Regulatory Networks Underlying Plant Nutrient Status

Plants require 14 mineral nutrients for growth and development. Nutrient imbalances remain major constraints on agricultural productivity. Conventional approaches for diagnosing plant nutritional status often fail to identify the primary limiting element, particularly in acidic soils where multiple deficiencies and toxicities co-occur. Non-invasive tools estimate nutrient status indirectly and are affected by other stresses, leading to misdiagnosis, excessive fertilization, and environmental harm. Thus, precise and element-specific diagnostic strategies are required for sustainable crop production. At the molecular level, nutrient homeostasis is tightly regulated by signaling networks. In this context, inositol pyrophosphates (PP-InsPs) are signaling molecules regulating phosphate signaling and diverse processes, while pyrophosphate (PPi), a by-product of biosynthetic reactions, plays key roles in cellular metabolism. Despite their importance, the link between PP-InsPs and PPi homeostasis has remained poorly understood. In this thesis, integrative ionomic, transcriptomic, and qPCR approaches were used to identify molecular sentinels that respond specifically to distinct nutrient imbalance conditions in barley. In addition, co-regulated genes were identified, revealing coordinated nutrient interactions. A set of reference genes with stable expression across treatments is also defined, providing a valuable resource for future qPCR studies. These findings demonstrate that molecular sentinels have the potential to overcome key limitations of conventional nutrient analyses and enable early, precise diagnosis of nutritional disorders before visible symptoms or yield losses occur. This thesis also uncovers a previously unrecognized role of PP-InsPs in regulating PPi homeostasis in plants. Genetic and biochemical studies in Arabidopsis show that canonical PPi-hydrolyzing systems are unlikely to be involved in InsP/PP-InsP metabolism. In contrast, disruption of InsP/PP-InsP biosynthetic enzymes causes marked PPi accumulation accompanied by extensive remodeling of InsP/PP-InsP profiles, adenylate pools, and the plant nutriome. Complementation restores PPi levels, nutrient balance, and shoot growth. Additionally, high PPi exacerbates shoot growth defects, and restoration of PPi levels with the kinase domain of VIH2 suggests that the kinase activity of VIH2 is important for PPi control. Biochemical analyses showed that PPi is not a phosphoryl donor for PP-InsP synthesis, but activates InsP/PP-InsP kinases, and that PP-InsPs stimulate ScIPP1. In addition, sulfur homeostasis is identified as a previously unrecognized macronutrient regulated by PP-InsPs. In conclusion, this thesis establishes molecular sentinels as a potentially precise diagnostic indicators of plant nutritional status and positions PP-InsPs as central integrators of nutrient signaling, PPi metabolism, and metabolic homeostasis, offering new avenues to improve nutrient use efficiency and support sustainable agriculture.