TPC2 regulates proteomic remodeling, bioenergetic phenotypes, and mitochondrial stress adaptation in melanoma.

Melanoma is a highly heterogeneous malignancy in which tumor behavior is shaped by complex signaling, trafficking, and metabolic programs. Two-pore channel 2 (TPC2), an endolysosomal ion channel, has been implicated in pigmentation, vesicular trafficking, and cancer-related cellular processes, but its role in melanoma remains incompletely understood. In this study, we investigated the impact of TPC2 loss in two melanoma models, the human cell line CHL-1 and the murine cell line B16, using previously validated CRISPR/Cas9-generated TPC2 knockout cells. Proteomic profiling, pathway enrichment analysis, extracellular flux measurements, and mitochondrial membrane potential assessment were used to define the molecular and functional consequences of TPC2 deletion. Proteomic analysis revealed marked but distinct changes in protein expression in CHL-1 and B16 cells, with clear separation between wild-type and TPC2 KO proteomic profiles in both models. Functional enrichment analysis showed that CHL-1 cells were preferentially associated with pathways related to metabolic regulation, morphogenesis, intracellular transport, extracellular matrix organization, and signaling networks including WNT and TGF-β, whereas B16 cells were enriched in immune/interferon-related pathways, protein homeostasis, intracellular trafficking, and stress-response programs. Bioenergetic profiling demonstrated that TPC2 KO altered cellular metabolism in a cell line-dependent manner. In CHL-1 cells, TPC2 loss reduced basal oxygen consumption rate (OCR) and the OCR/ extracellular acidification rate (ECAR) ratio, but enhanced oxidative adaptation under glucose-free conditions. In contrast, B16 TPC2 KO cells displayed increased OCR and ECAR under basal conditions, consistent with a more energetically active phenotype. Despite these differences in basal bioenergetics, TPC2 KO preserved mitochondrial membrane potential under stress in both cell lines, indicating enhanced mitochondrial stress resilience. Collectively, these findings identify TPC2 as a context-dependent regulator of melanoma cell physiology that influences proteomic remodeling, metabolic adaptation, and mitochondrial function. These results support further investigation of TPC2 as a potential therapeutic target in melanoma, while emphasizing that its biological effects depend on cellular context.