DEV021428 Supplementary Material

Files in this Data Supplement:

• Supplemental Figure S1 -

  Fig. S1. Apoptosis induction in lin mutant cells in the wing disc. (A-C) Wing discs containing lin^G1 clones induced at 48-72 hours AEL. (A) lin^G1 clones (marked by the lack of GFP and broken line) express high level of phospho-histone 3 (red) compared with the wild-type twin spots cells (bright green owing to double the GFP dose). (B) puc-LacZ expression in lin^G1 clones (lack of GFP). lin^G1 cells activate the Jun-kinase target puc-lacZ (red). Inset in B shows puc-LacZ expression in the wing disc. (C) Caspase 3 expression (red) in lin^G1 MARCM clones (labelled by GFP). Mutant cells in the border of the clones show lower GFP staining and higher levels of Caspase 3, indicating that cells start dying at the clone boundaries. (D) Wg expression (green) in a wing imaginal disc containing lin^G1 clones (lack of β-Gal) induced at 72-96 hours AEL (red). Note the wing disc duplication marked by duplicated Wg expression in the wing margin (arrowheads) and in the notum (asterisks). (E) When lin^G1 clones were induced at 24-48 hours AEL, only twin spots are observed (bright green due to double dose of GFP). A black spot indicates where the lin^G1 clone was originated (arrowheads).

• Supplemental Figure S2 -

  Fig. S2. Effects of Drm overexpression in the wing disc. (A-C) Wg, Dl and Vg (in green) in wing discs containing ectopic Drm random clones (red). Wg and Dl are ectopically expressed in all Drm ectopic clones. Vg expression is reduced in the mutant cells located far from the DV border (the vg quadrant enhancer is a target of Wg pathway, arrowheads), but it is maintained in those mutant cells close to DV border (the vg boundary enhancer is a target of N pathway, arrows). (D) Transient Drm expression in clones (red) using tubGal80^ts technique, activates Hth (green) after a 24-hour pulse of ectopic Drm expression and a further 24 hours at the restrictive temperature.

• Supplemental Figure S3 -

  Fig. S3. Lin-Gro genetic interaction. (A-F) Adult phenotypes, including duplications of the eye (A,B), leg (C,D) and antenna (E,F) are seen in lin^G1 clones (induced 48-72 hours AEL) (A,C,E) and in the heterozygous combination gro¹/+; lin^G1/+ (B,D,F). (G,H) Percentage of gro¹/+; lin^G1/+ flies showing phenotypic alterations. (I-I′) Imaginal wing disc containing lin^G1- MARCM clones expressing Gro labelled by GFP. The ectopic expression of Wg observed in lin⁻ cells is recovered by co-expression of Gro (red in I, grey in I′).

• Supplemental Figure S4 -

  Fig. S4. Effects of Gro, H and Mtv expression on Wg targets in the wing disc. (A-C) Sens expression (green) is repressed in ectopic Gro clones (red in A) but is not affected either by ectopic H clones (red in B) or by ectopic expression of Mtv using ptc-Gal4 driver. Ptc expression domain abuts the AP border and is marked with a broken line (C).

• Supplemental Figure S5 -

  Fig. S5. Alterations of EGFR, Dpp and JAK/STAT pathways in lin⁻ clones in the wing disc. (A-E) lin^G1 clones induced at 48-72 hours AEL in a wing disc. (A) Ectopic activation of Kekkon-lacZ (red) in lin^G1 clones (labelled by lack of GFP). Inset shows wild-type expression of Kekkon-lacZ. (B) Ectopic activation of Iro in lin^G1 clones (lack of β-Gal expression in red). Inset shows wild-type expression of Iro complex gene Caupolican (green). (C) Expression of the Dpp repressor Brk (red) is upregulated in lin^G1 clones (labelled by absence of GFP). Inset shows wild-type expression of Brk-LacZ. (D,D′) Two confocal sections of the same disc to show that the Dpp target gen Sal (green) is repressed in lin^G1 clones (lack of red).) Inset indicates wild-type expression of Sal. (E) STAT92E-lacZ activation (red) in lin^G1 clones (lack of the GFP in green). Inset shows expression of the STAT92E-lacZ reporter in the wing disc.