Angiogenesis and vascular remodeling are essential for the establishment of vascular networks during organogenesis. Here we show that the Hippo signaling pathway effectors YAP and TAZ are required, in a gene dosage-dependent manner, for the proliferation and migration of vascular endothelial cells (ECs) during retinal angiogenesis. Intriguingly, nuclear translocation of YAP and TAZ induced by Lats1/2-deletion blocked endothelial migration and phenocopied Yap/Taz-deficient mutants. Furthermore, overexpression of a cytoplasmic form of YAP (YAPS127D) partially rescued the migration defects caused by loss of YAP and TAZ function. Finally, we found that cytoplasmic YAP positively regulated the activity of the small GTPase CDC42, deletion of which caused severe defects in endothelial migration. These findings uncover a previously unrecognized role of cytoplasmic YAP/TAZ in promoting cell migration by activating CDC42 and provide insight into how Hippo signaling in ECs regulates angiogenesis.A ngiogenesis is a process of growth and remodeling in vascular networks that is essential for normal development. In adulthood, angiogenesis is activated as a reparative process, for example, during wound healing (1, 2). Aberrantly regulated angiogenesis can also be a component of disease (3) and can play a key role in tumor growth and metastasis (4), inflammatory diseases (5), diabetic retinopathy, and retinopathy of prematurity (6).Retinal angiogenesis in mice begins at postnatal day 0 (P0). The retinal vasculature initiates its expansion from the optic nerve head and migrates outwards along a preexisting network of astrocytes (7,8). This results in the formation of the superficial vascular plexus within the retinal ganglion cell layer during the first 8 d (9, 10). Endothelial cells (ECs) then migrate along nerve fibers to establish deep and intermediate vascular layers (9,11). Cell proliferation and migration are essential for angiogenesis and these cell responses are regulated by many different signaling pathways, including the VEGF, Notch, Wnt, FGF, BMP, and integrin signaling responses (9, 12-16). VEGFA and CDC42 are known to regulate extension of the angiogenic front and filopodia formation in angiogenic tip cells (2,17,18).The Hippo signaling pathway is an evolutionarily conserved, pivotal regulator of cell proliferation and organogenesis. YAP and TAZ are key components of the Hippo signaling pathway and function as transcription cofactors that regulate downstream gene expression via association with DNA binding proteins such as 20). Loss of Hippo signaling can drive the expression of genes that regulate cell proliferation and survival (diap1, bantam, cyclin E, and E2F1), the Hippo pathway (Kibra, Crb, and Fj), and cell-cell interaction (E-Cadherin, Serrate, Wingless, and Vein) (20). The activity of YAP and TAZ is regulated by the LATS1 and LATS2 kinases. These kinases phosphorylate YAP and TAZ, thus preventing their nuclear translocation and regulating transcriptional activity. Although the function of YAP and TAZ in the ...

Nonalcoholic steatohepatitis (NASH) is a progressive fibrotic disease, the pathogenesis of which has not been fully elucidated. Here, we report a molecular aspect of this disease elucidated using rabbits fed a cholesterol-rich high-fat diet and exhibiting insulin resistance. The liver in this model showed steatohepatitis with fibrosis and high mRNA expression for some cytokines, heme oxygenase-1, transforming growth factor-beta1, and collagen alpha1(I). Erythrocytes isolated from the model showed marked fragility and the externalization of phosphatidylserine (PS) on the outer leaflet of the membrane and were frequently engulfed by Kupffer cells/macrophages in the hepatic sinusoids. Expression of milk fat globule-epidermal growth factor (EGF)-factor 8, a PS-binding protein, was augmented in the liver. In culture, RAW 264.7 cells engulfed erythrocytes oxidized by tert-butyl hydroperoxide, a process that was inhibited by anti-milk fat globule-EGF-factor 8 antibody. In addition, PS-positive erythrocytes appeared entrapped in the model liver in ex vivo perfusion experiments. Finally, in specimens from NASH patients, the aggregation of erythrocytes in inflammatory hepatic sinusoids was notable. These results indicate that the engulfment of PS-externalized, apoptotic signal-positive, erythrocytes by hepatic macrophages may lead to the deposition of iron derived from hemoglobin in the liver and be involved in the pathogenesis of steatohepatitis.

Members of the transforming growth factor-β superfamily play essential roles in various aspects of embryonic development and physiological organ function. Among them, bone morphogenetic protein (BMP) 9 and BMP10 regulate embryonic vascular development by activating their endothelial receptor ALK1 (activin receptor-like kinase 1, also called Acvrl1). ALK1-mediated intracellular signaling is implicated in the etiologies of human diseases, but their downstream functional proteins are largely unknown. In this study, we identified Tmem100 , a gene encoding a previously uncharacterized intracellular transmembrane protein, to be an embryonic endothelium-enriched gene activated by BMP9 and BMP10 through the ALK1 receptor. Tmem100 null mice showed embryonic lethality due to impaired differentiation of arterial endothelium and defects of vascular morphogenesis, which phenocopied most of the vascular abnormalities observed with the Acvrl1/Alk1 deficiency. The activity of Notch- and Akt-mediated signaling, which is essential for vascular development, was down-regulated in Tmem100 null mice. Cre -mediated deletion of Tmem100 in endothelial cells was sufficient to recapitulate the null phenotypes. These data indicated that TMEM100 may play indispensable roles downstream of BMP9/BMP10-ALK1 signaling during endothelial differentiation and vascular morphogenesis.

Schwann cell (SC) myelination in the peripheral nervous system is essential for motor function, and uncontrolled SC proliferation occurs in cancer. Here, we show that a dual role for Hippo effectors TAZ and YAP in SC proliferation and myelination through modulating G-protein expression and interacting with SOX10, respectively. Developmentally regulated mutagenesis indicates that TAZ/YAP are critical for SC proliferation and differentiation in a stage-dependent manner. Genome-wide occupancy mapping and transcriptome profiling reveal that nuclear TAZ/YAP promote SC proliferation by activating cell cycle regulators, while targeting critical differentiation regulators in cooperation with SOX10 for myelination. We further identify that TAZ targets and represses Gnas, encoding Gαs-protein, which opposes TAZ/YAP activities to decelerate proliferation. Gnas deletion expands SC precursor pools and blocks peripheral myelination. Thus, the Hippo/TAZ/YAP and Gαs-protein feedback circuit functions as a fulcrum balancing SC proliferation and differentiation, providing insights into molecular programming of SC lineage progression and homeostasis.