Journal: Cell Death Discovery

Article Title: PPA1 promotes oxidative phosphorylation and malignant progression of colorectal cancer under glucose restriction via AMPK/ULK1/FUNDC1-mediated mitophagy

doi: 10.1038/s41420-025-02816-y

Figure Lengend Snippet: Under glucose-restricted conditions, ( A , B ) The expression levels of mitophagy-related proteins and phosphorylation sites of HCT8(A) and HCT116(B) cells were detected by Western blotting; ( C , D ) Western blotting detecting LC3II and LC3I expression levels in cytoplasmic and mitochondrial fractions of HCT8 (C) and HCT116 ( D ) cells; ( E ) Transmission electron microscopy images showing mitophagy and mitochondria in CRC cells after PPA1 knockdown (red arrows indicate mitophagosomes or mitochondria; left: 8000×, right: 25000× magnification); ( F ) Confocal microscopy imaging of LC3 fluorescence and its co-localization with mitochondria (left: 600×, right: 2400× magnification). G Flow cytometry detection of mitochondrial membrane potential in CRC cells (Mean ± SD, n = 3). H Flow cytometry analysis of apoptosis rates in HCT8 and HCT116 cells (Mean ± SD, n = 3). All data are presented as Mean ± SD. ** p < 0.01, *** p < 0.001, **** p < 0.0001.

Article Snippet: The ULK1 overexpression plasmids and phosphorylation site mutant plasmids (S467A, S556A, and S638A plasmids) were obtained from Sangon Biotech (Shanghai, China), with detailed sequences provided in Table .

Techniques: Expressing, Phospho-proteomics, Western Blot, Transmission Assay, Electron Microscopy, Knockdown, Confocal Microscopy, Imaging, Fluorescence, Flow Cytometry, Membrane

Journal: Cell Death Discovery

Article Title: PPA1 promotes oxidative phosphorylation and malignant progression of colorectal cancer under glucose restriction via AMPK/ULK1/FUNDC1-mediated mitophagy

doi: 10.1038/s41420-025-02816-y

Figure Lengend Snippet: Under glucose-restricted conditions, ( A ) Western blotting analysis of mitophagy-related proteins and phosphorylation levels in CRC cells with ULK1 mutations at Ser467 or Ser556 or Ser638 site; ( B , C ) Western blotting detection of LC3II and LC3I expression in cytoplasmic and mitochondrial fractions of HCT8 (B) and HCT116 ( C ) cells after mutating Ser467 or Ser556 or Ser638 site of ULK1; ( D – I ) Quantification of protein and phosphorylation site expression levels in HCT8 ( D − F ) and HCT116 ( G − I ) cells (Mean ± SD, n = 3); ( J ) Confocal microscopy imaging and quantitative analysis of LC3 fluorescence and its co-localization with mitochondria (left: 600×, right: 2400 × magnification); ( K ) Confocal microscopy analysis of mitochondrial membrane potential in CRC cells (Mean ± SD, n = 3, 200× magnification). All data are presented as Mean ± SD. * p < 0.05, ** p < 0.01, *** p < 0.001, ns not significant.

Article Snippet: The ULK1 overexpression plasmids and phosphorylation site mutant plasmids (S467A, S556A, and S638A plasmids) were obtained from Sangon Biotech (Shanghai, China), with detailed sequences provided in Table .

Techniques: Western Blot, Phospho-proteomics, Expressing, Confocal Microscopy, Imaging, Fluorescence, Membrane

Journal: Cell Death Discovery

Article Title: PPA1 promotes oxidative phosphorylation and malignant progression of colorectal cancer under glucose restriction via AMPK/ULK1/FUNDC1-mediated mitophagy

doi: 10.1038/s41420-025-02816-y

Figure Lengend Snippet: Under glucose-restricted conditions, ( A ) Western blotting analysis of mitophagy-related proteins and phosphorylation levels in CRC cells treated with the ULK1 agonist LYN-1604 (1 μM); ( B , C ) Western blotting detection of LC3II and LC3I expression in cytoplasmic and mitochondrial fractions of HCT8 ( B ) and HCT116 ( C ) cells treated with LYN-1604 (1 μM); ( D – I ) Quantification of protein and phosphorylation site expression levels in HCT8 (D-F) and HCT116 ( G − I ) cells (Mean ± SD, n = 3); ( J ) Confocal microscopy imaging of LC3 fluorescence and its co-localization with mitochondria in cells treated with or without LYN-1604 (1 μM) (left: 600×, right: 2400 × magnification); ( K ) Impact of LYN-1604 (1 μM) on mitochondrial OXPHOS activity in CRC cells(oligomycin (1 μM), FCCP (1.5 μM), rotenone (0.5 μM), and antimycin A (0.5 μM)); ( L , M ) Quantification of basal OCR(L) and ATP production(M) (Mean ± SD, n = 3); ( N , O ) Flow cytometry analysis of mitochondrial ROS levels in HCT8 (N) and HCT116 ( O ) cells treated with LYN-1604 (Mean ± SD, n = 3). All data are presented as Mean ± SD. * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001.

Article Snippet: The ULK1 overexpression plasmids and phosphorylation site mutant plasmids (S467A, S556A, and S638A plasmids) were obtained from Sangon Biotech (Shanghai, China), with detailed sequences provided in Table .

Techniques: Western Blot, Phospho-proteomics, Expressing, Confocal Microscopy, Imaging, Fluorescence, Activity Assay, Flow Cytometry

Journal: Cell Death Discovery

Article Title: PPA1 promotes oxidative phosphorylation and malignant progression of colorectal cancer under glucose restriction via AMPK/ULK1/FUNDC1-mediated mitophagy

doi: 10.1038/s41420-025-02816-y

Figure Lengend Snippet: Under glucose-restricted conditions, ( A , B ) The protein levels and phosphorylation status of both mTOR and AMPK after PPA1 knockdown; ( C , D ) The mitophagy-related proteins and phosphorylation levels in CRC cells treated with or without the AMPK agonist GSK621 (30 μM); ( E ) Confocal microscopy imaging of LC3 fluorescence and its co-localization with mitochondria in cells treated with or without GSK621 (30 μM) (left: 600×, right: 2400 × magnification). All data are presented as Mean ± SD. * p < 0.05, ** p < 0.01, ns not significant.

Article Snippet: The ULK1 overexpression plasmids and phosphorylation site mutant plasmids (S467A, S556A, and S638A plasmids) were obtained from Sangon Biotech (Shanghai, China), with detailed sequences provided in Table .

Techniques: Phospho-proteomics, Knockdown, Confocal Microscopy, Imaging, Fluorescence