Hippo signaling pathway

The Hippo signaling pathway, also known as the Salvador/Warts/Hippo (SWH) pathway, controls organ size in animals through the regulation of cell proliferation and apoptosis. The pathway takes its name from one of its key signaling components—the protein kinase Hippo (Hpo). Mutations in this gene lead to tissue overgrowth, or a “hippopotamus”-like phenotype.

A fundamental question in developmental biology is how an organ knows to stop growing after reaching a particular size. Organ growth relies on several processes occurring at the cellular level, including cell division and programmed cell death (or apoptosis). The Hippo signaling pathway is involved in restraining cell proliferation and promoting apoptosis. As many cancers are marked by unchecked cell division, this signaling pathway has become increasingly significant in the study of human cancer.

The Hippo signaling pathway appears to be highly conserved. While most of the Hippo pathway components were identified in the fruit fly (Drosophila melanogaster) using mosaic genetic screens, orthologs to these components (genes that function analogously in different species) have subsequently been found in mammals. Thus, the delineation of the pathway in Drosophila has helped to identify many genes that function as oncogenes or tumor suppressors in mammals.

Mechanism
The Hippo pathway consists of a core kinase cascade in which Hpo phosphorylates the protein kinase Warts (Wts). Hpo (MST1/2 in mammals) is a member of the Ste-20 family of protein kinases. This highly conserved group of serine/threonine kinases regulates several cellular processes, including cell proliferation, apoptosis, and various stress responses. Once phosphorylated, Wts (LATS1/2 in mammals) becomes active. Wts is a nuclear DBF-2-related kinase. These kinases are known regulators of cell cycle progression, growth, and development. Two proteins are known to facilitate the activation of Wts: Salvador (Sav) and Mob as tumor suppressor (Mats). Sav (WW45 in mammals) is a WW domain-containing protein, meaning that this protein contains a sequence of amino acids in which a [tryptophan] and an invariant proline are highly conserved. Hpo can bind to and phosphorylate Sav, which may function as a scaffold protein because this Hpo-Sav interaction promotes phosphorylation of Wts. Hpo can also phosphorylate and activate Mats (MOBKL1A/B in mammals), which allows Mats to associate with and strengthen the kinase activity of Wts.

Activated Wts can then go on to phosphorylate and inactivate the transcriptional coactivator Yorkie (Yki). Yki is unable to bind DNA by itself. In its active state, Yki binds to the transcription factor Scalloped (Sd), and the Yki-Sd complex becomes localized to the nucleus. This allows for the expression of several genes that promote organ growth, such as cyclin E, which promotes cell cycle progression, and diap1 (Drosophila inhibitor of apopotosis protein-1), which, as its name suggests, prevents apoptosis. Yki also activates expression of the bantam microRNA, a positive growth regulator that specifically affects cell number. Thus, the inactivation of Yki by Wts inhibits growth through the transcriptional repression of these pro-growth regulators. By phosphorylating Yki at serine 168, Wts promotes the association of Yki with 14-3-3 proteins, which help to anchor Yki in the cytoplasm and prevent its transport to the nucleus. In mammals, the two Yki orthologs are Yes-associated protein (YAP) and transcriptional coactivator with PDZ-binding motif (TAZ). When activated, YAP and TAZ can bind to several transcription factors including p73, Runx2 and several TEADs.

The upstream regulators of the core Hpo/Wts kinase cascade include the transmembrane protein Fat and several membrane-associated proteins. As an atypical cadherin, Fat (FAT1-4 in mammals) may function as a receptor, though an extracellular ligand has not been positively identified. While Fat is known to bind to another atypical cadherin, Dachsous (Ds), during tissue patterning, it is unclear what role Ds has in regulating tissue growth. Nevertheless, Fat is recognized as an upstream regulator of the Hpo pathway. Fat activates Hpo through the apical protein Expanded (Ex; FDM6 in mammals). Ex interacts with two other apically-localized proteins, Kibra (KIBRA in mammals) and Merlin (Mer; NF2 in mammals), to form the Kibra-Ex-Mer (KEM) complex. Both Ex and Mer are FERM domain-containing proteins, while Kibra, like Sav, is a WW domain-containing protein. The KEM complex physically interacts with the Hpo kinase cascade, thereby localizing the core kinase cacade to the plasma membrane for activation. Fat may also regulate Wts independently of Ex/Hpo, through the inhibition of the unconventional myosin Dachs. Normally, Dachs can bind to and promote the degradation of Wts.

The Hippo Signaling Pathway in Cancer
Many of the genes involved in the Hippo signaling pathway are recognized as tumor suppressors, while Yki/YAP/TAZ is identified as an oncogene. In fact, YAP has been found to be elevated in some human cancers, including breast cancer, colorectal cancer, and liver cancer. This may be explained by YAP’s recently defined role in overcoming contact inhibition, a fundamental growth control property of normal cells in culture in which proliferation stops after cells reach confluence. This property is typically lost in cancerous cells, allowing them to proliferate in an uncontrolled manner. In fact, YAP overexpression antagonizes contact inhibition.

Many of the pathway components recognized as tumor suppressor genes are mutated in human cancers. For example, mutations in Fat4 have been identified found in breast cancer, while NF2 is mutated in familial and sporadic schwanomas. Additionally, several human cancer cell lines invoke mutations of the WW45 and MOBK1B proteins.