Signaling through the Hippo pathway, which regulates organ size and cell proliferation in both the fruit fly Drosophila melanogaster and in mammals, culminates in regulated activity of the transcriptional coactivator Yorkie (Yki). The FERM domain proteins Merlin (Mer) and Expanded (Ex) modulate the activity of the kinase Hippo (Hpo), Hpo regulates the activity of the kinase Warts (Wts), and activated Wts phosphorylates Yki to promote retention of Yki in the cytoplasm. In addition to the Wts phosphorylation sites that repress Yki activity, Yki also contains two WW domains that enhance Yki’s ability to promote transcription of target genes, and Oh et al. report that these WW domains may also mediate a phosphorylation-independent mechanism for repressing Yki. Expression of a transgene encoding a mutant form of Yki lacking all three Wts phosphorylation sites (Yki:V53SA) in cells of the fly eye disc resulted in an overgrowth phenotype that could be partially suppressed by coexpression of transgenes encoding Hpo, Wts, and Ex. The presence of Wts, Hpo, and Ex reduced nuclear accumulation of Yki:V53SA in eye discs and reduced the ability of Yki:V53SA to activate expression of a reporter gene in cultured S2 cells. Transgenically encoded, tagged forms of Wts, Hpo, and Ex coimmunoprecipitated with both wild-type Yki and Yki:V53SA from S2 cell extracts but did not coimmunoprecipitate with a form of Yki in which the WW domains were mutated (Yki:V5-WW). In wts loss-of-function clones in the wing disc, a portion of endogenous Yki colocalized with endogenous Ex, suggesting that Yki and Ex can interact under physiological conditions, at least when Yki repression by Wts is eliminated. Although it is not clear whether this phosphorylation-independent mechanism for repressing Yki occurs in vivo, it is an enticing possibility that Wts and other Hippo signaling pathway components may repress Yki activity through more than one mechanism.
H. Oh, B. V. Reddy, K. D. Irvine, Phosphorylation-independent repression of Yorkie in Fat-Hippo signaling. Dev. Biol. 335, 188–917 (2009). [PubMed]