Fig. 2.

TR binding to target genes leads to the recruitment of RNA Pol II and loss of core histones during T₃-induced metamorphosis. Tadpoles at stage 54 were treated with or without T₃ for 2 d, and the intestine was isolated for ChIP assay with the anti-TR (A), anti-Pol II (B), anti-H3 (C), anti-H2B (D), or anti-Id14 (extracellular protein, as a negative control) (E) antibody. The immunoprecipitated DNA was analyzed by qPCR for the presence of the TRE regions of the TRβ and TH/bZIP promoters. A region of exon 5 of the TRβ gene was analyzed as a negative control. The promoter region of the IFABP gene was also analyzed as a non-TR direct target gene control. Note that TR bound to both genes in the absence of T₃ in premetamorphic tadpoles. In the presence of T₃, TR binding to the TRE was increased, accompanied by the recruitment Pol II and reduction in histones at the TRE regions of both TH/bZIP and TRβ promoters. There was no TR binding to exon 5 in the presence or absence of T₃. However, increased Pol II was observed in the exon region due to increased transcription in the presence of T₃. Only background signals were observed with the anti-ID14 antibody. Error bars indicate sem (n = 3). Student's t test was carried out between pairs of control and T₃-treated groups. The one and two stars indicate pairs of samples with significant differences (P < 0.05 and P < 0.01, respectively).