Integrator complex subunit 15 controls mRNA splicing and is critical for eye development.

The eye and brain are composed of elaborately organized tissues, development of which is supported by spatiotemporally precise expression of a number of transcription factors and developmental regulators. Here we report the molecular and genetic characterization of Integrator complex subunit 15 (INTS15). INTS15 was identified in search for the causative gene(s) for an autosomal-dominant eye disease with variable individual manifestation found in a large pedigree. While homozygous Ints15 knockout mice are embryonic lethal, mutant mice lacking a small C-terminal region of Ints15 show ocular malformations similar to the human patients. INTS15 is highly expressed in the eye and brain during embryogenesis and stably interacts with the Integrator complex to support small nuclear RNA 3' end processing. Its knockdown resulted in missplicing of a large number of genes, probably as a secondary consequence, and substantially affected genes associated with eye and brain development. Moreover, studies using human iPS cells-derived neural progenitor cells showed that INTS15 is critical for axonal outgrowth in retinal ganglion cells. This study suggests a new link between general transcription machinery and a highly specific hereditary disease.

A global in vivo Drosophila RNAi screen identifies NOT3 as a conserved regulator of heart function.

Heart diseases are the most common causes of morbidity and death in humans. Using cardiac-specific RNAi-silencing in Drosophila, we knocked down 7061 evolutionarily conserved genes under conditions of stress. We present a first global roadmap of pathways potentially playing conserved roles in the cardiovascular system. One critical pathway identified was the CCR4-Not complex implicated in transcriptional and posttranscriptional regulatory mechanisms. Silencing of CCR4-Not components in adult Drosophila resulted in myofibrillar disarray and dilated cardiomyopathy. Heterozygous not3 knockout mice showed spontaneous impairment of cardiac contractility and increased susceptibility to heart failure. These heart defects were reversed via inhibition of HDACs, suggesting a mechanistic link to epigenetic chromatin remodeling. In humans, we show that a common NOT3 SNP correlates with altered cardiac QT intervals, a known cause of potentially lethal ventricular tachyarrhythmias. Thus, our functional genome-wide screen in Drosophila can identify candidates that directly translate into conserved mammalian genes involved in heart function.

Hippo-TAZ signaling is the master regulator of the onset of triple-negative basal-like breast cancers.

Breast cancer is the most frequent malignancy in women worldwide. Basal-like breast cancer (BLBC) is the most aggressive form of this disease, and patients have a poor prognosis. Here, we present data suggesting that the Hippo-transcriptional coactivator with PDZ-binding motif (TAZ) pathway is a key driver of BLBC onset and progression. Deletion of Mob1a/b in mouse mammary luminal epithelium induced rapid and highly reproducible mammary tumorigenesis that was dependent on TAZ but not yes-associated protein 1 (YAP1). In situ early-stage BLBC-like malignancies developed in mutant animals by 2 wk of age, and invasive BLBC appeared by 4 wk. In a human estrogen receptor+ luminal breast cancer cell line, TAZ hyperactivation skewed the features of these luminal cells to the basal phenotype, consistent with the aberrant TAZ activation frequently observed in human precancerous BLBC lesions. TP53 mutation is rare in human precancerous BLBC but frequent in invasive BLBC. Addition of Trp53 deficiency to our Mob1a/b-deficient mouse model enhanced tumor grade and accelerated cancer progression. Our work justifies targeting the Hippo-TAZ pathway as a therapy for human BLBC, and our mouse model represents a powerful tool for evaluating candidate agents.

Lethal Phenotype-Based Database Screening Identifies Ceramide as a Negative Regulator of Primitive Streak Formation.

