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. 2021 May 5;40(13):e105770. doi: 10.15252/embj.2020105770

Figure 8. Irradiation induces YAP‐TEAD activity to drive crypt expansion, which also requires Wnt signalling.

Figure 8

  1. Control and irradiated murine small intestines analysed with RNAscope in situ hybridisation for Myc mRNA (black arrows point to mRNA granules in the control) and by immunostaining for Sox9 protein reveals an expansion in the expression domain of both genes. Nuclear localisation of beta‐catenin is readily detectable after irradiation. n = 3 samples per condition, quantification shown in the insets, error bars = 1 SD. Statistical analysis was performed with an unpaired Student's t‐test. Similar results were obtained at 3 and 5 dpi.
  2. qPCR analysis of YAP, TEADs and their target genes Ctgf and Cyr61 over a 4 days post‐irradiation time course at 12 Gy. Note the strong upregulation of TEAD4, Ctgf and Cyr61. In contrast, Axin2, TEAD2 and Sox9 mRNA expression levels are only moderately altered, confirming that Wnt signalling remains active upon irradiation‐induced YAP‐TEAD activation. n = 4 biological replicates per time point. Statistical analysis was performed with unpaired t‐test compared to the control 0 Gy condition, *P < 0.05, **P < 0.01, ***P < 0.001.
  3. Control and irradiated murine small intestines analysed by immunostaining for Sox9 protein reveals that treatment with Porcupine inhibitor LGK974 strongly reduces the Sox9 expression domain induced by 3 days after 12 Gy irradiation. Phosphorylated Src immunostaining reveals an upregulation by 3 days after irradiation which is also lost upon treatment with Porcupine inhibitor LGK974 (n = 4 animals in each condition).
  4. qPCR analysis of Wnt and YAP target gene expression in intestinal crypts isolated at 1 day after 12 Gy irradiation. Note the inhibition of Axin2, Sox9, YAP, Ctgf, Cyr61 and TEAD family expression upon treatment with Porcupine inhibitor LGK974. The mild reduction of Sox9 mRNA expression by LGK974 at 1 day after irradiation (1 dpi) becomes more pronounced at 3 days after irradiation (3 dpi). n = 4 biological replicates per condition, statistical analysis was performed with Student's two‐tailed t‐test, error bars = 1 SD, *P < 0.05, **P < 0.01, ***P < 0.001.