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Fig. 4. N-cadherin functions in the first layer-selection stage to promote R7 axons reaching and remaining in the R7-temporary layer. R7 targeting was assessed at 17% APF (A-D'), at 35% APF (E-H'), and in adult flies (I-L'). Single mutant R7 cells were generated using GMR-Flp-mediated mitotic recombination (see text), and labeled using the MARCM system and mCD8-GFP (A-H') or synb-GFP (I-L'). R7 and R8 axons were visualized with Mab24B10 (red). (A,A',E,E',I,I') Wild-type; (B,B',F,F',J,J') Ncad⁴⁰⁵ mutant; (C,C',G,G',K,K') LAR²¹²⁷ mutant; (D,D',H,H',L,L') Ncad⁴⁰⁵ LAR²¹²⁷ double mutant. (A,A') At 17% APF, single R7 cells, homozygous for a wild-type FRT40 chromosome arm, project axons into the medulla. The wild-type axons first project past the R8 layer (arrow), then expand their growth cones (double arrow) just below the R8 growth cones, and separate (arrowhead) from the R8 growth cones. (B,B') At 17% APF, single Ncad⁴⁰⁵ mutant R7 axons project into the medulla. Approximately 21% of these mutant axons expand their growth cones incorrectly at the R8-temporary layer or between the R7- and R8-temporary layers (arrows). Over half of these mutant growth cones show severe morphological defects (arrowheads). (C,C') Single LAR²¹²⁷ mutant R7 axons project into the medulla as in wild-type at 17% APF. (D,D') At 17% APF, single Ncad⁴⁰⁵ LAR²¹²⁷ double mutant R7 axons exhibit targeting (arrows) and morphological defects (arrowhead) as seen in Ncad⁴⁰⁵ mutants. (E,E') Single wild-type R7 axons (arrow) terminate at the R7-temporary layer at 35% APF. (F,F') At 35% APF, 55% of the Ncad mutant R7 axons (arrow) terminate at the R8-temporary layer or between the R7- and R8-temporary layers. Some of them leave a small filapodium (arrowhead) connecting to the R7-temporary layer. (G,G') At 35% APF, most single LAR²¹²⁷ mutant R7 axons terminate correctly at the R7-temporary layer in the younger part of the medulla. Approximately 9% of the mutant R7 growth cones exhibit abnormal morphology (arrowhead). Assessment of older R7 axons at this stage is limited because Elav-Gal4 driver is expressed at a low level in the older R7 cells. (H,H') At 35% APF, single Ncad⁴⁰⁵ LAR²¹²⁷ double mutant R7 axons targeted to incorrect layers (arrows), as seen in the Ncad⁴⁰⁵ mutant. (I,I') In adult flies, single wild-type R7 axons terminate at the R7-recipient layer. (J-L') At the adult stage, single Ncad⁴⁰⁵ or LAR²¹²⁷ or Ncad⁴⁰⁵ LAR²¹²⁷ double mutant R7 axons terminate incorrectly at the R8-recipient layer. Note that in the region where mutant R7 axons mistarget to the R8-recipient layer, the corresponding R8 axons target correctly to the R8-recipient layer, leaving the R7-recipient layer uninnervated by any R-cell afferent. The presumptive R7- and R8-temporary layers are indicated by dotted lines in A'-D',I'-L'). (A'-L') High-magnification views of A-L, respectively. Scale bars: in A, 30 µm for A-L; in A', 10 µm for A'-L'. Abbreviations as in Fig. 1.

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