Hippo signaling regulates Yorkie nuclear localization and activity through 14-3-3 dependent and independent mechanisms

The Hippo (Hpo) signaling pathway controls cell growth, proliferation and apoptosis in both Drosophila and vertebrates. In Drosophila, Hpo signaling regulates gene expression by inhibiting a transcription complex consisting of the transcriptional coactivator Yorkie (Yki) and the TEAD/TEF family of transcription factor Scalloped (Sd). Here we provide genetic evidence that both isoforms of 14-3-3, 14-3-3ɛ and 14-3-3ζ, regulate Yki activity through modulating its subcellular localization. Inactivation of 14-3-3 by RNAi or genetic mutations enhanced whereas overexpression of 14-3-3 suppressed tissue overgrowth induced by Yki overexpression. Loss of 14-3-3 resulted in the accumulation of Yki in the nucleus. We found that regulation of Yki by 14-3-3 was mediated by phosphorylation of Yki at S168. In addition, we found that Hpo signaling also inhibited Yki nuclear localization and activity by phosphorylating Yki at S111 and S250, and this inhibition appears to be independent of 14-3-3. Finally, we provided evidence that Hpo signaling restricted Yki nuclear localization depending on CRM1-mediated nuclear export.