In early embryogenesis, the primitive streak (PrS) generates the mesendoderm and is essential for organogenesis. However, because the PrS is a minute and transient tissue, elucidating the mechanism of its formation has been challenging. We performed comprehensive screening of 2 knockout mouse databases based on the fact that failure of PrS formation is lethal. We identified 812 genes involved in various cellular functions and responses that might be linked to PrS formation, with the category of greatest abundance being "Metabolism." In this study, we focused on genes of sphingolipid metabolism and investigated their roles in PrS formation using an in vitro mouse ES cell differentiation system. We show here that elevated intracellular ceramide negatively regulates gene expression essential for PrS formation and instead induces neurogenesis. In addition, sphingosine-1-phosphate (a ceramide derivative) positively regulates neural maturation. Our results indicate that ceramide regulates both PrS formation and the induction of neural differentiation.

CSE1L promotes nuclear accumulation of transcriptional coactivator TAZ and enhances invasiveness of human cancer cells.

The transcriptional coactivator with PDZ-binding motif (TAZ) (WWTR1) induces epithelial-mesenchymal transition and enhances drug resistance in multiple cancers. TAZ has been shown to interact with transcription factors in the nucleus, but when phosphorylated, translocates to the cytoplasm and is degraded through proteasomes. Here, we identified a compound TAZ inhibitor 4 (TI-4) that shifted TAZ localization to the cytoplasm independently of its phosphorylation. We used affinity beads to ascertain a putative target of TI-4, chromosomal segregation 1 like (CSE1L), which is known to be involved in the recycling of importin α and as a biomarker of cancer malignancy. We found that TI-4 suppressed TAZ-mediated transcription in a CSE1L-dependent manner. CSE1L overexpression increased nuclear levels of TAZ, whereas CSE1L silencing delayed its nuclear import. We also found via the in vitro coimmunoprecipitation experiments that TI-4 strengthened the interaction between CSE1L and importin α5 and blocked the binding of importin α5 to TAZ. WWTR1 silencing attenuated CSE1L-promoted colony formation, motility, and invasiveness of human lung cancer and glioblastoma cells. Conversely, CSE1L silencing blocked TAZ-promoted colony formation, motility, and invasiveness in human lung cancer and glioblastoma cells. In human cancer tissues, the expression level of CSE1L was found to correlate with nuclear levels of TAZ. These findings support that CSE1L promotes the nuclear accumulation of TAZ and enhances malignancy in cancer cells.

Physiological and pathological roles of the Hippo-YAP/TAZ signaling pathway in liver formation, homeostasis, and tumorigenesis.

The liver plays central homeostatic roles in metabolism and detoxification, and has a remarkable capacity to fully recover from injuries caused by the various insults to which it is constantly exposed. To fulfill these functions, the liver must maintain a specific size and so must regulate its cell numbers. It must also remove senescent, transformed, and/or injured cells that impair liver function and can lead to diseases such as cirrhosis and liver cancer. Despite their importance, however, the mechanisms governing liver size control and homeostasis have resisted delineation. The discovery of the Hippo intracellular signaling pathway and its downstream effectors, the transcriptional coactivators Yes-associated protein (YAP) and transcriptional coactivator with PDZ-binding motif (TAZ), has provided partial elucidation of these mechanisms. The Hippo-YAP/TAZ pathway is considered to be a cell's sensor of its immediate microenvironment and the cells that surround it, in that this pathway responds to changes in elements such as the ECM, cell-cell tension, and cell adhesion. Once triggered, Hippo signaling negatively regulates the binding of YAP/TAZ to transcription factors such as TEAD and Smad, controlling their ability to drive gene expression needed for cellular responses such as proliferation, survival, and stemness. Numerous KO mouse strains lacking YAP/TAZ, as well as transgenic mice showing YAP/TAZ hyperactivation, have been generated, and the effects of these mutations on liver development, size, regeneration, homeostasis, and tumorigenesis have been reported. In this review, I summarize the components and regulation of Hippo-YAP/TAZ signaling, and discuss this pathway in the context of liver physiology and pathology.

Protein kinase Cα activation switches YAP1 from TEAD-mediated signaling to p73-mediated signaling.

