Figure 3. The BRV118 enhancer is necessary for wg and Wnt6 expression following tissue damage.

(A) Schematic of the major WNT locus showing the BRV118 enhancer (blue rectangle). Approximate intergenic distances are labeled. (B) RNA in situ hybridization to detect wg, Wnt4, Wnt6 and Wnt10 RNA in day 7 unablated (top row) and rn^ts>egr ablated (bottom row) wing discs. Only wg and Wnt6 demonstrate significant upregulation of RNA in response to damage. (C) RNA in situ hybridization to detect wg, and Wnt6 RNA in discs ablated with rn^ts>egr on day 7 and day 9. Transcription of both wg and Wnt6 in response to damage is absent in a day 9 disc. (D) Schematic of the lesion in the wg¹ allele that deletes most of the BRV118 enhancer. (E) RNA in situ hybridization to detect wg and Wnt6 RNA and Wg protein, in wild type (top row) and homozygous wg¹ (bottom row) day 7 rn^ts>egr ablated discs. The damage-specific expression of wg and Wnt6 RNA, and Wg protein levels, are reduced in wg¹ discs compared to wild type. (F) Adult wing sizes following ablation with rn^ts>egr on day 7 in wild type and wg¹ homozygotes, indicating the percentage of animals that eclose with fully regenerated wings. Regeneration of wg¹ homozygous discs is significantly reduced compared to wild type. Error bars indicate SD of at least three independent experiments, scoring a total of >200 animals in each experiment. Only untransformed wg¹ wings were scored (for explanation of untransformed see Figure 3—figure supplement 1).

DOI: http://dx.doi.org/10.7554/eLife.11588.011

Figure 3—figure supplement 1. The wg¹ allele causes transformation of wing to notum in a proportion of the population.

(A-A’) Wing discs dissected from day 7 wg¹ homozygous larvae and stained for Wg (red) and DAPI (blue), showing a transformed disc (A), in which presumptive wing tissue is replaced by notum, indicated by the duplicated wg notum stripe (arrowheads), and an untransformed wing disc (A’) that is morphologically normal and has wild type wg expression. Transformation is thought to occur at L2 when the wing pouch is specified by wg expression (Sharma and Chopra, 1976; Whitworth and Russell, 2003), and thus ablation of the pouch by rn^ts>rpr or rn^ts>egr does not occur in these discs, (B-B’) A wg¹ homozygous adult with one untransformed left wing and a transformed right wing. Wing tissue is replaced by additional notum and associated bristles following transformation (arrowheads). (C-C’) For comparison, a wild type adult with two non-regenerated wings resulting from rn^ts>rpr ablation. The presence of hinge tissue and normal bristle formation allows clear distinction between ablated versus transformed wings; only untransformed wings were scored in our assays. (D) Quantification of the wing to notum transformation frequency in the wg¹ homozygous stock used in our experiments. Error bars are SD, n>300 flies. On average 58% of animals eclose with two untransformed wings. The frequency of the variably penetrant transformation phenotype can be shifted by selective breeding of two wing, one wing or no wing flies, but not eliminated (Sharma and Chopra, 1976).

DOI: http://dx.doi.org/10.7554/eLife.11588.012