@article{hornstein_charles_franklin_edwards_vintila_kleiner_sederoff_2024, title={<i>IPD3</i>, a master regulator of arbuscular mycorrhizal symbiosis, affects genes for immunity and metabolism of non-host <i>Arabidopsis</i> when restored long after its evolutionary loss}, volume={114}, ISSN={["1573-5028"]}, url={https://doi.org/10.1007/s11103-024-01422-3}, DOI={10.1007/s11103-024-01422-3}, abstractNote={Abstract Arbuscular mycorrhizal symbiosis (AM) is a beneficial trait originating with the first land plants, which has subsequently been lost by species scattered throughout the radiation of plant diversity to the present day, including the model Arabidopsis thaliana . To explore if elements of this apparently beneficial trait are still present and could be reactivated we generated Arabidopsis plants expressing a constitutively active form of Interacting Protein of DMI3 , a key transcription factor that enables AM within the Common Symbiosis Pathway, which was lost from Arabidopsis along with the AM host trait. We characterize the transcriptomic effect of expressing IPD3 in Arabidopsis with and without exposure to the AM fungus (AMF) Rhizophagus irregularis , and compare these results to the AM model Lotus japonicus and its ipd3 knockout mutant cyclops-4 . Despite its long history as a non-AM species, restoring IPD3 in the form of its constitutively active DNA-binding domain to Arabidopsis altered expression of specific gene networks. Surprisingly, the effect of expressing IPD3 in Arabidopsis and knocking it out in Lotus was strongest in plants not exposed to AMF, which is revealed to be due to changes in IPD3 genotype causing a transcriptional state, which partially mimics AMF exposure in non-inoculated plants. Our results indicate that molecular connections to symbiosis machinery remain in place in this nonAM species, with implications for both basic science and the prospect of engineering this trait for agriculture.}, number={2}, journal={PLANT MOLECULAR BIOLOGY}, author={Hornstein, Eli D. and Charles, Melodi and Franklin, Megan and Edwards, Brianne and Vintila, Simina and Kleiner, Manuel and Sederoff, Heike}, year={2024}, month={Apr} }
 @article{parnell_vintila_tang_wagner_kleiner_2023, title={Evaluation of ready-to-use freezer stocks of a synthetic microbial community for maize root colonization}, volume={12}, ISSN={["2165-0497"]}, url={https://doi.org/10.1128/spectrum.02401-23}, DOI={10.1128/spectrum.02401-23}, abstractNote={ABSTRACT Synthetic microbial communities (SynComs) are a valuable tool to study community assembly patterns, host–microbe interactions, and microbe–microbe interactions in a fully controllable setting. Constructing the SynCom inocula for plant–microbe experiments can be time-consuming and difficult because a large number of isolates with different medium requirements and growth rates are grown in parallel and mixed to appropriate titers. A potential workaround to assembling fresh SynCom inocula for every experiment could be to prepare and freeze SynComs on a large scale, creating ready-to-use inocula. The objective of this study was to compare the reproducibility, stability, and colonization ability of freshly prepared versus frozen SynCom inocula. We used a community of seven species known to colonize maize roots. The results from inoculation with the frozen SynCom were as consistent as those of standardized de novo construction of fresh SynCom. Our results indicate that creating frozen SynCom inocula for repeated use in experiments not only saves time but could also improve cross-experiment reproducibility. Although this approach was only validated with one SynCom, it demonstrates a principle that can be tested for improving approaches in constructing other SynComs. IMPORTANCE Synthetic communities (SynComs) are an invaluable tool to characterize and model plant–microbe interactions. Multimember SynComs approximate intricate real-world interactions between plants and their microbiome, but the complexity and time required for their construction increase enormously for each additional member added to the SynCom. Therefore, researchers who study a diversity of microbiomes using SynComs are looking for ways to simplify the use of SynComs. In this manuscript, we evaluate the feasibility of creating ready-to-use freezer stocks of a well-studied seven-member SynCom for maize roots. The frozen ready-to-use SynCom stocks work according to the principle of “just add buffer and apply to sterilized seeds or seedlings” and thus can save time applied in multiple days of laborious growing and combining of multiple microorganisms. We show that ready-to-use SynCom stocks provide comparable results to those of freshly constructed SynComs and thus allow for significant time savings when working with SynComs. Synthetic communities (SynComs) are an invaluable tool to characterize and model plant–microbe interactions. Multimember SynComs approximate intricate real-world interactions between plants and their microbiome, but the complexity and time required for their construction increase enormously for each additional member added to the SynCom. Therefore, researchers who study a diversity of microbiomes using SynComs are looking for ways to simplify the use of SynComs. In this manuscript, we evaluate the feasibility of creating ready-to-use freezer stocks of a well-studied seven-member SynCom for maize roots. The frozen ready-to-use SynCom stocks work according to the principle of “just add buffer and apply to sterilized seeds or seedlings” and thus can save time applied in multiple days of laborious growing and combining of multiple microorganisms. We show that ready-to-use SynCom stocks provide comparable results to those of freshly constructed SynComs and thus allow for significant time savings when working with SynComs.}, journal={MICROBIOLOGY SPECTRUM}, author={Parnell, J. Jacob and Vintila, Simina and Tang, Clara and Wagner, Maggie R. and Kleiner, Manuel}, editor={Hockett, Kevin LorenEditor}, year={2023}, month={Dec} }
