IHSS&IWA26 / BRNO / CZECHIA / 23–28 August 2026 BOOK OF ABSTRACTS 78 Research Frontiers Tuesday, 25 August 2026 / Hall B+C SL33 The Eustress Signature: Visualizing Proximal Signaling Events in Plant Roots Triggered by Electron Accepting and Donating Capacities Akshara Bathini1, Hiarhi Monda1, Richard T. Lamar1 1 Huma Inc., Gilbert, Arizona USA E-mail: akshara@huma.us Humic substances (HS) have been traditionally valued for their role in soil conditioning. However, current research has emphasized their bio stimulant effects on plants. The introduction of HS to plants initiates a sophisticated array of molecular signaling events that shape the development and physiology of plants. Particularly, the carboxyl, phenolic, and quinonoid functional groups within humics, potentially serve as primary reactive chemistries for interaction with plant membranes. Plants are known to perceive HS as environmental cues, triggering a response that is analogous to a mild stress or eustress (i.e. beneficial) response. Humic acid (HA) signaling is marked by activation of plasma membrane H+ ATPase activity, which establishes an electrochemical gradient required for nutrient uptake and cell expansion. While the electron accepting quinone moieties (pro-oxidants) appear to initiate a series of proximal events including membrane depolarization, Ca2+ flux from apoplast into cytoplasm and reactive oxygen species (ROS) production; the electron donating phenol groups (antioxidants) scavenge the ROS and modulate the stress intensity [1] [2]. Determining the spatiotemporal signatures of these proximal events and messengers, characterized by their unique amplitude, frequency, and duration, is required for identifying the roles they play in the HA mode of action. The current study hypothesizes that the ratio between electron accepting pro-oxidants and electron donating antioxidants determines the net bio stimulant effect of HA. To overcome the “black box” challenge of humic complexity, we employed a reductionist synthetic approach using purified quinones and flavonoids, which represented a range of redox potentials, to isolate the specific contributions of their respective chemistries. To test this, Arabidopsis thaliana was employed as the model organism for high resolution live cell fluorescence imaging. Real-time physiological responses were captured using transgenic R-GECO1 mutants. R-GECO1 is a red-shifted, intensity-based genetically encoded calcium indicator (GECI) that offers superior sensitivity for reporting [Ca2+] cyt oscillations at a cellular scale, allowing for the observation of transient calcium waves in response to redox stimuli. Sterilized R-GECO1 seeds were germinated on the edge of agar
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