Yes-associated protein 1 (YAP1) interacts with TEAD transcription factor in the nucleus and upregulates TEAD-target genes. YAP1 is phosphorylated by large tumor suppressor (LATS) kinases, the core kinases of the Hippo pathway, at 5 serine residues and is sequestered and degraded in the cytoplasm. In human cancers with the dysfunction of the Hippo pathway, YAP1 becomes hyperactive and confers malignant properties to cancer cells. We have observed that cold shock induces protein kinase C (PKC)-mediated phosphorylation of YAP1. PKC phosphorylates YAP1 at 3 serine residues among LATS-mediate phosphorylation sites. Importantly, PKC activation recruits YAP1 to the cytoplasm even in LATS-depleted cancer cells and reduces the cooperation with TEAD. PKC activation induces promyelocytic leukemia protein-mediated SUMOylation of YAP1. SUMOylated YAP1 remains in the nucleus, binds to p73, and promotes p73-target gene transcription. Bryostatin, a natural anti-neoplastic reagent that activates PKC, induces YAP1/p73-mediated apoptosis in cancer cells. Bryostatin reverses malignant transformation caused by the depletion of LATS kinases. Therefore, bryostatin and other reagents that activate PKC are expected to control cancers with the dysfunction of the Hippo pathway.

MKK7 deficiency in mature neurons impairs parental behavior in mice.

c-Jun N-terminal kinases (JNKs) are constitutively activated in mammalian brains and are indispensable for their development and neural functions. MKK7 is an upstream activator of all JNKs. However, whether the common JNK signaling pathway regulates the brain's control of social behavior remains unclear. Here, we show that female mice in which Mkk7 is deleted specifically in mature neurons (Mkk7flox/flox Syn-Cre mice) give birth to a normal number of pups but fail to raise them due to a defect in pup retrieval. To explore the mechanism underlying this abnormality, we performed comprehensive behavioral tests. Mkk7flox/flox Syn-Cre mice showed normal locomotor functions and cognitive ability but exhibited depression-like behavior. cDNA microarray analysis of mutant brain revealed an altered gene expression pattern. Quantitative RT-PCR analysis demonstrated that mRNA expression levels of genes related to neural signaling pathways and a calcium channel were significantly different from controls. In addition, loss of neural MKK7 had unexpected regulatory effects on gene expression patterns in oligodendrocytes. These findings indicate that MKK7 has an important role in regulating the gene expression patterns responsible for promoting normal social behavior and staving off depression.

Characterization of mouse embryonic fibroblasts derived from Rassf6 knockout mice shows the implication of Rassf6 in the regulation of NF-κB signaling.

RASSF6 is a member of the tumor suppressor Ras association domain family (RASSF) proteins. We have reported using human cancer cell lines that RASSF6 induces apoptosis and cell cycle arrest via p53 and plays tumor suppressive roles. In this study, we generated Rassf6 knockout mice by CRISPR/Cas technology. Contrary to our expectation, Rassf6 knockout mice were apparently healthy. However, Rassf6-null mouse embryonic fibroblasts (MEF) were resistant against ultraviolet (UV)-induced apoptosis/cell cycle arrest and senescence. UV-induced p53-target gene expression was compromised, and DNA repair was delayed in Rassf6-null MEF. More importantly, KRAS active mutant promoted the colony formation of Rassf6-null MEF but not the wild-type MEF. RNA sequencing analysis showed that NF-κB signaling was enhanced in Rassf6-null MEF. Consistently, 7,12-dimethylbenz(a)anthracene (DMBA) induced skin inflammation in Rassf6 knockout mice more remarkably than in the wild-type mice. Hence, Rassf6 deficiency not only compromises p53 function but also enhances NF-κB signaling to lead to oncogenesis.

Hepatocyte Mitogen-Activated Protein Kinase Kinase 7 Contributes to Restoration of the Liver Parenchyma Following Injury in Mice.

