Metaproteomics reveals insights into microbial structure, interactions, and dynamic regulation in defined communities as they respond to environmental disturbance

Him K Shrestha,Dale A Pelletier,Manasa R Appidi,Robert L Hettich,Jia Wang,Leah Burdick,Dana L Carper,Manuel I Villalobos Solis,Paul E Abraham,Mitchel J Doktycz

doi:10.1186/s12866-021-02370-4

Abstract

BackgroundMicrobe-microbe interactions between members of the plant rhizosphere are important but remain poorly understood. A more comprehensive understanding of the molecular mechanisms used by microbes to cooperate, compete, and persist has been challenging because of the complexity of natural ecosystems and the limited control over environmental factors. One strategy to address this challenge relies on studying complexity in a progressive manner, by first building a detailed understanding of relatively simple subsets of the community and then achieving high predictive power through combining different building blocks (e.g., hosts, community members) for different environments. Herein, we coupled this reductionist approach with high-resolution mass spectrometry-based metaproteomics to study molecular mechanisms driving community assembly, adaptation, and functionality for a defined community of ten taxonomically diverse bacterial members of Populus deltoides rhizosphere co-cultured either in a complex or defined medium.ResultsMetaproteomics showed this defined community assembled into distinct microbiomes based on growth media that eventually exhibit composition and functional stability over time. The community grown in two different media showed variation in composition, yet both were dominated by only a few microbial strains. Proteome-wide interrogation provided detailed insights into the functional behavior of each dominant member as they adjust to changing community compositions and environments. The emergence and persistence of select microbes in these communities were driven by specialization in strategies including motility, antibiotic production, altered metabolism, and dormancy. Protein-level interrogation identified post-translational modifications that provided additional insights into regulatory mechanisms influencing microbial adaptation in the changing environments.ConclusionsThis study provides high-resolution proteome-level insights into our understanding of microbe-microbe interactions and highlights specialized biological processes carried out by specific members of assembled microbiomes to compete and persist in changing environmental conditions. Emergent properties observed in these lower complexity communities can then be re-evaluated as more complex systems are studied and, when a particular property becomes less relevant, higher-order interactions can be identified.

Highlights

Microbe-microbe interactions between members of the plant rhizosphere are important but remain poorly understood
The 10-member defined community (DefCom) co-culture was grown in liquid defined media (MOPS + 0.2 % glucose) and complex media (R2A) subjected to growth/dilution cycles every two days until the community reached a stable state (Fig. 1). These two media were selected to determine the effect of environmental filtering based on a single carbon source with limited essential nutrients and a more complex media with increased carbon resources and nutrients designed for long incubation periods
Proteome-wide as well as protein-centric modifications identified in this study provide the proof of concept that optimized bioinformatics pipelines and high-resolution mass spectrometry affords the ability to broadly characterize PostTranslational Modifications (PTM) in a biological system and provides a level of sensitivity capable of revealing regulatory mechanisms influencing the activity of single proteins and we expect to continue mining this wealth of information to find novel and/or less-studied PTM regulatory mechanisms

Summary

Introduction

Microbe-microbe interactions between members of the plant rhizosphere are important but remain poorly understood. One strategy to address this challenge relies on studying complexity in a progressive manner, by first building a detailed understanding of relatively simple subsets of the community and achieving high predictive power through combining different building blocks (e.g., hosts, community members) for different environments We coupled this reductionist approach with high-resolution mass spectrometrybased metaproteomics to study molecular mechanisms driving community assembly, adaptation, and functionality for a defined community of ten taxonomically diverse bacterial members of Populus deltoides rhizosphere co-cultured either in a complex or defined medium. Constraint based genome-scale metabolic models are used in predicting the extent of resource competition and microbial metabolic interactions, which is facilitated by flux balance analysis (FBA) [7] Besides these approaches, data-driven inferences of microbial interaction networks, such as ecology guided models that predict the metabolite cross-feeding interactions based on metagenomic and metabolomic measurements, are valuable in understanding the community dynamics [8]. Despite the usefulness of these tools, the key to fully understand the dynamics of microbial communities and the underlying principles within them, is to integrate predictive models with experimental data such as those obtained from experiments using defined microbial consortia [5,6,7,8]

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