In vivo analysis of Yorkie phosphorylation sites

Oncogene. 2009 Apr 30;28(17):1916-27. doi: 10.1038/onc.2009.43. Epub 2009 Mar 30.

Abstract

The co-activator Yorkie (Yki) mediates transcriptional regulation effected by the Drosophila Fat-Warts (Wts)-Hippo (Hpo) pathways. Yki is inhibited by Wts-mediated phosphorylation, and a Wts phosphorylation site at Ser168 has been identified. Here we identify two additional Wts phosphorylation sites on Yki, and examine the respective contribution of all three sites to Yki nuclear localization and activity. Our results show that although Ser168 is the most critical site, all three phosphorylation sites influence Yki localization and activity in vivo, and can be sites of regulation by Wts. Thus, investigations of the role of Yki and its mammalian homolog Yes-associated protein (YAP) in development and oncogenesis should include evaluations of additional sites. The WW domains of Yki are not required for its phosphorylation, but instead are positively required for its activity. We also identify two potential sites of phosphorylation by an unknown kinase, which could influence phosphorylation of Ser168 by Wts, suggesting that there are additional mechanisms for regulating Yki/YAP activity.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Substitution
  • Animals
  • Animals, Genetically Modified
  • Binding Sites
  • Blotting, Western
  • Cell Line
  • Cell Nucleus / metabolism
  • Drosophila Proteins / genetics
  • Drosophila Proteins / metabolism*
  • Drosophila melanogaster / cytology
  • Drosophila melanogaster / genetics
  • Drosophila melanogaster / metabolism
  • Eye / cytology
  • Eye / metabolism
  • Female
  • Green Fluorescent Proteins / genetics
  • Green Fluorescent Proteins / metabolism
  • Immunoprecipitation
  • Intracellular Signaling Peptides and Proteins / genetics
  • Intracellular Signaling Peptides and Proteins / metabolism
  • Luciferases / genetics
  • Luciferases / metabolism
  • Male
  • Nuclear Proteins / genetics
  • Nuclear Proteins / metabolism*
  • Phosphorylation
  • Protein Binding
  • Protein Kinases / genetics
  • Protein Kinases / metabolism
  • Protein-Serine-Threonine Kinases / genetics
  • Protein-Serine-Threonine Kinases / metabolism
  • Recombinant Fusion Proteins / genetics
  • Recombinant Fusion Proteins / metabolism
  • Serine / genetics
  • Serine / metabolism
  • Trans-Activators / genetics
  • Trans-Activators / metabolism*
  • Wings, Animal / cytology
  • Wings, Animal / metabolism

Substances

  • Drosophila Proteins
  • Intracellular Signaling Peptides and Proteins
  • Nuclear Proteins
  • Recombinant Fusion Proteins
  • Trans-Activators
  • Yki protein, Drosophila
  • Green Fluorescent Proteins
  • Serine
  • Luciferases
  • Protein Kinases
  • wts protein, Drosophila
  • Protein-Serine-Threonine Kinases
  • hpo protein, Drosophila