BACKGROUND AND AIMS: Mitogen-activated protein kinase kinase (MKK) 7 and MKK4 are upstream activators of c-Jun NH2 -terminal kinases (JNKs) and have been shown to be required for the early development of the liver. Although it has been suggested that MKK7 might be involved in the regulation of hepatocyte proliferation, the functional role of MKK7 in the liver has remained unclear. APPROACH AND RESULTS: Here, we examined phenotypic alterations in liver-specific or hepatocyte/hematopoietic cell-specific MKK7 knockout (KO) mice, which were generated by crossing MKK7LoxP/LoxP with albumin-cyclization recombination (Alb-Cre) or myxovirus resistance protein 1-Cre mice, respectively. The livers of Alb-Cre-/+ MKK7LoxP/LoxP mice developed without discernible tissue disorganization. MKK7 KO mice responded normally to liver injuries incurred by partial hepatectomy or injection of CCl4 . However, tissue repair following CCl4 -induced injury was delayed in MKK7 KO mice compared with that of control mice. Furthermore, after repeated injections of CCl4 for 8 weeks, the liver in MKK7 KO mice showed intense fibrosis with increased protractive hepatocyte proliferation, suggesting that MKK7 deficiency might affect regenerative responses of hepatocytes in the altered tissue microenvironment. MKK7 KO hepatocytes demonstrated normal proliferative activity when cultured in monolayers. However, MKK7 KO significantly suppressed branching morphogenesis of hepatocyte aggregates within a collagen gel matrix. Microarray analyses revealed that suppression of branching morphogenesis in MKK7 KO hepatocytes was associated with a reduction in mRNA expression of transgelin, glioma pathogenesis related 2, and plasminogen activator urokinase-type (Plau); and forced expression of these genes in MKK7 KO hepatocytes partially recovered the attenuated morphogenesis. Furthermore, hepatocyte-specific overexpression of Plau rescued the impaired tissue repair of MKK7 KO mice following CCl4 -induced injury. CONCLUSIONS: MKK7 is dispensable for the regenerative proliferation of hepatocytes but plays important roles in repair processes following parenchymal destruction, possibly through modulation of hepatocyte-extracellular matrix interactions.

Heat shock induces the nuclear accumulation of YAP1 via SRC.

Yes-associated protein 1 (YAP1), a co-transcription activator, shuttles between the cytoplasm and the nucleus. Phosphorylation by large tumor suppressor kinases (LATS1/2) is the major determinant of YAP1 subcellular localization. Unphosphorylated YAP1 interacts with transcription factors in the nucleus and regulates gene transcription, while phosphorylated YAP1 is trapped in the cytoplasm and is degraded. We found that when U2OS and HeLa cells are exposed to 42 °C, YAP1 enters the nucleus within 30 min and returns to the cytoplasm at 4 h. SRC and HSP90 are involved in nuclear accumulation and return to the cytoplasm, respectively. Upon heat shock, LATS2 forms aggregates including protein phosphatase 1 and is dephosphorylated and inactivated. SRC activation is necessary for the formation of aggregates, while HSP90 is required for their dissociation. YAP1 is involved in heat shock-induced NF-κB signaling. Mechanistically, YAP1 is implicated in strengthening the interaction between RELA and DPF3, a component of SWI/SNF chromatin remodeling complex, in response to heat shock. Thus, YAP1 plays a role as a thermosensor.

The RAS-interacting chaperone UNC119 drives the RASSF6-MDM2-p53 axis and antagonizes RAS-mediated malignant transformation.

