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. 2023 Oct 16;19(10):e1010979. doi: 10.1371/journal.pgen.1010979

Fig 4. The conserved trafficking regulators Arf1 and AP-1 are essential for DCG biogenesis in SCs.

Fig 4

(A-E) Representative images of SCs expressing the DCG marker GFP-GPI together with a control RNAi (A) or RNAis targeting Arf1 (RNAi #1; B), AP-1γ (C), AP-1μ (D) or AP-1σ (E). Cellular organisation was assessed using DIC imaging, GFP-GPI fluorescence and Lysotracker Red fluorescence; a merged image is also shown for each cell. Note that the knockdowns generally reduce the number of large non-acidic compartments and the number of DCG compartments, though some remaining compartments can be expanded in size. (F) Bar chart showing number of compartments containing GFP-labelled DCGs in control SCs and following knockdown of Arf1 and AP-1 subunits. (G) Bar chart showing number of non-acidic compartments in these different genotypes. (H) Bar chart showing the percentage of non-acidic compartments with diffuse GFP-GPI in these different genotypes. Approximate outlines of SCs are marked by dashed circles. Scale bars: 10 μm. Data are typically for three cells per accessory gland; accessory glands from ≥10 individual males were imaged during three to six separate imaging sessions, for this and subsequent knockdown experiments. For F-H, bars show mean ± SD; Control, n = 28; Arf1 #1, n = 31; Arf1 #2, n = 30; AP-1γ, n = 28; AP-1μ, n = 32; AP-1σ, n = 30. P<0.05: *, P<0.01: **, P<0.001: ***, P<0.0001: ****. Genotypes for images: (A) w1118; P{tub-GAL80ts}/P{ryTRiP.HMS02827}; dsx-GAL4, P{UAS-GFP.GPI}/+; (B) w1118; P{tub-GAL80ts}/+; dsx-GAL4, P{UAS-GFP.GPI}/P{Arf1GD12522}; (C) w1118; P{tub-GAL80ts}/+; dsx-GAL4, P{UAS-GFP.GPI}/P{AP-1γTRiP.JF02684}; (D) w1118; P{tub-GAL80ts}/+; dsx-GAL4, P{UAS-GFP.GPI}/P{AP-1μGD14206}; (E) w1118; P{tub-GAL80ts}/P{AP-1σKK108869}; dsx-GAL4, P{UAS-GFP.GPI}/+.