Extended Data Fig. 3: Generation and validation of conditional Taz gain-of-function mice.
From: A YAP/TAZ-TEAD signalling module links endothelial nutrient acquisition to angiogenic growth
a, Schematic representations of the wild-type Rosa26 locus, the targeting vector, the recombined as well as the excised allele are shown. A cassette containing the CAG promoter, a floxed STOP sequence, a cDNA encoding for 3xFLAG-tagged TAZS89A and IRES-nGFP was inserted into the Rosa26 locus. Triangles denote loxP sites. DTA, diphteria toxin negative selection marker; pA, polyA signal. b, Immunoblot analysis of total lung lysates from control (Ctrl, Rosa26-TazS89A fl/fl) and mutant mice (TaziEC-GOF, Pdgfb-creERT2;Rosa26-TazS89A fl/fl). 4OHT was administered from P1 to P4 and samples analyzed at P6. Arrow heads indicate expression of the FLAG-tagged TazS89A mutant. c, RT-qPCR analysis of retinal ECs at P6, showing increased expression of the canonical YAP/TAZ target gene Ctgf in TaziEC-GOF mice when compared to Ctrl (n = 4 independent samples). d, Immunofluorescence staining for GFP, ERG and PECAM in P6 retinas depicting a nuclear GFP signal in PECAM positive vessels of TaziEC-GOF mutants, which is absent in Ctrl mice. e, Higher magnification images of P6 retinas labelled for EdU, ERG and PECAM showing increased EC proliferation in the TaziEC-GOF mice. f, g, Confocal images (f) and quantification of endothelial coverage (g) in PECAM labelled P6 retinas obtained from Ctrl (Yapfl/fl;Tazfl/fl;Rosa26-TazS89A fl/wt), Yap/TaziEC-KO (Pdgfb-creERT2;Yapfl/fl;Tazfl/fl;Rosa26-TazS89A wt/wt) and Yap/TaziEC-KO;TaziEC-GOF (Pdgfb-creERT2;Yapfl/fl;Tazfl/fl;Rosa26-TazS89A fl/wt) mice (EC coverage: n = 16 (Ctrl), 7 (Yap/TaziEC-KO) and 8 (Yap/TaziEC-KO;TaziEC-GOF) independent samples). Western blot data in b are from the respective experiment, processed in parallel, and are representative of at least three independent experiments. For c, g, data represent mean ± s.e.m.; two-tailed unpaired t-test. *P < 0.05; **P < 0.01; ****P < 0.0001. The numerical data, unprocessed western blots and P values are provided as source data.
