Elucidating Drought-Tolerance Mechanisms in Plant Roots through ¹H NMR Metabolomics in Parallel with MALDI-MS, and NanoSIMS Imaging Techniques

Abstract

As direct mediators between plants and soil, roots play an important role in metabolic responses to environmental stresses such as drought, yet these responses are vastly uncharacterized on a plant-specific level, especially for co-occurring species. Here, we aim to examine the effects of drought on root metabolic profiles and carbon allocation pathways of three tropical rainforest species by combining cutting-edge metabolomic and imaging technologies in an in situ position-specific 13 C-pyruvate root-labeling experiment. Further, washed (rhizosphere-depleted) and unwashed roots were examined to test the impact of microbial presence on root metabolic pathways. Drought had a species-specific impact on the metabolic profiles and spatial distribution in Piper sp. and Hibiscus rosa sinensis roots, signifying different defense mechanisms; Piper sp. enhanced root structural defense via recalcitrant compounds including lignin, while H. rosa sinensis enhanced biochemical defense via secretion of antioxidants and fatty acids. In contrast, Clitoria fairchildiana, a legume tree, was not influenced as much by drought but rather by rhizosphere presence where carbohydrate storage was enhanced, indicating a close association with symbiotic microbes. This study demonstrates how multiple techniques can be combined to identify how plants cope with drought through different drought-tolerance strategies and the consequences of such changes on below-ground organic matter composition.

Seeing plants as never before

“…Arabidopsis roots under altered phosphate conditions showed differences in the exudation of organic acids citrate and malate (Gomez‐Zepeda et al ., 2021). Finally, a comparative MSI (and NMR) study showed different drought‐tolerance strategies utilized within roots of three tropical plant species ( Piper sp., Hibiscus rosa sinensis , Clitoria fairchildiana ), with heterogeneous spatial patterns in a range of metabolites associated with structural, redox, and microbial defense (Honeker et al ., 2022).…”

Image to insight: exploring natural products through mass spectrometry imaging

“…The roots were labelled with position-specic (C1 or C2) 13 C-pyruvate, which allows to precisely track the fate of 13 C-pyruvate into downstream 13 C labelled metabolites. 90 Coupling MSI with SIL also provides the opportunity for spatial ux analysis and kinetics studies. MSI is limited in its ability to determine the production and turnover rates of intermediates as it only captures a snapshot of metabolism at the time of sampling.…”

“…The roots were labelled with position-specific (C1 or C2) 13 C-pyruvate, which allows to precisely track the fate of 13 C-pyruvate into downstream 13 C labelled metabolites. 90…”

Drought induces soil microbial stress responses and emissions of volatile organic compounds in an artificial tropical rainforest

“…Furthermore, plant production of lignin-like compounds may have increased during drought, for example, by Piper sp. 45 .…”

“…Position-speci c (C1 and C2) 13 C pyruvate labeling Soil was labeled with position-speci c (C1 or C2) 13 C-pyruvate (henceforth referred to as C1- 13 C-pyruvate and C2-13 C-pyruvate, respectively) to track C allocation into CO 2 , VOCs, and primary metabolites, as done previously in plants 45,62 . Three replicates each of C1-13 C-pyruvate or C2- 13 .…”