Expression Levels Of Yes-associated Protein Research Articles

A role for YAP/FOXM1/Nrf2 axis in oxidative stress and apoptosis of cataract induced by UVB irradiation.

This study aims to investigate the hypothesis that Yes-associated protein (YAP) significantly regulates antioxidant potential and anti-apoptosis in UVB-induced cataract by exploring the underlying molecular mechanisms. To investigate the association between YAP and cataract, various experimental techniques were employed, including cell viability assessment, Annexin V FITC/PI assay, measurement of ROS production, RT-PCR, Western blot assay, and Immunoprecipitation. UVB exposure on human lens epithelium cells (HLECs) reduced total and nuclear YAP protein expression, increased cleaved/pro-caspase 3 ratios, decreased cell viability, and elevated ROS levels compared to controls. Similar Western blot results were observed in invivo experiments involving UVB-treated mice. YAP knockdown invitro demonstrated a decrease in the protein expression of FOXM1, Nrf2, and HO-1, which correlated with the mRNA expression, accompanied by an increase in cell apoptosis, caspase 3 activation, and the release of ROS. Conversely, YAP overexpression mitigated these effects induced by UVB irradiation. Immunoprecipitation revealed a FOXM1-YAP interaction. Notably, inhibiting FOXM1 decreased Nrf2 and HO-1, activating caspase 3. Additionally, administering the ROS inhibitor N-acetyl-L-cysteine (NAC) effectively mitigated the apoptotic effects induced by oxidative stress from UVB irradiation, rescuing the protein expression levels of YAP, FOXM1, Nrf2, and HO-1. The initial findings of our study demonstrate the existence of a feedback loop involving YAP, FOXM1, Nrf2, and ROS that significantly influences the cell apoptosis in HLECs under UVB-induced oxidative stress.

Regulation of H9C2 cell hypertrophy by 14-3-3η via inhibiting glycolysis.

It has been reported that Ywhah (14-3-3η) reduces glycolysis. However, it remains unclear about the downstream mechanism by which glycolysis is regulated by 14-3-3η in cardiac hypertrophy. As an important regulator, Yes-associated protein (YAP) interacts with 14-3-3η to participate in the initiation and progression of various diseases in vivo. In this study, the model of H9C2 cardiomyocyte hypertrophy was established by triiodothyronine (T3) or rotenone stimulation to probe into the action mechanism of 14-3-3η. Interestingly, the overexpression of 14-3-3η attenuated T3 or rotenone induced cardiomyocyte hypertrophy and decreased glycolysis in H9C2 cardiomyocytes, whereas the knockdown of 14-3-3η had an opposite effect. Mechanistically, 14-3-3η can reduce the expression level of YAP and bind to it to reduce its nuclear translocation. In addition, changing YAP may affect the expression of lactate dehydrogenase A (LDHA), a glycolysis-related protein. Meanwhile, LDHA is also a possible target for 14-3-3η to mediate glycolysis based on changes in pyruvate, a substrate of LDHA. Collectively, 14-3-3η can suppress cardiomyocyte hypertrophy via decreasing the nucleus translocation of YAP and glycolysis, which indicates that 14-3-3η could be a promising target for inhibiting cardiac hypertrophy.

Suppression of YAP Ameliorates Cartilage Degeneration in Ankle Osteoarthritis via Modulation of the Wnt/β-Catenin Signaling Pathway.

Ankle osteoarthritis is a relatively understudied condition and the molecular mechanisms involved in its development are not well understood. This investigation aimed to explore the role and underlying molecular mechanisms of Yes-associated protein (YAP) in rat ankle osteoarthritis. The results demonstrated that YAP expression levels were abnormally increased in the ankle osteoarthritis cartilage model. In addition, knockdown of YAP expression was shown to hinder the imbalance in ECM metabolism induced by IL-1β in chondrocytes, as demonstrated by the regulation of matrix metalloproteinase (MMP)-3, MMP-9, and MMP-13, a disintegrin, metalloprotease with thrombospondin motifs, aggrecan, and collagen II expression. Additional studies revealed that downregulation of YAP expression markedly inhibited the overexpression of β-catenin stimulated by IL-1β. Furthermore, inhibition of the Wnt/β-catenin signaling pathway reversed the ECM metabolism imbalance caused by YAP overexpression in chondrocytes. It is important to note that the YAP-specific inhibitor verteporfin (VP) significantly delayed the progression of ankle osteoarthritis. In conclusion, the findings highlighted the crucial role of YAP as a regulator in modulating the progression of ankle osteoarthritis via the Wnt/β-catenin signaling pathway. These findings suggest that pharmacological inhibition of YAP can be an effective and critical therapeutic target for alleviating ankle osteoarthritis.

