Fig. 4: Mitochondrial Drp1 induces HK2 dephosphorylation through recruiting LRRK2 after hypoxia.

A Human proteome microarray showing potential interacting proteins with Biotin-tagged Drp1 protein. Red arrow indicated the positive control (Biotin-BSA), yellow arrow indicated the negative control (BSA protein), and white arrow indicated LRRK2, our interested potential interacting proteins with Biotin-tagged Drp1 protein. B Co-IP results for the combination of Drp1 and LRRK2 in total, cytoplasmic and mitochondrial fractions of VSMCs after hypoxia. (n = 3 samples/group). C Representative immunofluorescence images showing the co-location of Drp1 (red), LRRK2 (green) and mitochondria (white) in VSMCs after hypoxia. (bar, 5 μm). D Molecular docking of Drp1 to LRRK2 in hypoxia conditions determined by GRAMM-X Protein-Protein Docking program. The blue dotted indicated the stable hydrogen bond between Drp1 Thr595 and LRRK2 Gly2019. E Co-IP results for the combination of Drp1 and LRRK2 in hypoxia-induced VSMCs after Drp1 T595A mutation. F The phosphorylation of HK2-Thr 473 and HK2 expression in total, cytoplasmic and mitochondrial fractions of hypoxia-induced VSMCs after Drp1 T595A mutation. β-actin, Tubulin and COX IV were used as interior references of total, cytoplasmic and mitochondrial fractions. G Representative confocal images of mPTP opening in hypoxia-induced VSMCs after Drp1 T595A mutation. (bar, 100 μm). a: P < 0.05 compared with Normal group.