triggered basal defense. MLA appears to interfere with the WRKY repressor function in the nucleus, thereby de-repressing MAMP-triggered defense. Collectively, our findings imply a mechanism by which the polymorphic MLA immune receptors integrate distinct pathogen signals and initiate a bifurcated signal transduction cascade. Our structural and biochemical work revealed a CC domain-dependent homodimerization of the immune sensor that is necessary for receptor function. Notably, the CC homodimer module defines a minimal functional unit, which is sufficient to activate host cell death signaling.
Lu, X., Kracher, B., Saur, I.M.-L., Bauer, S., Ellwood, S.R., Wise, R., Yaeno, T., Maekawa, T., and Schulze-Lefert, P.: Allelic barley MLA immune receptors recognize sequence-unrelated avirulence effectors of the powdery mildew pathogen. PNAS 113 (42), E6486–E6495, doi: 10.1073/pnas.1612947113 (2016) abstract
Jacob, F.; Vernaldi, S. and Maekawa, T: Evolution and conservation of plant NLR functions. Frontiers in Immunology 4:297, doi 10.3389/fimmu.2013.00297 (2013). abstract
Bai, S., Liu, J., Chang Ch., Zhang, L., Maekawa, T., Wang, Q., Xiao, W., Liu, Y., Chai, J., Takken, FLW., Schulze-Lefert, P., Shen, QH. (2012) Structure-Function Analysis of Barley NLR immune Receptor MLA10 Reveals Its Cell Compartment Specific Activity in Cell Death and Disease Resistance. PLoS Pathogens 8(6), e1002752
Maekawa, T., Kracher, B., Vernaldi, S., Ver Loren van Themaat, E. and Schulze-Lefert, P. (2012), Conservation of NLR-triggered immunity across plant lineages. PNAS 49, 20119-20123 (2012). abstract
Maekawa, T., Cheng, W., Spiridon, LN., Töller, A. Lukasik, E., Saijo, Y., Liu, P.; Shen, Q. Micluta, MA., Somssich, IE., Takken, FL., Petrescu, A.-J., Chai, J. and Schulze-Lefert, P. (2011) Coiled-coil domain-dependent homodimerization of intracellular MLA immune receptors defines a minimal functional module for triggering cell death. Cell Host & Microbe 9, 187-199.
Shen, Q.-H., Saijo Y., Mauch S., Biskup C., Bieri S, Keller B, Seki H., lker B., Somssich I.E., and Schulze-Lefert P. (2007): Nuclear Activity of MLA Immune Receptors Links Isolate-Specific and Basal Disease-Resistance Responses. Science 315: 1098-1103.
This work was featured in:
- Cell 128: 823-824, 2007. Jen Sheen and Ping He: Nuclear Actions in Innate Immune Signaling
- Science 315: 1088-1089, 2007. Jeff L. Dangl: Nibbling at the Plant Cell Nucleus
Bieri S, Mauch S, Shen Q-H, Peart J, Devoto A, Casais C, Ceron F, Schulze S, Steinbiss H.-H., Shirasu K, and Schulze-Lefert P (2004) RAR1 positively controls steady state levels of barley MLA resistance proteins and enables sufficient MLA6 accumulation for effective resistance. The Plant Cell 16, 3480-3495.
Shen Q.-H., Zhou F., Bieri S., Haizel T., Shirasu K., and Schulze-Lefert P. (2003) Recognition specificity and RAR1/SGT1 dependence in barley Mla disease resistance genes to the powdery mildew fungus. Plant Cell 15: 732-744.
Current and future work
We aim at a deep understanding of MLA immune receptor function and evolution. Our recent experiments revealed that MLA is fully functional in Arabidopsis which diverged from barley ~150 My ago, thereby enabling comparative studies on MLA function in the model plant Arabidopsis and the crop barley. We aim to resolve the structure of individual MLA domains and of the full-length receptor in collaboration with the structural biology group of Jijie Chai, Beijing. We explore cytoplasmic and nuclear functions of MLA and wish to understand how this immune sensor reconfigures host transcription upon recognition of powdery mildew effectors in barley and Arabidopsis (in collaboration with Imre Somssich). We will also clarify whether MLA receptors detect AVRA effectors directly or indirectly.
Takaki Maekawa (Senior scientist), Xunli Lu (post-doc), Isabel Saur (post-doc), Makoto Yoshikawa-Maekawa (post-doc), Barbara Kracher (bioinformatician), Florence Jacob (PhD), Saskia Bauer (PhD), Petra Köchner (technical assistant), Sabine Haigis (Technical assistant).