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. 2023 Apr 26;299(6):104758. doi: 10.1016/j.jbc.2023.104758

Figure 2.

Figure 2

LysMD proteins localize to cell membranes.A and B, biochemical separation of whole cell lysates from adult flies into cytoplasmic (cyt) and membrane (mem) subfractions. Fractions were analyzed for protein components using Western blotting; Syx1A, plasma membrane marker; Cnx99A, ER membrane marker; Tubulin, cytoplasm marker. Mann-Whitney, ∗p < 0.05. Band intensity is plotted in arbitrary fluorescence units. A, WT flies; anti-Ima; n = 4 independent trials. B, the human ima homolog, hlysMD3, was expressed in WT Drosophila using the ubiquitous da-gal4 promoter driving the UAS-hlysMD3 transgene; anti-hLysMD3; n = 4 independent trials. C and D, distribution of Ima-GFP using immuno-EM. Arrows show stacked Golgi membranes in larval salivary gland (sg) cells. The scale bar represents 100 nm. E–H, colocalization of Ima-GFP with endomembrane markers using immunohistocytochemistry and confocal microscopy in larval salivary gland (sg) cells. Golgin-84, marker of Golgi rims; Hrs, early endosome. E and G, the scale bar represents 20 μm. F and H, the scale bar represents 5 μm. ER, endoplasmic reticulum; ima, immune active; LysMD, Lysin motif domain.