Involvement of Tead4, an effector of Hippo signaling, in intestinal remodeling during Xenopus laevis metamorphosis

During amphibian metamorphosis, the intestine is remodeled from larval to adult form. In this study, we investigated the expression and localization of Xenopus laevis Tead4 (a key transcription factor in the Hippo pathway) in the metamorphosing intestine, examined its interaction, and analyzed its phylogenetic relationships. Quantitative RT-PCR revealed that both tead4.L and tead4.S (homeologs) transcripts are expressed throughout natural and thyroid hormone (TH)-induced metamorphosis, with modest but stage-specific changes, indicating sensitivity to TH. Triple hybridization chain reaction RNA-FISH showed that tead4 expression is broadly distributed in the intestine, with its signal intensity increasing from the metamorphic climax onward, whereas yap1 expression is enriched in the connective and muscle layers. There was a partial overlap between tead4 and yap1 expression, indicating context-dependent co-regulation, and incomplete co-localization of the expressed proteins, suggesting alternative co-factors. Oocyte co-immunoprecipitation assays demonstrated that both Tead4.L and Tead4.S physically associate with Yap1, supporting their role as Hippo pathway effectors. Phylogenetic analysis placed X. laevis Tead4 homeologs within the TEAD2 clade, indicating that the genes annotated as tead4 can more accurately be classified as tead2. These findings demonstrate that Tead4 homeologs function as Yap1 partners and contribute to transcriptional control during amphibian intestinal metamorphosis, highlighting their conserved role as effectors of Hippo signaling.