Plant epidermal cells serve as a barrier between them and their surroundings as well as key interaction interfaces between plants and beneficial microbes. Epidermal plastids of leaves and the small plastids of roots, collectively called sensory plastids, play a vital role during microbial interactions. These have largely been studied during responses to pathogens, but we hypothesize that their reprogramming is necessary for responses to beneficial microbes. Indeed, sensory plastid proteins that we have studied are dynamically regulated and crucial for adaptations to both beneficial and pathogenic bacteria. Recent research has shown a relationship between the intracellular dynamics of sensory plastids and plant signaling, suggesting that these processes may be coordinated to increase adaptation to microbes. These processes include the focal accumulation of sensory plastids at the microbial interface, stromule formation (extensions/projections), inter-organelle communication, and others. Thus, the identification of undiscovered microbial interaction-related events and elements in the plant sensory plastids is crucial for gaining a thorough understanding of adaptive responses to microbes. We are deploying a recently developed fast and accurate method for isolating intact sensory plastids from leaves and roots using affinity purification. By using this method, we will minimize degradation/loss of the components we wish to measure and also, for leaves, separate the sensory plastids of interest from the large mesophyll plastids that dominate conventional preparations and can obscure signals from the sensory plastids. Our aims are to investigate the metabolite, lipid and membrane proteome reprogramming of the sensory plastids during beneficial microbial interactions using mass spectrometry. In a parallel set of experiments, we aim to understand the dynamics, also using mass spectrometry, of key plastid membrane protein complexes needed for optimal responses to beneficial bacteria. Metabolite and lipid changes will give insight into potential signal molecules (or their precursors) that plants deploy, while membrane protein changes, including complexes with proteins that we know are important for microbial responses, will give leads for understanding how signals are mobilized and contacts with other organelles are made. We will start with analyzing Arabidopsis plastids and extend our work to the oil seed energy crop Camelina sativa. Changes in plastid composition and an understanding of the pathways modulated (inferred from the omics analyses) will be related to results from our DOE collaborative project in which we will be monitoring the root environment over time using genetically encoded biosensors in the beneficial microbes.