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Supplementary Figure S1. Expression and activation of YAP and other Hippo Pathway factors in CD4+ T cell subsets; Supplementary Figure S2. Characterization of the baseline immune profile of T cell-specific YAP-deficient mice; Supplementary Figure S3. Haematoxylin and eosin staining of lung, kidney, liver, small intestine and stomach sections from 21-day-old wild-type and YAPcKO mice; Supplementary Figure S4. The effect on iTreg generation under optimal TGFβ concentration; Supplementary Figure S5. Treg-specific YAP deficiency slows the growth of implanted MC38-colon tumors and boosts anti-tumor immunity; Supplementary Figure S6. Treg-specific YAP deficiency slows the growth of implanted EL4 thymomas and boosts anti-tumor immunity; Supplementary Figure S7. The effect of YAP inhibition monotherapy and combinational immunotherapy treatments on the immune constituents of the tumor microenvironment; Supplementary Figure S8. Correlation of gene expression in stimulated wild-type and YAP cKO derived Tregs; Supplementary Figure S9. Both Activin A and its receptor (ACVR1c) are upregulated during the course of iTreg differentiation; Supplementary Figure S10. Effect of AcVR1c deficiency on iTreg differentiation; Supplementary Figure S11. Supplemental Activin fails to rescue iTreg generation when SMAD2/3 levels are lacking; Supplementary Figure S12. Effects of GM-Vac and Anti-Activin treatment on B16 tumor progression in Wild Type and AcVR1c KO mice; Supplementary Figure S13. A model for YAP-mediated TGFβ/SMAD signaling enhancement

Regulatory T cells (Treg) are critical for maintaining self-tolerance and immune homeostasis, but their suppressive function can impede effective antitumor immune responses. FOXP3 is a transcription factor expressed in Tregs that is required for their function. However, the pathways and microenvironmental cues governing FOXP3 expression and Treg function are not completely understood. Herein, we report that YAP, a coactivator of the Hippo pathway, is highly expressed in Tregs and bolsters FOXP3 expression and Treg function in vitro and in vivo. This potentiation stemmed from YAP-dependent upregulation of activin signaling, which amplifies TGFβ/SMAD activation in Tregs. YAP deficiency resulted in dysfunctional Tregs unable to suppress antitumor immunity or promote tumor growth in mice. Chemical YAP antagonism and knockout or blockade of the YAP-regulated activin receptor similarly improved antitumor immunity. Thus, we identify YAP as an unexpected amplifier of a Treg-reinforcing pathway with significant potential as an anticancer immunotherapeutic target.Significance: Tregs suppress antitumor immunity, and pathways supporting their function can be novel immunotherapy targets. Here, the selective expression of YAP by Tregs, its importance for their function, and its unexpected enhancement of pro-Treg Activin/SMAD signaling are reported, as are validations of potential cancer-fighting antagonists of YAP and its regulatory targets. Cancer Discov; 8(8); 1026–43. ©2018 AACR.This article is highlighted in the In This Issue feature, p. 899

Regulatory T cells (Treg) are critical for maintaining self-tolerance and immune homeostasis, but their suppressive function can impede effective antitumor immune responses. FOXP3 is a transcription factor expressed in Tregs that is required for their function. However, the pathways and microenvironmental cues governing FOXP3 expression and Treg function are not completely understood. Herein, we report that YAP, a coactivator of the Hippo pathway, is highly expressed in Tregs and bolsters FOXP3 expression and Treg function in vitro and in vivo. This potentiation stemmed from YAP-dependent upregulation of activin signaling, which amplifies TGFβ/SMAD activation in Tregs. YAP deficiency resulted in dysfunctional Tregs unable to suppress antitumor immunity or promote tumor growth in mice. Chemical YAP antagonism and knockout or blockade of the YAP-regulated activin receptor similarly improved antitumor immunity. Thus, we identify YAP as an unexpected amplifier of a Treg-reinforcing pathway with significant potential as an anticancer immunotherapeutic target. Significance: Tregs suppress antitumor immunity, and pathways supporting their function can be novel immunotherapy targets. Here, the selective expression of YAP by Tregs, its importance for their function, and its unexpected enhancement of pro-Treg Activin/SMAD signaling are reported, as are validations of potential cancer-fighting antagonists of YAP and its regulatory targets. Cancer Discov; 8(8); 1026–43. ©2018 AACR. This article is highlighted in the In This Issue feature, p. 899

