QSOX1 inhibits autophagic flux.

<p>(<b>A</b>) MCF-7 C, QSOX1S-1, QSOX1S-2 and (<b>B</b>) MDA-MB-231 shC, shQSOX1-2, shQSOX1-1 cells were cultured in complete medium with or without 100 nM bafilomycin A1 for 8 h. Cells were then lysed and total proteins (40 µg) were separated on 12.5% SDS-PAGE followed by immunoblotting with anti-p62, anti-LC3 and anti-actin antibodies. p62 and LC3-II levels were quantified using the Image Lab software. (<b>C and D</b>) The autophagic flux, observed in A and B respectively, was determined as the ratio of LC3-II protein levels in the presence of bafilomycin A1 versus the levels in the absence of bafilomycin A1. Data are means ± S.D. of three independent experiments. *P<0.05 compared to the control. (<b>E</b>) MCF-7 C, QSOX1S-1, QSOX1S-2 and (<b>F</b>) MDA-MB-231 shC, shQSOX1-1, shQSOX1-2 cells were transfected with the pGFP-LC3 vector. 24 h after transfection, cells were incubated with or without 100 nM bafilomycin A1 for 8 h. GFP-LC3 puncta were then analyzed by confocal microscopy. Each picture is representative of a typical cell staining observed in 10 fields chosen at random. Scale bar represents 20 µm. (<b>G and H</b>) GFP-LC3 puncta, observed in E and F respectively, were counted using the ImageJ software. For each group, 20 cells were randomly selected. Data are means ± S.D. of three independent experiments. *P<0.05 compared to the control.