(A) Schematic of the BRV-B-Myc transgene. Numbers indicate nucleotide positions. (B-D) Basal sections of discs following rnts>rpr ablation on day 7 (left panel) and day 9 (center panel), and on day 9 in discs bearing BRV-B-Myc transgene (right panel), imaged at the time of downshift. (B) Myc expression following ablation. Myc is expressed in basal wound periphery cells of a day 7 disc, but is absent in a day 9 ablated disc. The BRV-B-Myc transgene causes expression of Myc protein in an older disc. (C- D) Discs bearing the AP-1 GFP reporter (C) or stained with anti-Mmp1 (D), and imaged as in (B). AP-1 reporter expression is strongly expressed in basal cells of the pouch in day 7 ablated discs, coincident with Mmp1 expression. AP-1 reporter expression is found in the ring of wound periphery cells in day 9 ablated discs (arrowheads), while Mmp1 staining is reduced. Day 9 ablated discs bearing BRV-B-Myc have AP-1 GFP expressing cells covering the basal wound surface, resembling the expression pattern of day 7 discs, while Mmp1 expression is stronger than a wild type day 9 disc. (E-F) Discs stained for Wg (E) or bearing the BRV118-GFP reporter (F) following ablation with rnts>egr, and imaged as in (B-D). Declining expression of the reporter on day 9 is rescued in discs bearing the BRV-B-Myc transgene. Similarly, wg expression is stronger on a day 9 bearing the BRV-B-Myc transgene, comparable to expression level in an ablated day 7 disc. (G-G’) A day 9 disc bearing the BRV-BC enhancer with heat shock induced flip-out clones (marked by RFP, red) expressing Myc, irradiated with 45 Gy and dissected 16 hr later. Clones expressing Myc respond to damage by activating the BRV-BC enhancer (G’, green), while the enhancer remains inactive in neighboring damaged tissue not expressing Myc. (H-H’) Day 9 discs as in (G-G’), in the absence of irradiation. Expression of Myc in clones (red) causes low level of BRV-BC enhancer activity (H’, green), even in the absence of damage. (I) Assay of adult wing sizes that develop following ablation with rnts>rpr on day 7, 8, 9 and 10 in wild type and BRV-B-Myc discs, quantifying animals that eclose with fully regenerated wings. The BRV-B-Myc expressing animals consistently eclose with more of the full regenerated wings compared to wild type. Mean differences between wild type and BRV-B-Myc data sets is statistically significant, (p-value calculated by two way ANOVA). Error bars are SEM of at least 3 biological repeats, scoring a total of >200 animals in each condition. (J) Developmental timing of wild type and BRV-B-Myc expressing larvae following rnts>rpr ablation on day 7, 8, 9 and 10. Delay is measured as hours at which 50% of larvae have pupariated compared to unablated wild type. The BRV-B-Myc transgene (red bars) does not alter developmental timing following ablation compared to wild type (gray bars) on any day ablated. Mean differences between wild type and BRV-B-Myc data sets is not statistically significant (N.S.), calculated by two way ANOVA. Error bars are SEM of 3 biological repeats, n>100 flies for each genotype/condition.