Pre-symptomatic proteomic and metabolomic profiling identifies compensated ER-redox-metabolic adaptation and early nuclear vulnerability in neuronal ERO1L toxicity

Aging progressively challenges neuronal proteostasis, redox homeostasis, and metabolism, yet the molecular changes that precede functional decline remain poorly understood. Endoplasmic reticulum oxidoreductin 1 (ERO1), a key regulator of oxidative protein folding, links endoplasmic reticulum (ER) proteostasis with cellular redox balance and is elevated in aging and neurodegenerative contexts. Here, we investigated how neuronal ERO1L elevation reshapes cellular homeostasis before overt dysfunction in Drosophila melanogaster. Endogenous ERO1L expression increased with age, and neuronal ERO1L elevation shortened lifespan and caused progressive locomotor decline. This effect was strongly cell-type dependent, as ERO1L elevation in glia, muscle, or fat body did not produce a comparable survival phenotype. At day 5 post-eclosion, locomotor performance remained preserved and major brain reactive-oxygen-species (ROS) accumulation was not yet detectable, defining a pre-symptomatic stage. Multi-omic profiling at this stage revealed selective remodelling of ER proteostasis, redox defence, and mitochondrial-energy pathways, together with changes in central-carbon, nitrogen, and purine metabolism. In contrast to these broadly adaptive responses, chromatin- and RNA-homeostasis-associated proteins were selectively reduced, accompanied by decreased HP1, dFmr1, and Piwi expression and increased transposable-element transcripts. Thus, neuronal ERO1L elevation establishes a pre-symptomatic state in which proteostatic and metabolic adaptation coexists with early vulnerability of nuclear and RNA-homeostasis pathways, preceding overt oxidative stress and behavioural decline. These findings provide an in vivo framework to investigate how age-associated ERO1L elevation may progressively reduce neuronal resilience during brain aging.