The gene encoding the proto-oncogene GTPase RAS is frequently mutated in human cancers. Mutated RAS proteins trigger antiapoptotic and cell-proliferative signals and lead to oncogenesis. However, RAS also induces apoptosis and senescence, which may contribute to the eradication of cells with RAS mutations. We previously reported that Ras association domain family member 6 (RASSF6) binds MDM2 and stabilizes the tumor suppressor p53 and that the active form of KRAS promotes the interaction between RASSF6 and MDM2. We also reported that Unc-119 lipid-binding chaperone (UNC119A), a chaperone of myristoylated proteins, interacts with RASSF6 and regulates RASSF6-mediated apoptosis. In this study, using several human cancer cell lines, quantitative RT-PCR, RNAi-based gene silencing, and immunoprecipitation/-fluorescence and cell biology assays, we report that UNC119A interacts with the active form of KRAS and that the C-terminal modification of KRAS is required for this interaction. We also noted that the hydrophobic pocket of UNC119A, which binds the myristoylated peptides, is not involved in the interaction. We observed that UNC119A promotes the binding of KRAS to RASSF6, enhances the interaction between RASSF6 and MDM2, and induces apoptosis. Conversely, UNC119A silencing promoted soft-agar colony formation, migration, and invasiveness in KRAS-mutated cancer cells. We conclude that UNC119A promotes KRAS-mediated p53-dependent apoptosis via RASSF6 and may play a tumor-suppressive role in cells with KRAS mutations.

Alantolactone is a natural product that potently inhibits YAP1/TAZ through promotion of reactive oxygen species accumulation.

Yes-associated protein 1 (YAP1) and its paralogue PDZ-binding motif (TAZ) play pivotal roles in cell proliferation, migration, and invasion, and abnormal activation of these TEAD transcriptional coactivators is found in diverse cancers in humans and mice. Targeting YAP1/TAZ signaling is thus a promising therapeutic avenue but, to date, few selective YAP1/TAZ inhibitors have been effective against cancer cells either in vitro or in vivo. We screened chemical libraries for potent YAP1/TAZ inhibitors using a highly sensitive luciferase reporter system to monitor YAP1/TAZ-TEAD transcriptional activity in cells. Among 29 049 low-molecular-weight compounds screened, we obtained nine hits, and the four of these that were the most effective shared a core structure with the natural product alantolactone (ALT). We also tested 16 other structural derivatives of ALT and found that natural ALT was the most efficient at increasing ROS-induced LATS kinase activities and thus YAP1/TAZ phosphorylation. Phosphorylated YAP1/TAZ proteins were subject to nuclear exclusion and proteosomic degradation such that the growth of ALT-treated tumor cells was inhibited both in vitro and in vivo. Our data show for the first time that ALT can be used to target the ROS-YAP pathway driving tumor cell growth and so could be a potent anticancer drug.

YAP drives cell competition by activating choline metabolism.

Cell competition is a phenomenon that eliminates unfit cells from cell society, a function vital for maintaining cellular and organismal homeostasis. We previously showed that Madin-Darby canine kidney (MDCK) epithelial cells expressing the active form of the transcriptional coactivator Yes-associated protein (YAP) are apically extruded when surrounded by normal MDCK cells. Although we demonstrated that the arachidonic acid (AA) cascade is involved in YAP-dependent apical extrusion, the metabolic events leading to this outcome remained unclear. Here, we present the results of metabolomic analysis that identified phosphatidylcholine (PC) biosynthesis as the most significant player in this process. Removal of the PC biosynthetic components choline and methionine from culture medium inhibited YAP-dependent apical extrusion. Inhibition of either choline uptake or metabolic cycles involving choline or methionine also decreased YAP-dependent apical extrusion. At the molecular level, active YAP induced expression of the genes encoding glycerophosphocholine phosphodiesterase 1 (GPCPD1) and lecithin-cholesterol acyltransferase (LCAT), which are involved in choline metabolism. Our results indicate that YAP-dependent cell competition depends on YAP-mediated activation of the choline metabolic cycle.

Loss of Mob1a/b in mice results in chondrodysplasia due to YAP1/TAZ-TEAD-dependent repression of SOX9.

