Figure 5. PAM‐DANs are the source of DIP‐δ in the γ4/5 zones.

• A–D
  Confocal z‐projections of brains expressing DIP‐δ^GFSTF (DIP‐δ‐GFP) together with the indicated Gal4s and transgenes. Expressing diphtheria toxin (DTi) and membrane‐bound RFP (mCD8‐RFP; CD8) driven by the γ4 > γ1γ2 MBON driver MB294B‐Gal4 (MBONγ4‐Gal4) did not affect DIP‐δ‐GFP expression (A, n = 16/16; B, n = 14/14). In contrast, similar expression of DTi and membrane‐bound tomato (CD4‐tdT; CD4) in PAM‐DANs (using the GMR58E02‐Gal4; PAM‐DAN‐Gal4) abolished the normal DIP‐δ‐GFP expression in the γ4/5 zone (compare D, n = 18/18, to C, n = 16/16) and within the PAM‐DAN cell bodies (compare D’ to C’). Magenta is CD8‐RFP (A, B) and CD4‐mtdT (C, D). Green is GFP, and grayscale depicts individual channels as labeled. Scale bar is 20 µm. Yellow dashed line demarcates the γ‐lobe based on FasII staining (not shown).
• E–G
  Confocal z‐projections of MARCM clones labeled by DIP‐δ^T2A‐Gal4 (DIP‐δ Gal4) driving the expression of membrane‐bound GFP (mCD8‐GFP; CD8) and heat shocked at 24 h after egg laying. Clones innervate the γ4/5 zones at 24 h APF (E; n = 8), 48 h APF (F; n = 8), and adult (G; n = 16). Clones become tyrosine hydroxylase (TH) positive only at 48 h APF onwards (F, G). Magenta is FasII, green is mCD8‐GFP, cyan is TH antibody staining, and grayscale single channels are shown as indicated. White dashed line demarcates the γ‐lobe. Scale bar is 20 µm.