Fig. 3

MCU-induced mitochondrial Ca²⁺ uptake promotes mitochondrial biogenesis in CRC cells. a Relative mitochondrial content (µm³ mitochondria per µm³ cell) was determined based on a mitochondrial three-dimensional model of confocal microscope images in LS174T cells, treated as indicated. MitoTracker (Green) was used to label mitochondria. b Relative mtDNA copy number was determined by RT-qPCR in LS174T cells, treated as indicated. c Western blotting analysis to measure the expression of oxidative phosphorylation related proteins in LS174 cells treated as indicated. d Mitochondrial ATP levels were determined using the ATP Determination Kit in LS174T cells, treated as indicated. e Correlation between relative mtDNA copy number and relative MCU expression in CRC tissue. The mtDNA copy number was divided into four levels based on the quartering of RT-qPCR results. As the quartiles are 3.21 (upper quartile), 4.12 (median) and 5.22 (lower quartile), RT-qPCR value <3.21, 3.21–4.12, 4.12–5.22, and >5.22 were defined as (+), (++), (+++), and (++++), respectively. f Correlation between relative cytochrome c oxidase subunit 4 expression and relative MCU expression in CRC tissue. g Correlation between relative mitochondrial content and relative MCU expression in CRC tissue. Mitochondrial content was divided into four groups based on the quartering of the relative mitochondrial content levels. As the quartiles are 0.15 (upper quartile), 0.21 (median) and 0.25 (lower quartile), relative mitochondrial content level <0.15, 0.15–0.21, 0.21–0.25, and >0.25 were defined as (+), (++), (+++), and(++++), respectively. *P < 0.05; **P < 0.01. MCU mitochondrial calcium uniporter, CRC colorectal cancer, mtDNA mitochondrial DNA, RT-qPCR reverse transcription-quantitative PCR, Ca²⁺ calcium