Supplementary Figure 6: Minibrain phosphorylates six amino acids in the Warts protein, including a DYRK1A consensus site.

(a and b) Wts1 (amino acids 1-318 of Wts) peptides that possessed amino acids that were phosphorylated when incubated with either wild-type Mnb or kinase-dead Mnb (Mnb-KD) immunoprecipitates. Experiments 1 and 2 were performed independently and analyzed by different mass spectrometry facilities. In experiment 1, five Wts amino acids were phosphorylated by wild-type Mnb with high confidence, whereas Mnb-KD induced no observable phosphorylation. In experiment 2, six Wts amino acids were phosphorylated by wild-type Mnb with high confidence, whilst two amino acids were phosphorylated in the presence of Mnb-KD. Amino acids that were phosphorylated by Mnb, but not Mnb-KD, are highlighted; amino acids that were phosphorylated in both independent experiments are red and amino acids that were phosphorylated in only one experiment are blue. (c) A summary of mass spectrometry data outlined in (a) and (b). The amino acid sequence of Wts1 is shown and the degree of conservation with human LATS1, as assessed by CLUSTAL, is indicated below: * indicates perfect conservation, : indicates strongly similar;. indicates weakly similar. As in (a) and (b), amino acids that were phosphorylated in both independent experiments by Mnb, but not Mnb-KD, are highlighted in red and amino acids that were phosphorylated by Mnb in only one experiment are in blue. The Wts sequence that was phosphorylated by Mnb and closely resembles a DYRK1A consensus site (RPXS/TP; ref. 2) is in bold font. Of note, this site was phosphorylated in each experiment and is conserved between Wts and LATS1.