The regenerative capacity of Schwann cells in the PNS underlies functional repair after injury. In this study, Mindos et al. show a new function for the tumor suppressor Merlin and Hippo/YAP signaling in the generation of repair-competent Schwann cells after injury., Loss of the Merlin tumor suppressor and activation of the Hippo signaling pathway play major roles in the control of cell proliferation and tumorigenesis. We have identified completely novel roles for Merlin and the Hippo pathway effector Yes-associated protein (YAP) in the control of Schwann cell (SC) plasticity and peripheral nerve repair after injury. Injury to the peripheral nervous system (PNS) causes a dramatic shift in SC molecular phenotype and the generation of repair-competent SCs, which direct functional repair. We find that loss of Merlin in these cells causes a catastrophic failure of axonal regeneration and remyelination in the PNS. This effect is mediated by activation of YAP expression in Merlin-null SCs, and loss of YAP restores axonal regrowth and functional repair. This work identifies new mechanisms that control the regenerative potential of SCs and gives new insight into understanding the correct control of functional nerve repair in the PNS.

The Hippo pathway effector Yes-associated protein (YAP) regulates liver size by promoting cell proliferation and inhibiting apoptosis. However, recent in vivo studies suggest that YAP has important cellular functions other than controlling proliferation and apoptosis. Transgenic YAP expression in mouse hepatocytes results in severe jaundice. A possible explanation for the jaundice could be defects in adherens junctions that prevent bile from leaking into the blood stream. Indeed, immunostaining of E-cadherin and electron microscopic examination of bile canaliculi of Yap transgenic livers revealed abnormal adherens junction structures. Using primary hepatocytes from Yap transgenic livers and Yap knockout livers, we found that YAP antagonizes E-cadherin-mediated cell-cell junction assembly by regulating the cellular actin architecture, including its mechanical properties (elasticity and cortical tension). Mechanistically, we found that YAP promoted contractile actin structure formation by upregulating nonmuscle myosin light chain expression and cellular ATP generation. Thus, by modulating actomyosin organization, YAP may influence many actomyosin-dependent cellular characteristics, including adhesion, membrane protrusion, spreading, morphology, and cortical tension and elasticity, which in turn determine cell differentiation and tissue morphogenesis.

Cdc14 is an evolutionarily conserved serine/threoninephosphatase. Originally identified in S. cerevisiae as a cell cycle regulator, its role in other eukaryotic organisms remains unclear. In Drosophila melanogaster, Cdc14 is encoded by a single gene, thus facilitating its study. We found that Cdc14 expression is highest in the testis of adult flies and that cdc14 null flies are viable. cdc14 null female and male flies do not display altered fertility. cdc14 null males, however, exhibit decreased sperm competitiveness. Previous studies have shown that Cdc14 plays a role in ciliogenesis during zebrafish development. In Drosophila, sensory neurons are ciliated. We found that the Drosophila cdc14 null mutants have defects in chemosensation and mechanosensation as indicated by decreased avoidance of repellant substances and decreased response to touch. In addition, we show that cdc14 null mutants have defects in lipid metabolism and resistance to starvation. These studies highlight the diversity of Cdc14 function in eukaryotes despite its structural conservation.

SummaryAlthough Merlin/NF2 was discovered two decades ago as a tumor suppressor underlying Neurofibromatosis type II, its precise molecular mechanism remains poorly understood. Recent studies in Drosophila revealed a potential link between Merlin and the Hippo pathway by placing Merlin genetically upstream of the kinase Hpo/Mst. In contrast to the commonly depicted linear model of Merlin functioning through Hpo/Mst, here we show that in both Drosophila and mammals, Merlin promotes downstream Hippo signaling without activating the intrinsic kinase activity of Hpo/Mst. Instead, Merlin directly binds and recruits the effector kinase Wts/Lats to the plasma membrane. Membrane recruitment, in turn, promotes Wts phosphorylation by the Hpo-Sav kinase complex. We further show that disruption of the actin cytoskeleton promotes Merlin-Wts interactions, which implicates Merlin in actin-mediated regulation of Hippo signaling. Our findings elucidate an important molecular function of Merlin and highlight the plasma membrane as a critical subcellular compartment for Hippo signal transduction.

