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. 2020 Dec 29;40(4):e104347. doi: 10.15252/embj.2019104347

Figure EV4. Identification of transcription factors required for the activity of E3N and 9CJ2 svb enhancers in the embryo.

Figure EV4

  1. Drawing of the E3N svb enhancer, with position of putative binding sites for Pnt (red) and TCF (green) factors, and evolution of DNA sequences across Drosophila species. Nucleotides in red represent point mutations introduced to disrupt either Pnt‐ or TCF‐binding sites.
  2. Consequences of knocking out Pnt‐ or TCF‐binding sites on expression of the E3N svb enhancer in the embryonic epidermis. Pictures show ventral views of stage‐15 embryos. Scale bar is 50 µm.
  3. Trichome rescue assays (Crocker et al, 2015) showing the influence of TCF‐binding sites on E3N function. Picture show cuticle preparations of wild‐type and svb‐mutant embryos, focusing on the ventral region of A6 segments. svb mutants display strong reduction in the number of trichomes, remaining ones being highly abnormal. Consistent with its expression pattern, E3N driving svb cDNA (E3N‐wt::svb) rescues formation of the anterior‐most trichome row (arrow). Knocking out TCF‐binding sites (E3N‐TCF‐mt::svb) disrupts rescuing ability of the E3N enhancer. The graph plots the number of trichomes in the anterior‐most row. Boxes extend from the 25th to 75th percentiles, whiskers from min to max, the horizontal line in each box is plotted at the median; data were collected from three independent replicates. P values from one‐way ANOVA are ns > 0.5, **** < 0.0001. Scale bar is 15 µm.
  4. Drawing of the 9CJ2 svb enhancer, with position of putative binding sites for Pdm‐1 (orange) and evolutionary conservation of the DNA sequence. Nucleotides in red show mutations that have been introduced to disrupt Pdm‐binding sites.
  5. Consequences of knocking down Pdm‐1‐binding sites on expression of the 9CJ2 svb enhancer in the embryonic epidermis. Pictures show lateral (left), ventral (middle), and dorsal (right) views of stage‐15 embryos. Scale bar are 50 µm.

Data information: Dmel, Drosophila melanogaster; Dsim, Drosophila simulans; Dyac, Drosophila yacuba; Dere, Drosophila erecta; Dfic Drosophila ficusphila; Dtak, Drosophila takahashii; Dana, Drosophila ananassae.