Fig. 5: Interface and phosphomotif conservation in Fat and Dachsous.

a Conservation analysis of the Fat4 and Dchs1 binding interface. The EC1-4 domains of each protein are depicted in surface representation and interacting residues are colored based on their conservation across 18 different Fat4 or Dchs1 homologs. b A multiple sequence alignment of the Fj phosphorylation motif (D-X-N-D-[X]₇-S/T) in EC3 of Fat4 and Dchs1 is shown as a sequence logo. The predicted phosphoserine residue is indicated with a red arrow. The motif is mostly conserved in Fat4 and Dchs1 homologs. c,d. SDS-PAGE analysis of Fat4(EC1–4), Dchs1(EC1–4), Ft(EC1–4), and Ds(EC1–4) proteins following incubation with Fj and FJX1. The gels were stained with Coomassie to monitor total protein (bottom panel, colored blue) and with a phosphostain (top panels, grayscale) to monitor phosphorylated protein levels. The Drosophila Ft and Ds proteins were robustly phosphorylated with Fj but not FJX1, and there was no detectable signal in the phosphomutants. Phosphorylation of human Fat4 was not detected following treatment with Fj or FJX1. Dchs1 was endogenously phosphorylated at low levels, and this phosphorylation signal was diminished in the phosphomutant. Unphosphorylated and phosphorylated protein ladders were used as controls. Asterisks indicate fragmented Dchs1 protein bands that reproducibly arise following treatment with Fj. SDS-PAGE analysis was performed three times using different batches of recombinant protein each time. Source data are provided as a Source Data file.