Search

The Hippo signaling pathway restricts organ size by inactivating the Yorkie (Yki)/Yes-associated protein (YAP) family proteins. The oncogenic Yki/YAP transcriptional coactivator family promotes tissue growth by activating target gene transcription, but the regulation of Yki/YAP activation remains elusive. In mammalian cells, we identified Brg1, a major subunit of chromatin-remodeling SWI/SNF family proteins, which interacts with YAP. This finding led us to investigate the in vivo functional interaction of Yki and Brahma (Brm), the Drosophila homolog of Brg1. We found that Brm functions at the downstream of Hippo pathway and interacts with Yki and Scalloped (Sd) to promotes Yki-dependent transcription and tissue growth. Furthermore, we demonstrated that Brm is required for the Crumbs (Crb) dysregulation-induced Yki activation. Interestingly, we also found that crb is a downstream target of Yki–Brm complex. Brm physically binds to the promoter of crb and regulates its transcription through Yki. Together, we showed that Brm functions as a critical regulator of Hippo signaling during tissue growth and plays an important role in the feedback loop between Crb and Yki.

Tissue growth is achieved through coordinated cellular growth, cell division and apoptosis. Hippo signaling is critical for monitoring tissue growth during animal development. Loss of Hippo signaling leads to tissue overgrowth due to continuous cell proliferation and block of apoptosis. As cells lacking Hippo signaling are similar in size compared to normal cells, cellular growth must be properly maintained in Hippo signaling-deficient cells. However, it is not clear how Hippo signaling might regulate cellular growth. Here we show that loss of Hippo signaling increased Akt (also called Protein Kinase B, PKB) expression and activity, whereas activation of Hippo signaling reduced Akt expression in developing tissues in Drosophila. While yorkie (yki) is sufficient to increase Akt expression, Akt up-regulation caused by the loss of Hippo signaling is strongly dependent on yki, indicating that Hippo signaling negatively regulates Akt expression through Yki inhibition. Consistently, genetic analysis revealed that Akt plays a critical role in facilitating growth of Hippo signaling-defective tissues. Thus, Hippo signaling not only blocks cell division and promotes apoptosis, but also regulates cellular growth by inhibiting the Akt pathway activity.

The insulin/IGF signaling (IIS) pathway is a potent inducer of cell proliferation in normal development and in cancer. The mechanism by which this occurs, however, is not completely understood. The Hippo signaling pathway regulates cell proliferation via the transcriptional co-activator Yorkie/YAP, however the signaling inputs regulating Hippo activity are not fully elucidated. Here we present evidence linking these two conserved, oncogenic pathways in Drosophila and in mammalian cells. We find that activation of IIS and of Yorkie signaling correlate positively in hepatocellular carcinoma. We show that IIS activates Yorkie in vivo, and that Yorkie plays an important role in the ability of IIS to drive cell proliferation. Interestingly, we also find the converse—that Yorkie signaling activates components of the insulin/TOR pathway. In sum, this crosstalk between IIS and Yorkie leads to coordinated regulation of these two oncogenic pathways.

When cells undergo apoptosis, they can stimulate the proliferation of nearby cells, a process referred to as compensatory cell proliferation. The stimulation of proliferation in response to tissue damage or removal is also central to epimorphic regeneration. The Hippo signaling pathway has emerged as an important regulator of growth during normal development and oncogenesis from Drosophila to humans. Here we show that induction of apoptosis in the Drosophila wing imaginal disc stimulates activation of the Hippo pathway transcription factor Yorkie in surviving and nearby cells, and that Yorkie is required for the ability of the wing to regenerate after genetic ablation of the wing primordia. Induction of apoptosis activates Yorkie through the Jun kinase pathway, and direct activation of Jun kinase signaling also promotes Yorkie activation in the wing disc. We also show that depletion of neoplastic tumor suppressor genes, including lethal giant larvae and discs large, or activation of aPKC, activates Yorkie through Jun kinase signaling, and that Jun kinase activation is necessary, but not sufficient, for the disruption of apical-basal polarity associated with loss of lethal giant larvae. Our observations identify Jnk signaling as a modulator of Hippo pathway activity in wing imaginal discs, and implicate Yorkie activation in compensatory cell proliferation and disc regeneration.

During tissue regeneration, cell proliferation replaces missing structures to restore organ function. Regenerative potential differs greatly between organs and organisms; for example some amphibians can regrow entire limbs whereas mammals cannot. The process of regeneration relies on several signaling pathways that control developmental tissue growth, and implies the existence of organ size-control checkpoints that regulate both developmental, and regenerative, growth. Here we explore the role of one such checkpoint, the Salvador–Warts–Hippo pathway, in tissue regeneration. The Salvador–Warts–Hippo pathway limits tissue growth by repressing the Yorkie transcriptional co-activator. Several proteins serve as upstream modulators of this pathway including the atypical cadherins, Dachsous and Fat, whilst the atypical myosin, Dachs, functions downstream of Fat to activate Yorkie. Using Drosophila melanogaster imaginal discs we show that Salvador–Warts–Hippo pathway activity is repressed in regenerating tissue and that Yorkie is rate-limiting for regeneration of the developing wing. We show that regeneration is compromised in dachs mutant wing discs, but that proteins in addition to Fat and Dachs are likely to modulate Yorkie activity in regenerating cells. In conclusion our data reveal the importance of Yorkie hyperactivation for tissue regeneration and suggest that multiple upstream inputs, including Fat–Dachsous signaling, sense tissue damage and regulate Yorkie activity during regeneration of epithelial tissues.

The Fat-Hippo signaling pathway plays an important role in the regulation of normal organ growth during development, and in pathological growth during cancer. Fat-Hippo signaling controls growth through a transcriptional co-activator protein, Yorkie. A Fat-Hippo pathway has been described in which Yorkie is repressed by phosphorylation, mediated directly by the kinase Warts and indirectly by upstream tumor suppressors that promote Warts kinase activity. We present here evidence for an alternate pathway in which Yorkie activity is repressed by direct physical association with three other pathway components: Expanded, Hippo, and Warts. Each of these Yorkie repressors contains one or more PPXY sequence motifs, and associates with Yorkie via binding of these PPXY motifs to WW domains of Yorkie. This direct binding inhibits Yorkie activity independently from effects on Yorkie phosphorylation, and does so both in vivo and in cultured cell assays. These results emphasize the importance of the relative levels of Yorkie and its upstream tumor suppressors to Yorkie regulation, and suggest a dual repression model, in which upstream tumor suppressors can regulate Yorkie activity both by promoting Yorkie phosphorylation and by direct binding.