Yap1 is an important regulator of cardiomyocyte proliferation and embryonic heart development, yet the function of endogenous Yap1 in the adult heart remains unknown. We studied the role of Yap1 in maintaining basal cardiac function and in modulating injury after chronic myocardial infarction (MI). Cardiomyocyte-specific homozygous inactivation of Yap1 in the postnatal heart (Yap(F/F)Cre) elicited increased myocyte apoptosis and fibrosis, dilated cardiomyopathy, and premature death. Heterozygous deletion (Yap(+/F)Cre) did not cause an overt cardiac phenotype compared with Yap(F/F) control mice at base line. In response to stress (MI), nuclear Yap1 was found selectively in the border zone and not in the remote area of the heart. After chronic MI (28 days), Yap(+/F)Cre mice had significantly increased myocyte apoptosis and fibrosis, with attenuated compensatory cardiomyocyte hypertrophy, and further impaired function versus Yap(+/F) control mice. Studies in isolated cardiomyocytes demonstrated that Yap1 expression is sufficient to promote increased cell size and hypertrophic gene expression and protected cardiomyocytes against H(2)O(2)-induced cell death, whereas Yap1 depletion attenuated phenylephrine-induced hypertrophy and augmented apoptosis. Finally, we observed a significant decrease in cardiomyocyte proliferation in Yap(+/F)Cre hearts compared with Yap(+/F) controls after MI and demonstrated that Yap1 is sufficient to promote cardiomyocyte proliferation in isolated cardiomyocytes. Our findings suggest that Yap1 is critical for basal heart homeostasis and that Yap1 deficiency exacerbates injury in response to chronic MI.

SummaryCoordination of cell proliferation and cell death is essential to attain proper organ size during development and for maintaining tissue homeostasis throughout postnatal life. In Drosophila, these two processes are orchestrated by the Hippo kinase cascade, a growth-suppressive pathway that ultimately antagonizes the transcriptional coactivator Yorkie (Yki). Here we demonstrate that a single phosphorylation site in Yki mediates the growth-suppressive output of the Hippo pathway. Hippo-mediated phosphorylation inactivates Yki by excluding it from the nucleus, whereas loss of Hippo signaling leads to nuclear accumulation and therefore increased Yki activity. We further delineate a mammalian Hippo signaling pathway that culminates in the phosphorylation of YAP, the mammalian homolog of Yki. Using a conditional YAP transgenic mouse model, we demonstrate that the mammalian Hippo pathway is a potent regulator of organ size, and that its dysregulation leads to tumorigenesis. These results uncover a universal size-control mechanism in metazoan.

The Hippo signaling pathway converges on YAP to regulate growth, differentiation, and regeneration. Previous studies with overexpressed proteins have shown that YAP is phosphorylated by its upstream kinase, Lats1/2, on multiple sites, including an evolutionarily conserved 14-3-3-binding site whose phosphorylation is believed to inhibit YAP by excluding it from the nucleus. Indeed, nuclear localization of YAP or decreased YAP phosphorylation at this site (S168 in Drosophila, S127 in humans, and S112 in mice) is widely used in current literature as a surrogate of YAP activation even though the physiological importance of this phosphorylation event in regulating endogenous YAP activity has not been defined. Here we address this question by introducing a YapS112A knock-in mutation in the endogenous Yap locus. The YapS112A mice are surprisingly normal despite nuclear localization of the mutant YAP protein in vivo and profound defects in cytoplasmic translocation in vitro. Interestingly, the mutant YapS112A mice show a compensatory decrease in YAP protein levels due to increased phosphorylation at a mammalian-specific phosphodegron site on YAP. These findings reveal a robust homeostatic mechanism that maintains physiological levels of YAP activity and caution against the assumptive use of YAP localization alone as a surrogate of YAP activity.

Zinc finger proteins play important roles in various cellular functions, including cell proliferation, differentiation, and apoptosis. Mitogen-activated protein kinase (MAPK) signal transduction pathways are one of the most common mechanisms in eukaryotic cell regulation. Many transcription factors are important targets of MAPKs. In this study, we identified a novel gene encoding a zinc finger protein named ZNF445. The ZNF445 mRNA consists of 9105 nucleotides and has a 1031-amino acid open reading frame. The predicted 119-kDa protein contains a leucine-rich region (LER or SCAN domain) at the N-terminus, followed by a well-conserved Krüppel-associated box (KRAB) domain. At the C-terminus of the protein, there are 14 C2H2 (Cys2-His2) zinc finger motifs. ZNF445 gene is mapped to chromosome 3p21.32. Northern blot analysis indicates that a 9.1 kb transcript specific for ZNF445 is expressed in uterus, thymus, small intestine, colon, pancreas, peripheral blood leukocyte, and especially at a higher level in the testis and skeletal muscle in human adult tissues. ZNF445 protein was located in the nucleus when overexpressed in cultured cells. Reporter gene assays showed that ZNF445 is a transcriptional repressor, and overexpression of ZNF445 in the HEK 293T cells activates the transcriptional activities of AP1 and SRE. Deletion studies showed that the SCAN domain of ZNF445 may be involved in this activation. Furthermore, we found that expression of ZNF445 can increase p42/44 MAPK, MEK and Raf-1 phosphorylation. These results clearly indicate that ZNF445 is a member of the zinc finger transcription factor family and may function in MAPK pathway through Raf-1/MEK/p42/44 MAPK signals.

