Fig. 9
From: Targeting cancer stem cell OXPHOS with tailored ruthenium complexes as a new anti-cancer strategy
Ru1 binds the mtDNA D-loop and inhibits transcription. A Representative IF confocal images of MitoGreen (mitochondrial mass), Ru1-TMR (red) and DAPI (Blue) staining in Ru1 (100µM)-treated Panc185 cells (24 h). White arrows indicate co-localization of Ru1-TMR and MitoGreen. Scale bar = 20 µm. B Amount of Ru1 molecule (µg/L) in untreated (UT), Ru1-treated (red) or Ru1-met-treated (green) Panc185 and PancA6L, determined by ICP at the indicated times post-treatment with 100µM of compounds. C Amount of Ru1 molecule (µg/L), determined by ICP, in gradient-purified mitochondria isolated from untreated (UT) and Ru1-treated Panc185 (100µM, 24 h). D Diagram of the mitochondrial genome, indicating the protein-coding genes (CI, yellow; CIII, blue, CIV, green; CV, red), and ribosomal RNA (rRNA)-coding genes (light blue). The D-loop region is magnified, and the predicted GQs and their positions are show in purple. TAS = termination associated sequence; HSP = heavy strand promoter; LSP = light strand promoter; OH = origin of replication – heavy; OL = origin of replication – light. Adapted from [52] and created in part with BioRender.com. E Agarose gel resolved D-loop or RNR2 PCR products amplified from 0.1 or 0.01ng of mtDNA pre-incubated for 1 h with diluent (Ctl) or 10µM of Ru1 or Ru1-met. F Mean fold-change ± SD in the expression of the indicated mtDNA-encoded genes as a function of increasing concentrations of Ru1 in Panc185 or PancA6L cells (48 h treatment). Values were normalized to ß-actin levels. * p < 0.05, ** p < 0.01, *** p < 0.001, as determined by one-way ANOVA with Dunnett post-test, compared to untreated (Ctl) set as 1.0