Fig. 3: Increased cysteine acquisition causes excess cysteine stress, which increases sugar–CYS conjugates and impairs cancer cell proliferation.
From: Excess cysteine drives conjugate formation and impairs proliferation of NRF2-activated cancer cells
a, Relative abundance of intracellular sugar–CYS conjugates as measured by LC–MS metabolomics from SNU308 cells upon treatment with different medium concentrations of CYS2, with or without 0.5 μM erastin. Abundances are relative ion counts to cells cultured in 200 μM CYS2. n = 3 replicate wells per condition. 3GC: 50 μM CYS2 versus 100 μM CYS2 P = 0.0498, 50 μM CYS2 versus 200 CYS2 P < 0.0001, 50 μM CYS2 versus 400 CYS2 P < 0.0001, 50 μM CYS2 versus 800 μM CYS2 P < 0.0001, 800 μM CYS2 versus 800 μM CYS2 + 0.5 μM erastin P < 0.0001. 1DC: 50 μM CYS2 versus 100 μM CYS2 P = 0.4228, 50 μM CYS2 versus 200 μM CYS2 P = 0.0014, 50 μM CYS2 versus 400 CYS2 P < 0.0001, 50 μM CYS2 versus 800 μM CYS2 P < 0.0001, 800 μM CYS2 versus 800 μM CYS2 + 0.5 μM erastin P < 0.0001. b,c, Cell proliferation rates of the NRF2on cell line SNU308 (b) or NRF2off cell line CCLP1 (c) treated with different medium concentrations of CYS2, with vehicle control (dimethylsulfoxide (DMSO)) or with 0.5 μM erastin. n = 3 replicate wells per condition. SNU308: 50 μM CYS2 versus 100 μM CYS2 P = 0.6832, 50 μM CYS2 versus 200 μM CYS2 P = 0.1468, 50 μM CYS2 versus 400 μM CYS2 P = 0.0007, 50 μM CYS2 versus 800 μM CYS2 P < 0.0001, 800 μM CYS2 versus 800 μM CYS2 + 0.5 μM erastin P < 0.0003. d, Schematic depicting an xCT-independent route of CYS acquisition, where treatment with β-mercaptoethanol (BME) reacts with medium CYS2 to either reduce it to CYS or generate the mixed disulfide, CYS-BME. One or both compounds are imported through the neutral amino acid transporter family (ASCT), resulting in intracellular CYS delivery uncoupled from glutamate export. e, Cell proliferation rates of CCLP1 cells treated with different medium concentrations of BME, in medium containing either 200 or 800 μM CYS2. n = 3 replicate wells per condition, 0 μM BME P = 0.8164, 100 μM BME P = 0.4121, 200 μM BME P = 0.0001, 400 μM BME P = 0.0001. f, Abundance of intracellular sugar–CYS conjugates as measured by LC–MS metabolomics from CCLP1 cells upon treatment with different medium concentrations of BME, in medium containing either 200 or 800 μM CYS2 for 24 h. Abundances are relative ion counts to cells cultured in 200 μM CYS2 with 0 μM BME for each metabolite. n = 3 replicate wells per condition. 3GC: 0 μM BME P = 0.9421, 100 μM BME P = 0.0396, 200 μM BME P = 0.0001, 400 μM BME P = 0.0001. 1DC: 0 μM BME P = 0.8724, 100 μM BME P = 0.0013, 200 μM BME P < 0.0001, 400 μM BME P < 0.0001. Error bars show s.e.m. Statistical significance was assessed using one-way ANOVA with Sidak’s correction for multiple comparisons (a–c) or by two-way ANOVA with Sidak’s correction for multiple comparisons (e,f). *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001. Panel d created in BioRender; Sullivan, L. https://biorender.com/wtofmpq (2026).