Background: The Hippo signaling pathway regulates organ size by restricting the activity of the transcriptional co-activator yes-associated protein (YAP), an oncogene implicated in pancreatic cancer. However, the role of YAP in mammalian pancreatic development and homeostasis is not well understood. Methods: Yap was conditionally ablated in the murine pancreas, either during development, using Pdx1-Cre as a “driver” or in the post-natal exocrine pancreas, using tamoxifen-inducible elastase (Ela)-Cre-ERT2 mice. Exocrine pancreatitis was induced by intraperitoneal caerulein administration. Results: Pdx1-Cre;Yapflox/flox mice demonstrated perinatal lethality (29/33 mice), secondary to a malformed pancreas, underscoring the importance of an intact Yap effector for pancreatic development. In wild-type mice, caerulein pancreatitis was associated with nuclear translocation of Yap protein, and upregulation of its bona fide transcriptional targets, survivin, and connective tissue growth factor (CTGF). Caerulein administration in tamoxifen-induced Ela-Cre-ERT2;Yapflox/flox mice resulted in markedly severe (necrotizing) pancreatitis, including the onset of mortality, compared to littermates with retained pancreatic Yap expression. Transcriptional profiling confirmed the failure to induce multiple anti-apoptotic molecules, including survivin, in the absence of Yap protein, which was accompanied by an increase in acinar cell apoptosis. However, neither exocrine regeneration itself, nor the upregulation of canonical signaling pathways implicated in regeneration (Notch and Hedgehog), was significantly abrogated in the absence of Yap. Conclusions: An intact Hippo pathway is essential for pancreatic morphogenesis. In the mature pancreas, absence of Yap profoundly exacerbates exocrine pancreatitis, with subsequent tissue regeneration remaining largely unaffected. Systemic therapeutics directed at negating YAP function in cancer cells need to be cognizant of the requirement for sustained YAP signaling in embryonic and mature tissues. Citation Format: Sonal Gupta, Anirban Maitra, Ji-Kon Ryu, Nailing Zhang, Seungmin Bang, Meritxell Rovira, Hanno Matthaei, Seung-Mo Hong, Steven D. Leach, Duojia Pan. Loss of the Hippo pathway effector Yap disrupts pancreatic morphogenesis and exacerbates exocrine pancreatitis. [abstract]. In: Proceedings of the AACR Special Conference on Pancreatic Cancer: Progress and Challenges; Jun 18-21, 2012; Lake Tahoe, NV. Philadelphia (PA): AACR; Cancer Res 2012;72(12 Suppl):Abstract nr B83.

Human chronic cholestatic liver diseases are characterized by cholangiocyte proliferation, hepatocyte injury, and fibrosis. Yes-associated protein (YAP), the effector of the Hippo tumor-suppressor pathway, has been shown to play a critical role in promoting cholangiocyte and hepatocyte proliferation and survival during embryonic liver development and hepatocellular carcinogenesis. Therefore, the aim of this study was to examine whether YAP participates in the regenerative response after cholestatic injury. First, we examined human liver tissue from patients with chronic cholestasis. We found more-active nuclear YAP in the bile ductular reactions of primary sclerosing cholangitis and primary biliary cirrhosis patient liver samples. Next, we used the murine bile duct ligation (BDL) model to induce cholestatic liver injury. We found significant changes in YAP activity after BDL in wild-type mice. The function of YAP in the hepatic response after BDL was further evaluated with liver-specific Yap conditional deletion in mice. Ablating Yap in the mouse liver not only compromised bile duct proliferation, but also enhanced hepatocyte necrosis and suppressed hepatocyte proliferation after BDL. Furthermore, primary hepatocytes and cholangiocytes isolated from Yap-deficient livers showed reduced proliferation in response to epidermal growth factor in vitro. Finally, we demonstrated that YAP likely mediates its biological effects through the modulation of Survivin expression. Conclusion: Our data suggest that YAP promotes cholangiocyte and hepatocyte proliferation and prevents parenchymal damage after cholestatic injury in mice and thus may mediate the response to cholestasis-induced human liver disease. (Hepatology 2012;56:1097–1107)

