Journal: Nature Communications

Article Title: Senescent cells perturb intestinal stem cell differentiation through Ptk7 induced noncanonical Wnt and YAP signaling

doi: 10.1038/s41467-022-35487-9

Figure Lengend Snippet: a Schematic representation of canonical and noncanonical Wnt signaling pathways. While canonical and noncanonical Wnt signaling pathways share components such as FZD and Dvl, other components such as LRP5/6 and β-Catenin are specific for canonical Wnt signaling. The figure was created with BioRender.com. b Representative images of organoids cultured in corresponding conditioned media with DMSO or inhibitors. Scale bar, 200 μm. c Quantification of data from ( b ). The percentage of cystic organoids in organoids cultured in conditioned media is shown. d Representative images of organoids treated with DMSO (carrier), Wnt3a or Wnt5a. Scale bar, 200 μm. e Quantification of data from ( d ). The percentage of cystic organoids in organoids cultured in conditioned media is shown. f Representative images of organoids cultured in Wnt5a with DMSO or inhibitors. Scale bar, 200 μm. g Quantification of data from (f). mean ± s.d. percentage of cystic organoids in organoids cultured in conditioned media. h Representative images of organoids cultured in Wnt5a with IgG or anti-Ptk7 antibodies. Scale bar, 200 μm. i Quantification of data from (h). mean ± s.d. The percentage of cystic organoids in organoids cultured in conditioned media is shown. In ( c ), ( e ), ( g ), and ( i ), data are presented as means ± s.d. One-way ANOVA followed by Tukey’s post-hoc multiple comparison tests. Each dot represents a result from organoids established from a mouse, n = 3. j Representative ELISA binding curves showing binding of Ptk7 or FZD7 to Wnt ligands. k Representative ELISA binding curves showing binding of Ptk7 to Wnt ligands in the absence or presence of 500 nM FZD7. In ( j ) and ( k ), data are presented as means ± s.d. n = 3 ( l ) Relative Ptk7 and FZD7 binding to Wnt ligands from ( k ). Relative Ptk7 binding was calculated by normalization to Ptk7 and Wnt ligand binding at 1500 nM in the absence of FZD7 (black striped bar). Relative FZD7 binding was calculated by normalization to FZD7 binding to Wnt ligands in the absence of Ptk7 (white striped bar). Data are presented as means ± s.d. See also Fig. S . Source data are provided as a Source Data file.

Article Snippet: Proteins used in ELISAs were obtained from R&D Systems: PTK7-Fc (R&D Systems, #9799-TK), biotinylated Wnt3a (R&D Systems, #BT1324), biotinylated Wnt5a (R&D Systems, #BT645).

Techniques: Protein-Protein interactions, Cell Culture, Comparison, Enzyme-linked Immunosorbent Assay, Binding Assay, Ligand Binding Assay