Fig. 3: Gonococcal OMVs deliver PorB to mitochondria to induce mitophagy.
From: Gonococcal OMV-delivered PorB induces epithelial cell mitophagy
a TEM of HeLa cells expressing gonococcal PorB from strain ATCC 49226 show mitochondrial capture in mitophagy-like structures. Data are mean ± s.d.; n = 21 cells, two-tailed Mann–Whitney test, P = 2 × 10−10. b Western blots showing degradation of mitochondrial proteins TOM20 and TIM23 in HeLa cells expressing gonococcal PorB from strains ATCC 49226 and ZJXSH86, but not for PorB from Neisseria mucosa. c Western blots showing that gonococcal OMVs expressing PorB from N. mucosa lost the ability to induce degradation of TOM20 and TIM23 in HeLa cells. d Gonococcal OMVs expressing PorB from N. mucosa lost the ability to dissipate the mitochondrial membrane potential (MMP) in HeLa cells. Scale bar, 5 μm. Data are mean ± s.d.; n = 55 cells from 4 independent experiments, Kruskal–Wallis with posthoc Dunn test (Mock-N.g. PorB: P = 8 × 10−13; N.g. PorB-N.m. PorB: P < 10−15). e Flow cytometry analysis of TMRM fluorescence intensity in HeLa cells stimulated with gonococcal OMVs expressing gonococcal PorB or PorB from N. mucosa. f Gonococcal PorB structure (pdb entry 4AUI) and sequence of PorB from strain ATCC 49226. Lysines are indicated in blue, or in magenta when located within the PorB channel and associated with ATP (orange) binding. g Western blots showing reduced degradation of TOM20 and TIM23 in HeLa cells expressing PorB K117Q. h Reduced HSP60 and LC3 colocalization in HeLa cells expressing PorB K117Q compared with PorB WT. Scale bar, 5 μm. Data are mean ± s.d.; n = 51 cells, Kruskal–Wallis with posthoc Dunn test (Empty-WT PorB: P < 10−15; Empty-PorB K117Q: P = 2 × 10−9; WT PorB-PorB K117Q: P = 1 × 10−5). i Reduced HSP60 and LAMP1 colocalization in HeLa cells expressing PorB K117Q compared with PorB WT. Scale bar, 5 μm. Data are mean ± s.d.; n = 51 cells, Kruskal–Wallis with posthoc Dunn test (Empty-WT PorB: P < 10−15; Empty-PorB K117Q: P = 9 × 10−8). j Western blots of HeLa 4KO cells showing p62 expression restores PorB K117Q-dependent degradation of TOM20 and TIM23, while expression of OPTN or NDP52 restores PorB K5-dependent degradation of TOM20 and TIM23. k Western blots showing impaired degradation of TOM20 and TIM23 in HeLa cells expressing PorB K117Q, K5 or Knull. l Western blots showing that knock-down of PINK1 in HeLa cells inhibits PorB K5-dependent degradation of TOM20 and TIM23, but not for PorB K117Q. Cells in a, h, i are from 3 independent experiments. Western blots in b, c, g, j, k, l are representative of 3 independent experiments. Source data are provided as a Source Data file.
