Merlin, the protein product of the Neurofibromatosis type-2 gene, acts as a tumour suppressor in mice and humans. Merlin is an adaptor protein with a FERM domain and it is thought to transduce a growth-regulatory signal. However, the pathway through which Merlin acts as a tumour suppressor is poorly understood. Merlin, and its function as a negative regulator of growth, is conserved in Drosophila, where it functions with Expanded, a related FERM domain protein. Here, we show that Drosophila Merlin and Expanded are components of the Hippo signalling pathway, an emerging tumour-suppressor pathway. We find that Merlin and Expanded, similar to other components of the Hippo pathway, are required for proliferation arrest and apoptosis in developing imaginal discs. Our genetic and biochemical data place Merlin and Expanded upstream of Hippo and identify a pathway through which they act as tumour-suppressor genes.

Proliferation and apoptosis must be precisely regulated to form organs with appropriate cell numbers and to avoid tumour growth. Here we show that Hippo (Hpo), the Drosophila homologue of the mammalian Ste20-like kinases, MST1/2, promotes proper termination of cell proliferation and stimulates apoptosis during development. hpo mutant tissues are larger than normal because mutant cells continue to proliferate beyond normal tissue size and are resistant to apoptotic stimuli that usually eliminate extra cells. Hpo negatively regulates expression of Cyclin E to restrict cell proliferation, downregulates the Drosophila inhibitor of apoptosis protein DIAP1, and induces the proapoptotic gene head involution defective (hid) to promote apoptosis. The mutant phenotypes of hpo are similar to those of warts (wts), which encodes a serine/threonine kinase of the myotonic dystrophy protein kinase family, and salvador (sav), which encodes a WW domain protein that binds to Wts. We find that Sav binds to a regulatory domain of Hpo that is essential for its function, indicating that Hpo acts together with Sav and Wts in a signalling module that coordinately regulates cell proliferation and apoptosis.

Taken together, our data identify a cell-surface molecule that may act as a receptor of the Hippo signaling pathway.

Homeostatic mechanisms can eliminate abnormal cells to prevent diseases such as cancer. However, the underlying mechanisms of this surveillance are poorly understood. Here we investigated how clones of cells mutant for the neoplastic tumor suppressor gene scribble ( scrib ) are eliminated from Drosophila imaginal discs. When all cells in imaginal discs are mutant for scrib, they hyperactivate the Hippo pathway effector Yorkie (Yki), which drives growth of the discs into large neoplastic masses. Strikingly, when discs also contain normal cells, the scrib − cells do not overproliferate and eventually undergo apoptosis through JNK-dependent mechanisms. However, induction of apoptosis does not explain how scrib − cells are prevented from overproliferating. We report that cell competition between scrib − and wild-type cells prevents hyperproliferation by suppressing Yki activity in scrib − cells. Suppressing Yki activation is critical for scrib − clone elimination by cell competition, and experimental elevation of Yki activity in scrib − cells is sufficient to fuel their neoplastic growth. Thus, cell competition acts as a tumor-suppressing mechanism by regulating the Hippo pathway in scrib − cells.

During animal development, organ size is determined primarily by the amount of cell proliferation, which must be tightly regulated to ensure the generation of properly proportioned organs. However, little is known about the molecular pathways that direct cells to stop proliferating when an organ has attained its proper size. We have identified mutations in a novel gene, shar-pei, that is required for proper termination of cell proliferation during Drosophila imaginal disc development. Clones of shar-pei mutant cells in imaginal discs produce enlarged tissues containing more cells of normal size. We show that this phenotype is the result of both increased cell proliferation and reduced apoptosis. Hence, shar-pei restricts cell proliferation and promotes apoptosis. By contrast, shar-pei is not required for cell differentiation and pattern formation of adult tissue. Shar-pei is also not required for cell cycle exit during terminal differentiation, indicating that the mechanisms directing cell proliferation arrest during organ growth are distinct from those directing cell cycle exit during terminal differentiation. shar-pei encodes a WW-domain-containing protein that has homologs in worms, mice and humans, suggesting that mechanisms of organ growth control are evolutionarily conserved. and Ripoll, 1975). Notably, after such manipulation of proliferation rates, the final pattern and size of the adult structures are normal. Moreover, discs can regenerate missing parts after surgical manipulation (Bryant, 1978; Bryant and Simpson, 1984) and when ~75% of the progenitor cells of imaginal discs are killed by X-rays, the remaining cells proliferate and compensate for the loss of cells (Haynie and Bryant, 1977). Hence, cell proliferation is plastic and cells in a developing tissue adjust their proliferation depending on whether more cells are needed to build a normal sized structure (Day and Lawrence, 2000;French et al., 1976;Garcia-Bellido and Garcia-Bellido, 1998). However, the molecular mechanisms that direct cells to stop proliferating once the primordium of a structure has reached the correct size are poorly understood.In principle, defined organ size can be generated either by regulating the extent of cell proliferation or by eliminating superfluous cells through programmed cell death, or both. Only limited amounts of cell death are observed during imaginal disc growth (Milan et al., 1997;Wolff and Ready, 1991), indicating that disc size is primarily, albeit not exclusively, controlled at the level of cell division. Thus, factors must exist that regulate the decision of imaginal disc cells to re-enter or exit the cell cycle to mediate growth control.The Drosophila eye is particularly well suited to identify factors that regulate cell proliferation. First, the various stages of cell division and differentiation can be accurately followed in eye imaginal discs. Second, defects in growth control and differentiation can be easily scored. In the early growth phase of the eye disc, cell cycles are not synchronized and pro...