Figure 2.
Juxta-Golgi localization of Rop, Vps20, lysosomal markers, and endosomal Rab-GTPases. (A) Confocal image of L3 fat body (feeding stage) showing localization of Rop-GFP (green) and cis-Golgi marker Grasp65-RFP (magenta), both driven by Cg-GAL4. Magnified insets are shown below for both markers and for Rop alone. (B) Localization in fat body of Vps20-GFP (green, driven by Cg-GAL4) and ERES Tango1 (antibody staining, magenta). Magnified insets are shown for both markers and Vps20 alone. (C) Localization in fat body of Syx17-GFP (green) and trans-Golgi Golgin245 (antibody staining, magenta). Magnified insets show both markers and Syx17 alone. (D) Localization in fat body of Syx17-GFP (green) and trans-Golgi marker GalT-TagRFP (magenta), both driven by BM-40-SPARC-GAL4. Magnified insets show both markers and Syx17 alone. (E) Localization in fat body of Lamp1-YFP (green) and trans-Golgi Golgin245 (magenta). Magnified insets show both markers and Lamp1 alone. (F) Localization in fat body of Lamp1-YFP (green) and trans-Golgi GalT-TagRFP (magenta, driven by BM-40-SPARC-GAL4). Magnified insets show both markers and Lamp1 alone. Circles and arrows mark examples of tGA- and free lysosomes, respectively. (G) Quantification of the proximity between Lamp1-YFP and trans-Golgi Golgin245. The upper graph represents proximity in 10 cells measured in images like those in E. The lower graph represents average proximity values (n = 10, error bars indicate SD): 84.6% of Golgi elements display Lamp1 puncta, while 75.1% of Lamp1 puncta are Golgi-associated. (H) Quantification of the proximity between Lamp1-YFP and trans-Golgi GalT-TagRFP as in G, measured in images like those in F. 86.1% of Golgi elements display Lamp1 puncta, while 75.4% of Lamp1 puncta are Golgi-associated. (I) Confocal image of fat body showing localization of lysosome marker Lamp1-YFP (green), ERES Tango1 (antibody staining, blue), and trans-Golgi GalT-TagRFP (magenta, driven by BM-40-SPARC-GAL4). ERES-Golgi units are shown below at higher magnification. (J) Schematic drawing of a Drosophila ERES-Golgi unit displaying a tGA-lysosome. (K) Confocal images of fat body showing localization of trans-Golgi marker GalT-TagRFP (magenta, driven by BM-40-SPARC-GAL4) and endogenously expressed, YFP-tagged Rab-GTPases Rab4, Rab5, Rab7, and Rab11 (green; Dunst et al., 2015). Magnified insets in lower right corner of each image with separate channels below. (L) Proximity between trans-Golgi marker GalT-TagRFP and fluorescently tagged Rab-GTPases Rab4, Rab5, Rab7, and Rab11. The upper graphs represent puncta proximity in 10 cells measured in images like those in H. The lower graphs represent average proximity (n = 10, error bars indicate SD).

Juxta-Golgi localization of Rop, Vps20, lysosomal markers, and endosomal Rab-GTPases. (A) Confocal image of L3 fat body (feeding stage) showing localization of Rop-GFP (green) and cis-Golgi marker Grasp65-RFP (magenta), both driven by Cg-GAL4. Magnified insets are shown below for both markers and for Rop alone. (B) Localization in fat body of Vps20-GFP (green, driven by Cg-GAL4) and ERES Tango1 (antibody staining, magenta). Magnified insets are shown for both markers and Vps20 alone. (C) Localization in fat body of Syx17-GFP (green) and trans-Golgi Golgin245 (antibody staining, magenta). Magnified insets show both markers and Syx17 alone. (D) Localization in fat body of Syx17-GFP (green) and trans-Golgi marker GalT-TagRFP (magenta), both driven by BM-40-SPARC-GAL4. Magnified insets show both markers and Syx17 alone. (E) Localization in fat body of Lamp1-YFP (green) and trans-Golgi Golgin245 (magenta). Magnified insets show both markers and Lamp1 alone. (F) Localization in fat body of Lamp1-YFP (green) and trans-Golgi GalT-TagRFP (magenta, driven by BM-40-SPARC-GAL4). Magnified insets show both markers and Lamp1 alone. Circles and arrows mark examples of tGA- and free lysosomes, respectively. (G) Quantification of the proximity between Lamp1-YFP and trans-Golgi Golgin245. The upper graph represents proximity in 10 cells measured in images like those in E. The lower graph represents average proximity values (n = 10, error bars indicate SD): 84.6% of Golgi elements display Lamp1 puncta, while 75.1% of Lamp1 puncta are Golgi-associated. (H) Quantification of the proximity between Lamp1-YFP and trans-Golgi GalT-TagRFP as in G, measured in images like those in F. 86.1% of Golgi elements display Lamp1 puncta, while 75.4% of Lamp1 puncta are Golgi-associated. (I) Confocal image of fat body showing localization of lysosome marker Lamp1-YFP (green), ERES Tango1 (antibody staining, blue), and trans-Golgi GalT-TagRFP (magenta, driven by BM-40-SPARC-GAL4). ERES-Golgi units are shown below at higher magnification. (J) Schematic drawing of a Drosophila ERES-Golgi unit displaying a tGA-lysosome. (K) Confocal images of fat body showing localization of trans-Golgi marker GalT-TagRFP (magenta, driven by BM-40-SPARC-GAL4) and endogenously expressed, YFP-tagged Rab-GTPases Rab4, Rab5, Rab7, and Rab11 (green; Dunst et al., 2015). Magnified insets in lower right corner of each image with separate channels below. (L) Proximity between trans-Golgi marker GalT-TagRFP and fluorescently tagged Rab-GTPases Rab4, Rab5, Rab7, and Rab11. The upper graphs represent puncta proximity in 10 cells measured in images like those in H. The lower graphs represent average proximity (n = 10, error bars indicate SD).

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