Although a developmental role for Hippo signaling in organ size control is well appreciated, how this pathway functions in tissue regeneration is largely unknown. Here we address this issue using a dextran sodium sulfate (DSS)-induced colonic regeneration model. We find that regenerating crypts express elevated Yes-associated protein (YAP) levels. Inactivation of YAP causes no obvious intestinal defects under normal homeostasis, but severely impairs DSS-induced intestinal regeneration. Conversely, hyperactivation of YAP results in widespread early-onset polyp formation following DSS treatment. Thus, the YAP oncoprotein must be exquisitely controlled in tissue regeneration to allow compensatory proliferation and prevent the intrinsic oncogenic potential of a tissue regeneration program.

Glioblastoma (GBM) remains fatal despite intensive treatments including standard-of-care surgery, radiation, and chemotherapy. Modern targeted therapies have also failed to produce effective results in clinical trials, possibly due to a lack of relevant preclinical models for drug discovery/assessment. Approximately 90% GBMs harbor aberrations in RTK/RAS-signaling, including EGFR amplification/gain-of-function mutations (57%), PDGFRa amplification (10%), and NF1 deletion/loss-of-function mutations (10%). KRAS mutations also occur, but at low frequency (1%). Frequent alteration of RTK/RAS-signaling emphasizes a great need for several distinct RTK/RAS-network-driven preclinical GBM models that mirror the range of human GBM lesions, since therapeutic responses may differ depending on the major network driver in a given tumor. We have recently generated a genetically-engineered mouse model of GBM via adult-inducible astrocyte-specific perturbation of RB, KRAS and PTEN. The triple mutant mice develop GBMs that faithfully recapitulate the histopathology and molecular characteristics of human primary GBMs. Interestingly, without RAS activation, only low-grade disease is induced. We established astrocytes from the triple mutant mouse model [RB-inactivated/KRASG12D/+/PTEN+/-; RKP astrocytes)] that elicit GBM upon intracranial transplantation, as well as the RB/PTEN double mutant mice [RB-inactivated/PTEN+/-; RP astrocytes], which do not produce GBM. Thus, the RP cells provide an ideal platform for evaluation of multiple RAS-activating events in causation of GBM. We are currently introducing the most frequent events found in human GBMs, including EGFR, ERFRvIII and PDGFRa activation and NF1 deficiency, into RP cells in vitro. The ability of each event to convert the RP cells into GBM-forming astrocytes by orthotopic transplantation is being assessed. This novel in vitro-to-allograft approach provides direct comparisons of etiologies driven by various RAS-activating events in the same genetic background and will potentially lead to basic discovery of distinct mechanisms in GBM tumorigenesis. Furthermore, this approach will generate relevant panels of RTK/RAS-network-driven GBM models for robust preclinical therapeutic discovery and biomarker development.

