Biphasic Temporal Remodeling Of The Proteome In A Polyglutamine-Expanded Huntingtin In Vitro Aggregation Cell Model: From Early Rna-Regulatory Compensation To Selective Mitochondrial Energy Failure
Huntington disease (HD) is caused by a polyglutamine expanded huntingtin protein that exerts progressive cellular toxicity. However, the temporal sequence of pathogenic, particularly early and reversible versus late and irreversible events remain incompletely defined, despite their distinct therapeutic implications. To delineate this trajectory, we profiled the proteome of a huntingtin expressing cell model at early (72 h) and late (144 h) stages. Rather than a linear progression, pathogenicity unfolded in two discrete phases. At the early stage, cells exhibited a broad activation of RNA processing, splicing, and protein synthesis machinery, consistent with an adaptive response aimed at preserving gene expression fidelity under stress. By the late stage, this compensatory program had collapsed, giving rise to a dominant failure in mitochondrial energy metabolism. Notably, 85% of proteins altered at both time points reversed direction of change between stages, indicating that mutant huntingtin reprograms cellular function wholesale rather than amplifying a fixed set of perturbations. Detailed analysis of mitochondrial respiratory complexes revealed that terminal ATP generating components (cytochrome c oxidase and ATP synthase) were severely affected, whereas upstream electron transport elements were retained or upregulated. Leveraging this proteomic map, we applied an AI assisted, direction aware drug repurposing strategy. Of 1,712 differentially expressed proteins, 498 were druggable, and 89 mapped to approved agents with mechanisms concordant with the required correction. These included Complex I targeted agents (metformin, ME 344) and mitochondria directed therapeutics (SS 31, MitoQ), several of which have previously been evaluated in HD. Collectively, these findings define a biphasic course of huntingtin toxicity and highlight an early therapeutic window in which intervention is most likely to be applied, prior to irreversible deterioration of mitochondrial respiratory function.