Fig. 3: The EGFR/Ras1 pathway influences Ihog stabilized cytonemes.

A Cytonemes stabilized by Ihog^RFP in Hh-producing cells in hh.Gal4 tub.Gal80^ts > UAS.RHGmiRNA, UAS.Ihog^RFP wing discs. B–D Ihog^RFP stabilized cytonemes are reduced or nearly absent after transient inhibition of the EGFR by UAS.^EGFRRNAi (B) or UAS.EGFR^DN (C) and augmented by expressing a constitutively active form of EGFR (UAS.EGFR^CA) (D). E Quantification of cytonemes number within a selected window measuring the proportional 20% of the wing disc pouch size to normalize (wing discs represented by a dot). The statistical analysis performed using two-side pairwise comparisons with Wilcoxon rank-sum test shows a significant reduction after EGFR^RNAi (p < 0.0002). or UAS.EGFR^DN expression (p < 0.0005) while it marginally increased upon over-expression of UAS.EGFR^CA (p < 0.0005). F Quantification of the maximum lengths reached by cytonemes (measured as described above) reveals a significant difference between expressing UAS.Ihog^RFP alone and coexpressing EGFR^RNAi (p < 0.0001) or UAS.EGFR^DN (p < 0.0005), while it slightly increases with high variability upon co-expression of EGFR^CA (p < 0.0002). G Cytonemes of hh.Gal4, tub.Gal80^ts > UAS.Ihog^RFP wing disc as control. H, I Ihog^RFP stabilized cytonemes are reduced after transient inhibition (for 22 h) of the Ras1 (UAS.Ras1^RNAi) (H) or after transient expression (for 20 h) of a dominant negative form of Ras1 (UAS.Ras1^DN) (I). J, K Quantification of cytoneme number (control, Ras1^RNAi, Ras1^DN p < 0.002) (J) and of the maximum lengths that cytonemes as in (F) (Ras1^RNAi, Ras1^DN p < 0.002) (K). L–O Wing disc containing Ihog^RFP expressing clones (L, M) and coexpressing EGFR^RNAi in the dorsal compartment (N, O) showing in basal sections stabilized cytonemes emanating from a clone expressing Ihog^RFP under normal conditions (L) or co-expressing EGFR^RNAi in the dorsal side, keeping the ventral side as control (N). Drawing model shows clones in yellow and the EGFR^RNAi expression domain in purple. Note that cytonemes protruding from cells within the EGFR downregulated area are very short or absent and that Ihog^RFP levels membranes severely decrease after EGFR inhibition (N, O). P Statistic analysis for Ihog^RFP levels expressed alone or co-expressed with EGFR^CA, EGFR^RNAi Ras1^V12, Ras1^RNAi or Ras1^DN in the P compartment (p < 0.0001). The panels display the data point distribution with mean and SD whiskers. Scale bars = 10 μm. Source data are provided as a Source Data file.