Non-linear transcriptional responses suggest mechanisms of tributyltin toxicity in Triops longicaudatus

Tributyltin (TBT), a persistent biocide banned in 2008, continues to pose a significant threat to aquatic ecosystems. While its role as an endocrine disruptor is well established, its complete mechanistic cascade, particularly across environmentally relevant concentrations in non-target organisms, remains poorly characterised. Here, RNA-seq was used to characterise the transcriptional response of the non-model crustacean Triops longicaudatus following 90 min exposure to TBT at three nominal concentrations: 0.075 μg/L (96h-LC₅₀), 0.25 μg/L (a sub-acute concentration below the US EPA acute criterion of 0.46 μg/L), and 1.54 μg/L (a high environmental concentration). Rather than following a linear concentration-response pattern, TBT showed a quantitatively validated non-linear, tri-phasic mechanistic response characterised by three distinct signatures: (1) a U-shaped immunotoxic response at sub-lethal, acute (0.075 μg/L) and high concentrations (1.54 μg/L), with the downregulation of the complement cascade; (2) a unique middle concentration (0.25 μg/L) compensatory response associated with signatures of AMPK-mediated stress mitigation, pointing to potential systemic metabolic adaptation through crustacean hyperglycemic hormone signalling and upregulation of digestive pathways; and (3) high concentration (1.54 μg/L) multi-system failure, associated with DNA repair failure, chromatin degradation and suppression of cellular quality control mechanisms including autophagy and lysosomal acidification. This collapse can be mechanistically related to endocrine dysregulation: TBT's agonism of the RXR/PPAR heterodimer simultaneously coincides with changes in ecdysone signalling whilst attenuating juvenile hormone and cholesterol biosynthesis pathways. This study presents the first comprehensive transcriptomic evidence for concentration-dependent, non-monotonic mechanistic specificity in TBT toxicity, suggesting distinct modes of cellular failure that challenge conventional toxicological paradigms and carry significant implications for environmental risk assessment and regulatory frameworks.