Generation of Anterior Segment of the Eye Cells from hiPSCs in Microfluidic Platforms.

Ophthalmic diseases affect many people, causing partial or total loss of vision and a reduced quality of life. The anterior segment of the eye accounts for nearly half of all visual impairment that can lead to blindness. Therefore, there is a growing demand for ocular research and regenerative medicine that specifically targets the anterior segment to improve vision quality. This study aims to generate a microfluidic platform for investigating the formation of the anterior segment of the eye derived from human induced pluripotent stem cells (hiPSC) under various spatial-mechanoresponsive conditions. Microfluidic platforms are developed to examine the effects of dynamic conditions on the generation of hiPSCs-derived ocular organoids. The differentiation protocol is validated, and mechanoresponsive genes are identified through transcriptomic analysis. Several culture strategies is implemented for the anterior segment of eye cells in a microfluidic chip. hiPSC-derived cells showed anterior eye cell characteristics in mRNA and protein expression levels under dynamic culture conditions. The expression levels of yes-associated protein and transcriptional coactivator PDZ binding motif (YAP/TAZ) and PIEZO1, varied depending on the differentiation and growth conditions of the cells, as well as the metabolomic profiles under dynamic culture conditions.

YAP/TAZ-TEAD activity promotes the malignant transformation of cervical intraepithelial neoplasia through enhancing the characteristics and Warburg effect of cancer stem cells

A number of studies have confirmed that Yes-associated protein (YAP)/transcriptional co-activator with PDZ-binding motif (TAZ)-transcriptional enhanced associate domain (TEAD) activity is the driver of cancer development. However, the role and mechanism of the YAP/TAZ-TEAD pathway in cervical intraepithelial neoplasia (CIN) remain to be clarified. Therefore, this study was designed to observe the effect of YAP/TAZ-TEAD activity on the development of CIN and provide new ideas for the diagnosis and treatment of CIN. Firstly, cervical tissues were collected from CIN patients in different stages [CIN grade 1 (CIN1) tissue, CIN grade 2/3 (CIN 2/3) and squamous cell carcinoma (SCC)] and healthy volunteers. Next, the expression levels of YAP, TAZ and TEAD in cervical tissues and cells were observed by immunohistochemistry, qRT-PCR and western blot. Besides, Z172 and Z183 cells were transfected with siRNA-YAP/TAZ (si-YAP/TAZ) and YAP/TAZ overexpression vector (YAP-5SA). Also, Z172 cells were co-transfected with YAP-5SA and si-TEAD2/4. Subsequently, the stemness characteristics, glycolysis level and malignant transformation of cells in each group were observed by sphere-formation assay, commercial kit, MTT, Transwell, scratch experiment, xenotransplantation and western blot.The expression of YAP, TAZ and TEAD increased significantly in cervical cancer tissue and cell line at the stage of CIN2/3 and SCC. When YAP/TAZ was knocked down, the stemness characteristics, glycolysis level and malignant transformation of cancer cells were notably inhibited; while activating YAP/TAZ exhibited a completely opposite result. In addition, activating YAP/TAZ and knocking down the TEAD expression at the same time significant weakened the effect of activated YAP/TAZ signal on precancerous cells and reduced inhibitory effect of knocking down TEAD alone. YAP/TAZ-TEAD signal activates the characteristics and Warburg effect of cancer stem cells, thereby promoting the malignant transformation of CIN.

Mechanism of resveratrol in protecting ovary in poor ovarian response mice by regulating Hippo signaling pathway