Hippo signaling is modulated in response to cell density, external mechanical forces, and rigidity of the extracellular matrix (ECM). The Mps one binder kinase activator (MOB) adaptor proteins are core components of Hippo signaling and influence Yes-associated protein 1 (YAP1) and transcriptional co-activator with PDZ-binding motif (TAZ), which are potent transcriptional regulators. YAP1/TAZ are key contributors to cartilage and bone development but the molecular mechanisms by which the Hippo pathway controls chondrogenesis are largely unknown. Cartilage is rich in ECM and also subject to strong external forces - two upstream factors regulating Hippo signaling. Chondrogenesis and endochondral ossification are tightly controlled by growth factors, morphogens, hormones, and transcriptional factors that engage in crosstalk with Hippo-YAP1/TAZ signaling. Here, we generated tamoxifen-inducible, chondrocyte-specific Mob1a/b-deficient mice and show that hyperactivation of endogenous YAP1/TAZ impairs chondrocyte proliferation and differentiation/maturation, leading to chondrodysplasia. These defects were linked to suppression of SOX9, a master regulator of chondrogenesis, the expression of which is mediated by TEAD transcription factors. Our data indicate that a MOB1-dependent YAP1/TAZ-TEAD complex functions as a transcriptional repressor of SOX9 and thereby negatively regulates chondrogenesis.

Prostaglandin E2 and its receptor EP2 trigger signaling that contributes to YAP-mediated cell competition.

Cell competition is a biological process by which unfit cells are eliminated from "cell society." We previously showed that cultured mammalian epithelial Madin-Darby canine kidney (MDCK) cells expressing constitutively active YAP were eliminated by apical extrusion when surrounded by "normal" MDCK cells. However, the molecular mechanism underlying the elimination of active YAP-expressing cells was unknown. Here, we used high-throughput chemical compound screening to identify cyclooxygenase-2 (COX-2) as a key molecule triggering cell competition. Our work shows that COX-2-mediated PGE2 secretion engages its receptor EP2 on abnormal and nearby normal cells. This engagement of EP2 triggers downstream signaling via an adenylyl cyclase-cyclic AMP-PKA pathway that, in the presence of active YAP, induces E-cadherin internalization leading to apical extrusion. Thus, COX-2-induced PGE2 appears a warning signal to both abnormal and surrounding normal cells to drive cell competition.

YAP is essential for tissue tension to ensure vertebrate 3D body shape.

Vertebrates have a unique 3D body shape in which correct tissue and organ shape and alignment are essential for function. For example, vision requires the lens to be centred in the eye cup which must in turn be correctly positioned in the head. Tissue morphogenesis depends on force generation, force transmission through the tissue, and response of tissues and extracellular matrix to force. Although a century ago D'Arcy Thompson postulated that terrestrial animal body shapes are conditioned by gravity, there has been no animal model directly demonstrating how the aforementioned mechano-morphogenetic processes are coordinated to generate a body shape that withstands gravity. Here we report a unique medaka fish (Oryzias latipes) mutant, hirame (hir), which is sensitive to deformation by gravity. hir embryos display a markedly flattened body caused by mutation of YAP, a nuclear executor of Hippo signalling that regulates organ size. We show that actomyosin-mediated tissue tension is reduced in hir embryos, leading to tissue flattening and tissue misalignment, both of which contribute to body flattening. By analysing YAP function in 3D spheroids of human cells, we identify the Rho GTPase activating protein ARHGAP18 as an effector of YAP in controlling tissue tension. Together, these findings reveal a previously unrecognised function of YAP in regulating tissue shape and alignment required for proper 3D body shape. Understanding this morphogenetic function of YAP could facilitate the use of embryonic stem cells to generate complex organs requiring correct alignment of multiple tissues.

The Light-Inducible Genes Per2, Cry1a, and Cry2a Regulate Oxidative Status in Zebrafish.

