Fig. 4: PGAM5 exerts anti-apoptotic function in HCC cells via Bcl-xL mediated apoptosis inhibition signaling pathway.

a Depletion of PGAM5 expression substantially downregulated Bcl-xL expression in HCC cells detected by western blotting. b Co-immunopercipitated with PGAM5 antibody incubated shows that PGAM5 interacts with Bcl-xL in both 7402 and HepG2 cells. c Representative images for immune-fluorescent staining of PGAM5 and mitochondria. Nucleus was stained with DAPI (bule). Yellow dots indicate PGAM5 (green) on the mitochondria (red). d Immuno-staining images indicate that PGAM5 (red) and Bcl-xL (green) were co-localization (orange). e After 48 h treatment with 5-Fu at indicated dose, mitotracker pre-stained cells stained with Bcl-xL antibody, BAX antibody, Cyt.C antibody and PGAM-5 antibody. Representative images show that inhibition of PGAM5 decrease Bcl-xL (green) exhibition in mitochondria (upper), whereas enhances the translocation of BAX (green) from cytosol to the mitochondria (yellow dots indicated the co-localization of BAX and mitochondria, middle). In addition, silence of PGAM5 promoted the translocation of Cyt.C (green) from mitochondrial to cytosol (Yellow dots indicated the co-localization of Cyt.C and mitochondria, lower). Scale bars: 20 um. f Collected protein extraction respectively mitochondrial and cytosolic. Western blotting reveals that depletion of PGAM5 decrease Bcl-xL expression on mitochondrial and inhibits BAX translocation from cytoplasm to mitochondria. Proteins from the cytosolic fraction were used to determine Cyt.C release. After replenishment of PGAM5 in PGAM5-silenced 7402 and HepG2 cells, all of the altered phenotypes were recoved (a, e, and f). g Proposed schematic representation of a molecular mechanism of PGAM5, Bcl-xL, BAX, and Cyt.C in HCC chemoresistance to 5-Fu