This study observed the protective effect of resveratrol(Res) on ovarian function in poor ovarian response(POR) mice by regulating the Hippo signaling pathway and explored the potential mechanism of Res in inhibiting ovarian cell apoptosis. Female mice with regular estrous cycles were randomly divided into a blank group, a model group, and low-and high-dose Res groups(20 and 40 mg·kg~(-1)), with 20 mice in each group. The blank group received an equal volume of 0.9% saline solution by gavage, while the model group and Res groups received suspension of glycosides of Triptergium wilfordii(GTW) at 50 mg·kg~(-1) by gavage for two weeks to induce the model. After modeling, the low-and high-dose Res groups were continuously treated with drugs by gavage for two weeks, while the blank group and the model group received an equal volume of 0.9% saline solution by gavage. Ovulation was induced in all groups on the day following the end of treatment. Finally, 12 female mice were randomly selected from each group, and the remaining eight female mice were co-housed with male mice at a ratio of 1∶1. Changes in the estrous cycle of mice were observed using vaginal cytology smears. The number of ovulated eggs, ovarian wet weight, ovarian index, and pregnancy rate of mice were measured. The le-vels of anti-Mullerian hormone(AMH), follicle-stimulating hormone(FSH), estradiol(E_2), and luteinizing hormone(LH) in serum were determined using enzyme-linked immunosorbent assay(ELISA). Ovarian tissue morphology and ovarian cell apoptosis were observed using hematoxylin-eosin(HE) staining and terminal deoxynucleotidyl transferase dUTP nick end labeling(TUNEL) staining, respectively. The protein expression levels of yes-associated protein(YAP) 1 and transcriptional coactivator with PDZ-binding motif(TAZ) were detected by immunohistochemistry(IHC), while the changes in protein expression levels of mammalian sterile 20-like kinase(MST) 1/2, large tumor suppressor(LATS) 1/2, YAP1, TAZ, B-cell lymphoma-2(Bcl-2), and Bcl-2 associated X protein(Bax) were determined by Western blot. The results showed that compared with the blank group, the model group had an increased rate of estrous cycle disruption in mice, a decreased number of normally developing ovarian follicles, an increased number of blocked ovarian follicles, increased ovarian granulosa cell apoptosis, decreased ovulation, reduced ovarian wet weight and ovarian index, increased serum FSH and LH levels, decreased AMH and E_2 levels, decreased protein expression levels of YAP1 and TAZ in ovarian tissues, increased relative expression levels of MST1/2, LATS1/2, and Bax proteins, and decreased relative expression levels of YAP1, TAZ, and Bcl-2 proteins. Additionally, the number of embryos per litter significantly decreased after co-housing. Compared with the model group, the low-and high-dose Res groups exhibited reduced estrous cycle disruption rates in mice, varying degrees of improvement in the number and morphology of ovarian follicles, reduced numbers of blocked ovarian follicles, improved ovarian granulosa cell apoptosis, increased ovulation, elevated ovarian wet weight and ovarian index, decreased serum FSH and LH levels, increased AMH and E_2 levels, elevated protein expression levels of YAP1 and TAZ in ovarian tissues, decreased relative expression levels of MST1/2, LATS1/2, and Bax proteins, and increased relative expression levels of YAP1, TAZ, and Bcl-2 proteins. Furthermore, the number of embryos per litter increased to varying degrees after co-housing. In conclusion, Res effectively inhibits ovarian cell apoptosis in mice and improves ovarian responsiveness. Its mechanism may be related to the regulation of key molecules in the Hippo pathway.

Syndecan-2 modulates the YAP pathway in epithelial-to-mesenchymal transition-related migration, invasion, and drug resistance in colorectal cancer

Epithelial-to-mesenchymal transition (EMT) is associated with an invasive phenotype in colorectal cancer (CRC). Here, we examined the roles of YES-associated protein (YAP) and syndecan-2 (SDC2) in EMT-related progression, invasion, metastasis, and drug resistance in CRC. The expression levels of YAP and SDC2 in CRC patient tumor tissue were quantified by PCR and western blotting. EMT-associated characteristics were assessed using Transwell assays and immunohistochemistry. Co-immunoprecipitation, glutathione S-transferase pull-down, and luciferase reporter assays were used to assess interactions between YAP and SDC2. YAP was found to be highly expressed in tumor tissue from 13/16 CRC patients, while SDC2 was highly expressed in the tumor tissue of 12/16 CRC patients. Overexpression of YAP in colon cancer cells led to increased cell viability, invasion, migration, and oxaliplatin resistance demonstrating that YAP plays a role in EMT. In addition, overexpression of YAP led to decreased expression of the large tumor suppressor kinase 1 (LATS1) and mammalian sterile 20-like kinases (MST1/2). Decreased LATS1 expression was associated with increased levels of cell proliferation. Knockdown of YAP by shRNA interference led to decreased cell invasion, migration, and drug resistance in colon cancer cells and reduced tumorigenesis in a mouse xenograft model. Finally, we established that YAP interacted with SDC2, and demonstrated that SDC2 mediated the YAP pathway through the EMT-related factors BMP4, CTGF and FOXM1.

YES-associated protein-regulated Smad7 worsen epithelial barrier injury of chronic sinusitis with nasal polyps.