The circadian clock is a highly conserved 24 h biological oscillation mechanism and is affected by environmental stimuli such as light, food and temperature. Disruption of the circadian clock results in disorders of diverse biological processes, including the sleep-wake cycle and metabolism. Although we previously identified several components of the circadian clock in zebrafish, our understanding of the relationship between light-inducible clock genes and metabolism remains incomplete. To investigate how light-inducible clock genes regulate metabolism, we performed transcriptomic and metabolomic analyses of the light-inducible clock genes zPer2, zCry1a, and zCry2a in zebrafish. Transcriptomic analysis of zPer2/zCry1a double knockout (DKO) and zPer2/zCry1a/zCry2a triple knockout (TKO) mutants showed that their gene expression profiles differed from that of wild type (WT) zebrafish. In particular, mRNA levels of zKeap1a, which encodes an oxidative stress sensor, were increased in DKO and TKO mutants. Metabolomic analysis showed genotype-dependent alteration of metabolomic profiles. Principal component analysis (PCA) and partial least squares-discriminant analysis (PLS-DA) showed the alteration of cysteine/methionine metabolism and glutathione metabolism. Specifically, cysteine and glutathione were decreased but methionine sulfoxide was increased in TKO zebrafish. These results indicate that the light-inducible genes zPer2, zCry1a, and zCry2a are involved in regulating the oxidative status of zebrafish.

Endogenous YAP1 activation drives immediate onset of cervical carcinoma in situ in mice.

Cervical cancer (CC) is usually initiated by infection with high-risk types of human papillomavirus (HPV). The HPV E6 and E7 proteins target p53 and RB, respectively, but other cellular targets likely exist. We generated uterus-specific MOB1A/B double KO (uMob1DKO) mice, which immediately developed cervical squamous cell carcinoma in situ. Mutant cervical epithelial cells showed YAP1-dependent hyperproliferation, altered self-renewal, impaired contact inhibition, and chromosomal instability. p53 activation was increased in uMob1DKO cells, and additional p53 loss in uMob1DKO mice accelerated tumor invasion. In human CC, strong YAP1 activation was observed from the precancerous stage. Human cells overexpressing HPV16 E6/E7 showed inactivation of not only p53 and RB but also PTPN14, boosting YAP1 activation. Estrogen, cigarette smoke condensate, and PI3K hyperactivation all increased YAP1 activity in human cervical epithelial cells, and PTPN14 depletion along with PI3K activation or estrogen treatment further enhanced YAP1. Thus, immediate CC onset may initiate when YAP1 activity exceeds an oncogenic threshold, making Hippo-YAP1 signaling a major CC driver.

Hippo pathway controls cell adhesion and context-dependent cell competition to influence skin engraftment efficiency.

Cell competition is involved in mammalian embryogenesis and tumor elimination and progression. It was previously shown that, whereas NIH3T3 mouse fibroblasts expressing high levels of the yes-associated protein 1(YAP1) target TEA domain family (TEAD) transcription factors become "winners" in cell competitions, Madin-Darby canine kidney cells expressing activated YAP1 become "losers" and are eliminated from culture monolayers. Thus, YAP1's role in cell competitions is clearly context dependent. Here, we show that keratinocytes overexpressing a constitutively activated YAP1 mutant lose in in vitro competitions with control cells conducted in standard tissue culture dishes and undergo apical extrusion. Similarly, cells in which endogenous YAP1 is activated by NF2 knockdown become losers. The YAP1-overexpressing cells exhibit a decrease in cell-matrix adhesion because of defective expression of adhesion molecules such as fibronectin-1. Cell adhesion-mediated proliferation is also impaired. However, because of intrinsic factors, YAP1-expressing cells proliferate faster than control cells when cocultured in dishes impeding cell adhesion. In vivo, Mob1a/b-deficient (YAP1-activated) epidermis, which shows decreased expression of type XVII collagen, cannot be engrafted successfully onto donor mice. YAP1-activated skin grafts shrink away from surrounding control skin, and the epidermis peels off the basement membrane. Our data show that YAP1 activation controls cell competition in part by decreasing cell adhesion.-Nishio, M., Miyachi, Y., Otani, J., Tane, S., Omori, H., Ueda, F., Togashi, H., Sasaki, T., Mak, T. W., Nakao, K., Fujita, Y., Nishina, H., Maehama, T., Suzuki, A. Hippo pathway controls cell adhesion and context-dependent cell competition to influence skin engraftment efficiency.