Figure S4.
SNX1-GFP, CIP4-EGFP, and MIM-mScarlet localization. (A) Confocal intensity-projection and time-lapse sequence of a representative SG expressing SNX1-GFP (green) and LifeAcrt-Ruby (magenta) at increasing magnification. Yellow squares mark the magnified area. Left: Representative image of a cell (confocal intensity-projection); SNX1 is seen on cell membranes and in the cytoplasm. Right: Time-lapse sequence of representative secreting LSV (SRRF intensity-projection). Specific localization of SNX1-GFP to the fusion pore or to the LSV is not observed. (B) Confocal intensity-projection and time-lapse sequence of a representative SG expressing CIP4-EGFP (green) and LifeAct-Ruby (magenta), at increasing magnification. Yellow squares mark the magnified area. Left: Representative image of a cell (confocal intensity-projection); CIP4-EGFP is seen mostly on the apical and lateral membranes of the cell. Right: Time lapse of representative secreting LSV (SRRF intensity-projection). CIP4-EGFP is mostly apical and localizes to the LSV membrane after fusion, but specific localization to the pore was not detected. (C) Representative SG expressing MIM-mScarlet (Fig. 6 A; magenta). Left: Overview of a large area in the gland (confocal intensity-projection). The yellow square shows the area enlarged in the middle image. Middle: Enlarged cell overview. Like MIM-Emerald (Fig. 5 A), MIM-mScarlet localizes to cytoplasmic clusters and is absent from the apical membrane. Additionally, MIM-mScarlet localizes in the cytoplasm and to lateral membranes. Right: Representative LSVs from different vesicles, in various stages of secretion (confocal intensity-projections). MIM-mScarlet is observed at and around the pore (white arrowheads) and is localized to LSV membrane during secretion. UAS-based expression is driven by c135-GAL4. Scale bars in A (left) and B (left), 10 µm; in A (right) and B (right), 1 µm; in C (left), 40 µm; in C (middle), 10 µm; in C (right), 1 µm.

SNX1-GFP, CIP4-EGFP, and MIM-mScarlet localization. (A) Confocal intensity-projection and time-lapse sequence of a representative SG expressing SNX1-GFP (green) and LifeAcrt-Ruby (magenta) at increasing magnification. Yellow squares mark the magnified area. Left: Representative image of a cell (confocal intensity-projection); SNX1 is seen on cell membranes and in the cytoplasm. Right: Time-lapse sequence of representative secreting LSV (SRRF intensity-projection). Specific localization of SNX1-GFP to the fusion pore or to the LSV is not observed. (B) Confocal intensity-projection and time-lapse sequence of a representative SG expressing CIP4-EGFP (green) and LifeAct-Ruby (magenta), at increasing magnification. Yellow squares mark the magnified area. Left: Representative image of a cell (confocal intensity-projection); CIP4-EGFP is seen mostly on the apical and lateral membranes of the cell. Right: Time lapse of representative secreting LSV (SRRF intensity-projection). CIP4-EGFP is mostly apical and localizes to the LSV membrane after fusion, but specific localization to the pore was not detected. (C) Representative SG expressing MIM-mScarlet (Fig. 6 A; magenta). Left: Overview of a large area in the gland (confocal intensity-projection). The yellow square shows the area enlarged in the middle image. Middle: Enlarged cell overview. Like MIM-Emerald (Fig. 5 A), MIM-mScarlet localizes to cytoplasmic clusters and is absent from the apical membrane. Additionally, MIM-mScarlet localizes in the cytoplasm and to lateral membranes. Right: Representative LSVs from different vesicles, in various stages of secretion (confocal intensity-projections). MIM-mScarlet is observed at and around the pore (white arrowheads) and is localized to LSV membrane during secretion. UAS-based expression is driven by c135-GAL4. Scale bars in A (left) and B (left), 10 µm; in A (right) and B (right), 1 µm; in C (left), 40 µm; in C (middle), 10 µm; in C (right), 1 µm.

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