Fig. 7: Lysine maintains lysosomal function by antagonizing the effect of arginine on NO synthesis. | Nature Communications

Fig. 7: Lysine maintains lysosomal function by antagonizing the effect of arginine on NO synthesis.

From: Lysine-arginine imbalance overcomes therapeutic tolerance governed by the transcription factor E3-lysosome axis in glioblastoma

Fig. 7

A Lysosomal proteolytic activity was imaged and quantified in TGS04 cells after pretreatment with 25 mM arginine (Arg) for 16–24 h (n = 53 cells/group). Scale bars, 10 µm. B The representative images and quantification of galectin-3 punctation after treatment of cells with 50 μM LLOMe for 15 min (n = 3 independent samples). TGS04 cells expressing EGFP-Gal3 were pretreated with 25 mM arginine (Arg) for 16–24 h. Scale bars, 10 μm. C Number and relative number of spheres formed by TGS04 cells in the presence of different concentrations of LLOMe with or without arginine (Arg) (n = 3 technical replicates). D Detection of lysosomal proteolytic activity in the TGS04 cells treated with 50 µM Nor-NOHA for 16–24 h. E Sensitivity of TGS04 to TMZ with or without Nor-NOHA (n = 3 technical replicates). F The level of nitric oxide (NO) was detected in the TGS04 cells after treatment with 25 mM arginine (Arg) or lysine-depleted medium (w/o Lys) with flow cytometry. G Detection of lysosomal proteolytic activity in the TGS04 cells treated with 25 mM arginine (Arg) or 200 µM NOC-18 with flow cytometry. H Representative images and quantification of lysosomal proteolytic activity in the TGS04 cells pretreated with 4 mM L-NAME for 3 h before exposure to lysine-depleted medium (w/o Lys) for 16–24 h (Control medium n = 91; w/o Lys n = 58; L-NAME n = 72; Lys+L-NAME n = 83). Scale bars, 10 µm. I Cathepsin B activity was measured in the TGS04 cells after pretreatment with 4 mM L-NAME for 3 h, followed by exposure to a lysine-depleted medium for 16–24 h. J Illustration showing the mechanism, by which lysine-arginine imbalance induces nitric oxide production, leading to lysosomal dysfunction. Data are presented as the means ± SD. Statistic comparison (A) using unpaired two-tailed Student’s t-tests. Statistic comparison (B and H) using a one-way ANOVA. Statistic comparison (C and E) using a two-way ANOVA. ns, not significant, ****p < 0.0001. Experiments were repeated at least three times with similar results. Source data are provided with this paper.

Back to article page