Characterization of molecular and physiological responses to iron excess in sugarcane genotypes

Iron (Fe) is an essential micronutrient; however, in acidic or waterlogged soils its bioavailability increases and may reach toxic concentrations, triggering damage to plants, a problem that remains poorly explored in sugarcane. Excess Fe catalyses the formation of reactive oxygen species through Fenton reactions, compromising the photosynthetic apparatus and reducing crop productivity. Although the physiological effects of Fe toxicity are known, the molecular mechanisms that distinguish the contrasting responses among sugarcane genotypes remain poorly understood. Therefore, this study characterised the leaf proteomic responses to Fe excess in two sugarcane genotypes with contrasting morphophysiological behaviour, CTC9001 and RB855536, in order to elucidate the molecular basis of these responses. Plants were grown in a sand-based hydroponic system, in a randomised complete block design with eight biological replicates per genotype/treatment for the morphophysiological analyses and three biological replicates per genotype/treatment for the proteomic analyses, and were supplied for 42 days with Hoagland and Arnon nutrient solution containing 70 µmol L⁻¹ Fe-EDTA (control) or 500 µmol L⁻¹ (Fe excess). Morphophysiological analyses revealed distinct responses between the genotypes regarding growth and the functioning of the photosynthetic apparatus. The maximum photochemical efficiency of photosystem II (Fv/Fm) and the reaction centres were affected in a genotype-dependent manner, with an early photochemical response in RB855536 and a later one in CTC9001. Proteomic analysis identified 83 differentially accumulated proteins in CTC9001 and 201 in RB855536, of which 28 were shared. Fe excess promoted a remodelling of the chloroplast- and plastid-associated proteome, mainly involving pathways related to carbon and energy metabolism, including carbon fixation, the Calvin cycle, glycolysis and the tricarboxylic acid cycle. RB855536 exhibited a broader response, with enrichment of carbon metabolism, amino acid biosynthesis, glyoxylate and dicarboxylate metabolism and chaperone-mediated protein refolding, including HSP70, HSP60 and ClpB, indicating an intense metabolic adjustment. CTC9001 concentrated its response on proteins related to the generation of precursor metabolites and energy, carbon fixation and chlorophyll biosynthesis. In both genotypes, redox metabolism proteins such as peroxidases, catalase, peroxiredoxin and thioredoxin were differentially modulated. In RB855536, protein-protein interaction networks associated with proteostasis and with the regeneration of reducing power (NADPH) were observed, whereas in CTC9001 a reduction in the interactions between energy-respiratory metabolism and translation was observed. The results reveal molecular mechanisms associated with tolerance to Fe excess and identify candidate proteins that may support genotype selection strategies and accelerate the development of sugarcane cultivars adapted to soils with Fe toxicity.