Fig. 5

TATA-box binding protein associated Factor 1 (TAF1) regulated the Notch signaling pathway in vitro and in vivo. A: the chromatin immunoprecipitation (ChIP) assay was performed to detect the binding ability between TAF1 and the NOTCH1 promoter in HEK293T cell lines. B: the ratios of luciferase activity regulated by the NOTCH1 response element to Renilla luciferase in HEK293T cells transfected with vector alone, wild-type TAF1 (WT), mutant TAF1 (MU), or Notch intracellular domain (NICD) overexpression plasmid (NICD). C: the expression of SLC5A5 and ACTIN was determined by Western blot (n = 3). TAF1-inhibited Nthy-ori 3-1 cells were transfected with vector alone, wild-type TAF1 (WT), mutant TAF1 (MU), or NICD overexpression plasmid (NICD). D: the relative protein expression (SLC5A5/ACTIN) was quantified by ImageJ. E and F: the rescue effects of human NICD (Tol2:tg-NICD-2A-mCherry) on thyroid development are displayed. Vector control, embryos injected with Tol2:tg-mCherry. Whole mount RNA in situ hybridization (WISH) (E) and confocal imaging (F) were detected in 5 days postfertilization (dpf) embryos. G, left: under normal conditions, TAF1 could bind to the Notch1 promoter, promoting the activation of Notch signaling. HES1 transcription was then initiated. HES1 upregulated the expression of SLC5A5 and other genes associated with thyroid hormone biosynthesis. Furthermore, thyroid follicular cells can proliferate and mature. Right: the mutant TAF1 was incapable of binding to the Notch1 promoter and impaired Notch signaling. Inadequacy of Notch signaling led to the abnormal proliferation and maturation of thyroid follicular cells. This figure was created using Figdraw and PowerPoint. Scale bars: 100 μm. ***P < 0.001, ****P < 0.0001.