Local and distant invasion is a hallmark property of glioblastoma (GBM) that prevents curative surgical resection, and may be the key reason for GBM recurrence. Although cancer stem cells (CSCs) have been suggested to cause tumor recurrence based on their resistance to radiotherapy and chemotherapy, it is not clear whether CSCs are also involved in GBM invasiveness. We hypothesize that the invading cells in GBM have stem-like properties. To test our hypothesis, we propose to characterize cells at the invasive front of GBM using a syngeneic intracranial transplant tumor model. Three core regulatory networks are dysregulated in most human GBMs, including the RTK/RAS/PI3K/PTEN, INK/Cyclin D/CDK/RB, and ARF/p53 pathways. We have previously generated a genetically engineered mouse (GEM) model for GBM development via adult-inducible astrocyte-specific perturbation of RB, K-RAS and PTEN. These mice develop astrocytomas that progress to high grade GBM, and faithfully recapitulate the histopathology and molecular characteristics of most human GBMs. We have isolated primary tumor cells from these mice that are maintained in serum-free neural stem cell culture medium and expanded as neurospheres. These primary and low-passage cultures contain a mixed population of cancer stem cells and progenitor cells, and are highly invasive in vitro by matrigel invasion assay. With > 90% penetrance, intracranial injection of 25,000 cells leads to invasive GBM formation and a 32-day median survival in syngeneic host mice. Thus, the model provides an efficient approach for dissecting GBM cell invasion mechanisms in vivo. We will further trace the migration/invasion of GBM cells in the host brain after labeling with lentiviral-expressed mCherry. Host blood vessels are labeled with YFP via Tie2-Cre endothelial expression of a Rosa26-YFP allele. A time course analysis of stem cell marker expression will be used to assess properties of invading cells at the diffusive tumor margin. Laser capture microdissection will be used for invasive rim-cell and non-invasive core-cell isolation, and RNA samples will be compared by unbiased transcriptome studies. Results will provide hypotheses for invasion mechanisms and may identify potential biomarkers for invasive cells. Citation Format: Nailing Zhang, Yurong Song, Sanaz Jansen, Amit Adhikari, Sophie Wang, Lucy Lu, Teresa Sullivan, Terry Van Dyke. Characterization of the invading cells in glioblastoma using a syngeneic intracranial transplant tumor model. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 2626. doi:10.1158/1538-7445.AM2013-2626

Abstract 1592 The Yes-associated protein (YAP) is a transcriptional co-activator that interacts with many transcription factors, including RUNX2, p73, ERBB4, PEBP2α, p53BP2, SMAD7 and the TEAD/TEF proteins. Recently, it has been established that YAP is the major downstream molecule of the evolutionarily conserved Hippo signaling pathway. First elucidated in Drosophila, the Hippo signaling pathway controls organ size by regulating apoptosis and proliferation. Beginning a kinase cascade, the MST1/MST2 kinases (fly ortholog: Hippo) cooperate with the WW domain-containing SAV1 (fly ortholog: Salvador) to phosphorylate and activate the LATS1/LATS2 kinases (fly ortholog: Warts). In turn, activated LATS1/LATS2 phosphorylate and inactivate YAP (fly ortholog: Yorkie). Inactivated YAP binds to 14-3-3 and is kept in the cytosol. In contrast, unphosphorylated YAP binds to TEAD family transcription factors (fly ortholog: Scalloped) and the complex transactivates genes including cell cycle regulators (e.g. cyclin) and cell death inhibitors (e.g. IAPs, inhibitor of apoptosis). Therefore, inactivation of MST or LATS kinase, or overexpression of YAP results in organ/tissue overgrowth characterized by excessive cell proliferation and diminished apoptosis in both fly and mammals. In two transgenic mouse models, overexpression of YAP1 in liver results in reversible increase in liver size due to increased proliferation and decreased cell death of hepatocytes. Several pieces of evidence suggest that YAP regulates stem cell self-renewal and differentiation. First, transcriptional profiling of mouse stem cells (hematopoietic, neuronal and embryonic) identified both YAP1 and TEAD2 as 2 of the only 14 transcription factors commonly expressed in all 3 types of stem cell. Second, in many adult human tissues (e.g. intestine, lung, pancreas), YAP1 is preferentially expressed in stem-progenitor cell compartments. Third, in mouse intestine and chick neural tubes, overexpression of YAP results in expansion of intestinal or neural stem-progenitor cells, as well as inhibition of progenitor cell differentiation in vivo. Finally, it has been shown that YAP expression decreases during mouse ES cell differentiation. Ectopic expression of YAP can maintain ES cell pluripotency and prevent differentiation both in vitro and in vivo. Given that YAP has a proven role in regulating stem-progenitor cells in multiple tissue and multiple organisms, we set out to investigate whether YAP also regulate hematopoetic stem-progenitors in mouse and human. In conditional YAP knockout mice with a specific deletion of YAP expression in the hematopoetic system, we found no abnormalities at steady state in hemtaopoietic lineages, as assessed by CBC (complete blood counts) or immunophenotypic analysis. In addition, the YAP-null hematopietic stem-progenitors (HSPCs) had no change in vitro hematopoietic colony-forming cells. Furthermore, 5-fluorouracil treatment did not reveal a significant difference in blood cell numbers or types, between wild type and YAP-null mice. Although these results in mice suggest that YAP appears not to be required in hematopoiesis, overexpression of YAP, which promotes stem-progenitor cell proliferation in other tissues, may still provide an excellent opportunity to drive HSPC expansion. Disclosures: No relevant conflicts of interest to declare.