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Additional Supporting Information may be found in the online version of this article.

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DVDY_22340_sm_suppfig-S1.tif15920KSupporting Figure S1: Tankyrase is expressed in ares of active Wnt signaling during embryonic development. A–H: In situ hybridization evaluating the expression of Tnks1 mRNA (A–D) or Tnks2 mRNA (E–H) in embryonic kidneys at embryonic day (E) 11.5 (A,E), newborn kidneys (B and F), E12.5 lungs (C,G), or E11.5 neural tube (D,H). The ureteric epithelium is labeled with an asterisk, while arrows label the metanephric mesenchyme.
DVDY_22340_sm_suppfig-S2.tif1760KSupporting Figure S2: IW molecules have a dosage-dependent effect on branching morphogenesis. A–M: Immunohistochemistry with antibodies against E-cadherin (green) and Laminin (red) evaluating branching morphogenesis in kidneys cultured in varying concentrations of IWR1 (A–E), IWP2 (F–I), or XAV939 (J–M). The presence of renal vesicles are indicated by arrows, while the absence of renal vesicles are indicated by arrowheads.
DVDY_22340_sm_suppfig-S3.tif14122KSupporting Figure S3: Evaluation of branching morphogenesis in cultured kidneys. A,B: Graphical representation of the number of branching tips in kidneys cultured in the presence of Tankyrase inhibitors (IWR1 and XAV939) or the diasteriomer control IWR1-exo (A) or in the presence of IWP2 or a related compound IWP7 (B). n = 15 kidneys from three independent experiments.
DVDY_22340_sm_suppfig-S4.tif17350KSupporting Figure S4: Tankyrase inhibition blocks branching morphogenesis in cultured lungs similar to Wnt inhibition. A–C: Immunohistochemistry with an antibody against E-cadherin (green) to evaluate branching morphogenesis in left lung buds cultured in dimethyl sulfoxide (DMSO; A), IWP2 (B), or IWR1 (C). D: Graphical representation of the quantification of lung branches after 48 hr of IW treatment. n = 12 lung buds from three independent experiments.
DVDY_22340_sm_suppfig-S5.tif1967KSupporting Figure S5: A 24-hr pulse of LiCl reinitiates kidney development after IW treatment. A–X: Evaluation of branching morphogenesis in kidneys cultured for 96 hr in dimethyl sulfoxide (DMSO; A–D,Q,U), 5 μM IWP2 (E–H,R,V), or sequentially for 24 hr in 5 μM IWP2 followed by 72 hr in 15 mM LiCl (I–L,S,W), or for 24 hr in 5 μM IWP2 followed by 24 hr in 15 mM LiCl and 48 hr in DMSO (M–P,T,X). A–P:Branching morphogenesis was visualized using HoxB7Cre;RosaYFP mice. Q–X:In situ hybridization evaluating the expression of Pax8 mRNA (Q–T) or Wnt11 mRNA (U–X) was performed on kidneys on treated explants upon completion of the 96 hr experiment. Results are representative of those found from three different experiments.
DVDY_22340_sm_suppfig-S6.tif1962KSupporting Figure S6: Activation of β-catenin is sufficient to rescue 48-hr IW treatment. A–X: Evaluation of branching morphogenesis in kidneys cultured for 96 hr in dimethyl sulfoxide (DMSO; A–D,Q,U), 5 μM IWP2 (E–H,R,V), or sequentially for 48 hr in 5 μM IWP2 followed by 48 hr in 15 mM LiCl (I–L,S,W) or for 48 hr in 5 μM IWP2 followed by 24 hr in 15 mM LiCl, and 24 hr in DMSO (M–P,T,X). A–P: Branching morphogenesis was analyzed every 24 hr using HoxB7Cre;RosaYFP mice. Q–X: In situ hybridization evaluating the expression of Pax8 mRNA (Q–T) or Wnt11 mRNA (U–X) was performed on kidneys on treated explants upon completion of the 96 hr experiment. Results are representative of those found from three different experiments.

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