Fig. 7. Point mutations in a hydrophobic rich region of the Slug N terminus prevents interaction with Ppa and stabilizes the protein. A) Alignment showing the conserved hydrophobic stretch of amino acids found in Slug proteins. laevis, Xenopus laevis; tropicalis; Xenopus tropicalis; gallus, Gallus gallus; rattus, Rattus rattus; musculus, Mus musculus; sapiens, Homo sapiens. (B) Wild-type Slug, or Slug in which target hydrophobic amino acids had been mutated (LY33, 34 VW58, 59 AA) were assayed in co-IP experiments for interaction with Ppa. Whereas Slug, Slug¹ and Slug² can efficiently bind Ppa (arrow), the combined mutant (Slug^1,2) no longer interacts (^* indicates IgG background band). (C) Embryos injected with wild-type Slug or Slug^1,2 were collected at the stages indicated. Western blots from injected embryo lysates demonstrate that Slug^1,2 is significantly more stable than is wild-type Slug.