STUDYING THE ROLE OF CALCIUM SIGNALING IN PLANT STRESS RESPONSES: INVESTIGATING NEW TARGETS FOR PLANT BIOTECHNOLOGY

Abstract

Plants perceive and decode environmental stress signals through transient elevations in cytosolic calcium, yet the molecular components that translate these signatures into adaptive responses remain incompletely understood. Here, we combined high-resolution in vivo calcium imaging in GCaMP6f‐expressing Arabidopsis, rice, and tomato with genome-wide discovery and functional validation of calcium sensors to identify new targets for biotechnological enhancement of stress tolerance. Under drought, salinity, and heat treatments, calcium transient amplitudes reached up to ΔF/F₀ = 0.55 and frequencies peaked at 3.5 peaks·min⁻¹ (Table 1), with distinct temporal profiles for each stress. Genome scans and expression profiling revealed that calmodulin-like proteins and CDPKs were among the most strongly induced sensors (fold-changes up to 3.2; Table 2). Yeast two-hybrid assays confirmed high-affinity interactions between CPK5 and MAPK3/SnRK2 (interaction scores ≥ 0.90; Table 3). CRISPR-Cas9 knockouts and overexpression lines of CML3, CPK5, ND-F2, and C2D1 demonstrated that overexpression enhanced drought and salinity tolerance—evidenced by 27 % lower stomatal conductance, 30 % reduced electrolyte leakage, and up to 35 % greater biomass (Tables 4–5; Figures 5–7). Network reconstruction highlighted that 60 % of key hubs are calcium sensors with higher centrality metrics than kinases (Table 6; Figures 8–9), underscoring their regulatory prominence. A positive correlation between sensor expression, calcium signal amplitude, and biomass gain establishes a direct link between calcium decoding and stress adaptation. Our integrative framework not only elucidates core calcium-mediated pathways but also nominates validated sensor and effector genes as prime candidates for CRISPR‐based or transgenic enhancement of crop resilience under increasingly erratic climates.