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. 2018 Nov 1;29(22):2766–2783. doi: 10.1091/mbc.E18-07-0434

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© 2018 Di Cara et al. “ASCB®,” “The American Society for Cell Biology®,” and “Molecular Biology of the Cell®” are registered trademarks of The American Society for Cell Biology.

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FIGURE 1: — Dysfunctional peroxisomes lead to structural disorganization and increased lysosomal/autophagic activity in the fly gut. (A) Pex5 depletion in the midgut causes increased lethality during fly development. Embryos were followed through development, and survival to larval, pupal, and adult stages were scored for Mex >Pex5-i flies and control flies; n = 70 eggs for each genotype in a single experiment. Values reported represent the averages of three independent experiments ± SD. Statistical significance was determined using Student’s t test; ***p < 0.001. (B) Representative electron microscopy images of midguts from control flies and Mex >Pex5-i flies uninfected (top panels) or infected with Ecc15 (bottom panels). nu, nucleus; vm, visceral muscle. Scale bar, 2 µm. (C) Number of vesicles containing electron dense material per region of interest (ROI) observed in midguts from control flies and Mex >Pex5-i flies (left graphic); profile height of midgut longitudinal sections (µm ± SD) from control flies and Mex >Pex5-i flies (right graphic). Flies were subjected to chronic oral infection with Ecc15. Twenty-five cells of each genotype were analyzed. Statistical significance was determined using Student’s t test; ***p < 0.001. (D) Immunogold labeling of epithelial cells with anti-Lamp1 antibodies. Panels a and b show higher magnifications of the vesicular structures seen in epithelial cells of infected Mex >Pex5-i flies presented in B. Panels c and d show immunogold labeling of round, electron-dense vesicles with anti-Lamp1 antibodies, confirming that these structures are autophagosomes or lysosomes. Scale bar, 500 nm. (E) Fluorescence detection of the autophagosome markers, Lysotracker (red) and ATG1 (green), in midguts from infected control flies and Mex >Pex5-i flies. Scale bar, 10 µm. (F) Quantification of ATG1, ATG3, and ATG8a mRNA transcript levels in midguts from Mex >Pex5-i flies relative to midguts from control flies. The values reported represent the average of four independent experiments. Statistical significance was determined using Student’s t test; *p < 0.05.