Fig. 6
From: Sphingolipid-dependent Dscam sorting regulates axon segregation
Dscam[TM1]-FasII interaction defines axonal versus dendritic domain in MB neuron. a–f MB of lacek05/ + (a–c) and lace2/lacek05 (d–f) labeled for FasII with mCD8::GFP (a, d), Dscam[TM1]::GFP (b, e) or Apc2::GFP (c, f) expressed using OK107-Gal4. Inset represents FasII distribution from the area indicated by the dotted box. White dotted lines 1-3 in b, c, e, f represent three peduncular zones for analysis and the corresponding cross section of the peduncle (from a different image) is represented in the insets 1-3. In SPT mutants, membranous FasII shifts upwards into the dendritic compartment (inset (d), arrowhead). The expression of Dscam[TM1]::GFP causes axo-dendritic boundary of FasII to shift deeper into the peduncle, while retaining the exclusion from Dscam[TM1]::GFP (b). FasII partially regains its axo-dendritic boundary in SPT mutants owing to absence of membranous Dscam[TM1] ((e), arrowhead insets). Expression of the cytosolic dendritic marker Apc2::GFP causes no shift in FasII localization (c, f). g–l MB in the wild-type background with α′/β′ neuron-specific expression (R26E01-Gal4) of Dscam[TM1]::GFP (g), FasIIPEST− (h), Dscam[TM1]::GFP + FasIIPEST- (i), Dscam[TM1]::RNAi + FasIIPEST− (j), Dscam[TM1]::RFP + intra-FasII-YFP (k), and Dscam[TM1]::RFP + extra-FasII-YFP (l). Full length (FL) FasII and Dscam[TM1] maintains exclusion in α′/β′ neurons (g–i). Interestingly, Dscam[TM1] knockdown causes FasII to shift into the somato-dendritic compartment (j). On the other hand, intra-FasII-YFP (k) but not extra-FasII-YFP (l) shows membranous exclusion from Dscam[TM1], indicating that the membranous exclusion is mediated by interactions in the intra-cellular domain. The white arrowhead marks the axo-dendritic boundary, the white dotted circle marks calyx. n(g) = 28, n(h) = 40, n(i) = 24, n(j) = 36, n(k) = 28, n(l) = 28. a–l Green: GFP, Red: FasII, Blue: N-Cad, Scale Bar: 25 μm
