Phosphate availability modulates elemental homeostasis in rainbow trout hepatocytes: compositional ionomics illuminates system-wide adjustments

Phosphorus (P) is central to biology, yet it remains unclear whether P limitation acts as an isolated nutrient constraint or as a perturbation to a dynamical elemental network. We tested this using the rainbow trout liver cell line RTL-W1 by manipulating phosphorus supply (0, 10% and 100% of normal supply; P0, P10, P100) and quantifying proliferation, protein content, metabolic activity, membrane integrity, and multielement composition. Phosphorus supply significantly altered cell proliferation and protein accumulation, with higher P supporting greater growth. Metabolic activity was affected by P supply, whereas membrane integrity remained largely stable, indicating altered allocation rather than generalized cellular damage. Compositional ionomic analysis revealed that phosphorus perturbation restructured the multielement network. At Day 3, treatment effects were strongest for P, K, and S. By Day 6, additional elements, including Sr and Mn, exhibited coordinated shifts relative to the P100 reference. Compositional data analysis showed that elemental imbalances shifted through time, consistent with flux rebalancing in an open system rather than static homeostasis. If phosphorus limitation were mechanistically independent, multielement composition would remain stable aside from P itself. Instead, phosphorus perturbation induced coordinated shifts across the ionome, consistent with rebalancing in open material systems.