 @article{zvi-kedem_vintila_kleiner_tchernov_rubin-blum_2023, title={Metabolic handoffs between multiple symbionts may benefit the deep-sea bathymodioline mussels}, volume={3}, ISSN={["2730-6151"]}, DOI={10.1038/s43705-023-00254-4}, abstractNote={Bathymodioline mussels rely on thiotrophic and/or methanotrophic chemosynthetic symbionts for nutrition, yet, secondary heterotrophic symbionts are often present and play an unknown role in the fitness of the organism. The bathymodioline Idas mussels that thrive in gas seeps and on sunken wood in the Mediterranean Sea and the Atlantic Ocean, host at least six symbiont lineages that often co-occur. These lineages include the primary symbionts chemosynthetic methane- and sulfur-oxidizing gammaproteobacteria, and the secondary symbionts, Methylophagaceae, Nitrincolaceae and Flavobacteriaceae, whose physiology and metabolism are obscure. Little is known about if and how these symbionts interact or exchange metabolites. Here we curated metagenome-assembled genomes of Idas modiolaeformis symbionts and used genome-centered metatranscriptomics and metaproteomics to assess key symbiont functions. The Methylophagaceae symbiont is a methylotrophic autotroph, as it encoded and expressed the ribulose monophosphate and Calvin-Benson-Bassham cycle enzymes, particularly RuBisCO. The Nitrincolaceae ASP10-02a symbiont likely fuels its metabolism with nitrogen-rich macromolecules and may provide the holobiont with vitamin B12. The Urechidicola (Flavobacteriaceae) symbionts likely degrade glycans and may remove NO. Our findings indicate that these flexible associations allow for expanding the range of substrates and environmental niches, via new metabolic functions and handoffs.}, number={1}, journal={ISME COMMUNICATIONS}, author={Zvi-Kedem, Tal and Vintila, Simina and Kleiner, Manuel and Tchernov, Dan and Rubin-Blum, Maxim}, year={2023}, month={May} }
 @article{salvato_vintila_finkel_dangl_kleiner_2022, title={Evaluation of Protein Extraction Methods for Metaproteomic Analyses of Root-Associated Microbes}, volume={35}, ISSN={["1943-7706"]}, url={https://doi.org/10.1094/MPMI-05-22-0116-TA}, DOI={10.1094/MPMI-05-22-0116-TA}, abstractNote={Metaproteomics is a powerful tool for the characterization of metabolism, physiology and functional interactions in microbial communities, including plant-associated microbiota. However, the metaproteomic methods that have been used to study plant-associated microbiota are very laborious and require large amounts of plant tissue; hindering, wider application of these methods. We optimized and evaluated different protein extraction methods for metaproteomics of plant-associated microbiota in two different plant species (Arabidopsis and maize). Our main goal was to identify a method that would work with low amounts of input material (40-70 mg) and that would maximize the number of identified microbial proteins. We tested eight protocols, each comprising a different combination of physical lysis method, extraction buffer, and cell enrichment method on roots from plants grown with synthetic microbial communities. We assessed the performance of the extraction protocols by LC-MS/MS-based metaproteomics and found that the optimal extraction method differed between the two species. For Arabidopsis roots protein extraction by beating whole roots with small beads provided the highest number of identified microbial proteins and improved the identification of proteins from gram-positive bacteria. For maize, vortexing root pieces in the presence of large glass beads yielded the highest number of microbial proteins identified. Based on these data we recommend the use of these two methods for metaproteomics with Arabidopsis and maize. Furthermore, detailed descriptions of the eight tested protocols will enable future optimization of protein extraction for metaproteomics in other dicot and monocot plants.}, number={11}, journal={MOLECULAR PLANT-MICROBE INTERACTIONS}, author={Salvato, Fernanda and Vintila, Simina and Finkel, Omri M. and Dangl, Jeffery L. and Kleiner, Manuel}, year={2022}, month={Nov}, pages={977–988} }
 @article{wagner_tang_salvato_clouse_bartlett_vintila_phillips_sermons_hoffmann_balint-kurti_et al._2021, title={Microbe-dependent heterosis in maize}, volume={118}, ISSN={0027-8424 1091-6490}, url={http://dx.doi.org/10.1073/pnas.2021965118}, DOI={10.1073/pnas.2021965118}, abstractNote={Hybrids account for nearly all commercially planted varieties of maize and many other crop plants because crosses between inbred lines of these species produce first-generation [F1] offspring that greatly outperform their parents. The mechanisms underlying this phenomenon, called heterosis or hybrid vigor, are not well understood despite over a century of intensive research. The leading hypotheses-which focus on quantitative genetic mechanisms (dominance, overdominance, and epistasis) and molecular mechanisms (gene dosage and transcriptional regulation)-have been able to explain some but not all of the observed patterns of heterosis. Abiotic stressors are known to impact the expression of heterosis; however, the potential role of microbes in heterosis has largely been ignored. Here, we show that heterosis of root biomass and other traits in maize is strongly dependent on the belowground microbial environment. We found that, in some cases, inbred lines perform as well by these criteria as their F1 offspring under sterile conditions but that heterosis can be restored by inoculation with a simple community of seven bacterial strains. We observed the same pattern for seedlings inoculated with autoclaved versus live soil slurries in a growth chamber and for plants grown in steamed or fumigated versus untreated soil in the field. In a different field site, however, soil steaming increased rather than decreased heterosis, indicating that the direction of the effect depends on community composition, environment, or both. Together, our results demonstrate an ecological phenomenon whereby soil microbes differentially impact the early growth of inbred and hybrid maize.}, number={30}, journal={Proceedings of the National Academy of Sciences}, publisher={Proceedings of the National Academy of Sciences}, author={Wagner, Maggie R. and Tang, Clara and Salvato, Fernanda and Clouse, Kayla M. and Bartlett, Alexandria and Vintila, Simina and Phillips, Laura and Sermons, Shannon and Hoffmann, Mark and Balint-Kurti, Peter J. and et al.}, year={2021}, month={Jul} }