Chronic rhinosinusitis with nasal polyps (CRSwNP) was potentially due to the epithelial barrier injury. YES-associated protein (YAP) is a multifunctional transcriptional factor and plays versatile roles in the regulation and maintenance of epithelial barrier in different organs and tissues. The purpose of this study is to elucidate possible effect and mechanism of YAP on the epithelial barrier of CRSwNP. Patients were divided into CRSwNP group (n = 12) and control group (n = 9). The location of YAP, PDZ-binding transcriptional co-activator (TAZ), and Smad7 were estimated by immunohistochemistry and immunofluorescence. Meanwhile, the expression of YAP, TAZ, Zona occludens-1 (ZO-1), E-cadherin, and transforming growth factor-beta1 (TGF-β1) were detected by Western blot. After primary human nasal epithelial cells were treated with YAP inhibitor, the expression level of YAP, TAZ, ZO-1, E-cadherin, TGF-β1, and Smad7 were measured by Western blot. Compared with the control group, the protein levels of YAP, TAZ, and Smad7 were significantly upregulated, while TGF-β1, ZO-1, and E-cadherin were downregulated in CRSwNP. YAP and Smad7 demonstrated lower levels, while the expression of ZO-1, E-cadherin, and TGF-β1 rose slightly after YAP inhibitor treatment in primary nasal epithelial cells. Higher level of YAP may lead to CRSwNP epithelial barrier injury via the TGF-β1 signaling pathway, and the inhibition of YAP can partially reverse epithelial barrier function.

YAP1 suppression inhibits autophagy and improves the efficacy of anti-PD-1 immunotherapy in hepatocellular carcinoma

Anti-PD-1 immunotherapy is a promising treatment for hepatocellular carcinoma (HCC), but some patients with HCC do not experience clinical benefits. Autophagy promotes tumor progression and participates in drug resistance. Previous studies have revealed that suppressing the expression level of Yes-associated protein 1 (YAP1) improves anti-PD-1 therapy efficacy. Therefore, the relationship between YAP1 expression and autophagy activity during anti-PD-1 treatment was investigated in this study. A positive correlation was found between the expression level of YAP1 and LC3B by analyzing The Cancer Genome Atlas (TCGA), UALCAN databases, and HCC tissue microarray. Meanwhile, YAP1 expression and autophagy constituted positive feedback, in which YAP1 inhibition decreased the autophagy activity in liver tumor cells by hepatocyte-specific Yap1 knockout mice. Further, anti-PD-1 treatment increased autophagy and YAP1 expression levels in the cancer tissues from DEN/TCPOBOP-induced liver cancer mice. Finally, Yap1 knockout suppressed autophagy and improved anti-PD-1 therapy efficacy in hepatocyte-specific Yap1 knockout mice with liver tumors. These results suggested that YAP1 suppression was sensitized to anti-PD-1 treatment and inhibited autophagy activity in liver tumor cells. YAP1 is a promising target for improving the efficacy of anti-PD-1 immunotherapy in HCC.

HIF-1α/YAP Signaling Rewrites Glucose/Iodine Metabolism Program to Promote Papillary Thyroid Cancer Progression.

Background: The management of aggressive and progressive metastatic papillary thyroid cancer (PTC) is very difficult. An inverse relationship between radioiodine and F-18 fluorodeoxyglucose (FDG) uptake (''flip-flop'' phenomenon) is described for invasive PTC during dedifferentiation. However, no satisfactory biologic explanation for this phenomenon. Hypoxia is an important microenvironmental factor that promotes cancer progression and glycolysis. The Hippo-YAP is a highly conserved tumor suppressor pathway and contributes to cancer metabolic reprogramming. Thus, we investigated the underlying molecular mechanisms of glucose/iodine metabolic reprogramming in PTC, focusing on the tumor hypoxia microenvironment and Hippo-YAP signaling. Methods: Immunohistochemistry staining was conducted to evaluate the expressions of hypoxia-inducible factor 1α (HIF-1α), yes-associated protein (YAP), glucose transporters 1 (GLUT1) and sodium iodine symporter (NIS) in matched PTC and the adjacent noncancerous tissues. PTC cell lines were cultured under normoxic (20% O2) and hypoxic (1% O2) conditions and the glycolysis level and NIS expression were measured. Further, we characterized the molecular mechanism of glucose/iodine metabolic reprogramming in PTC cell. Finally, we validated the results in vivo by establishing subcutaneous xenografts in nude mice. Results: The expression levels of HIF1-α, YAP and GLUT1 were upregulated in PTC tissues and YAP expression was positively associated with HIF-1α, GLUT1 and TNM stages. Meanwhile, the expression of NIS was negatively correlated with YAP. Further, in vitro studies indicated that hypoxia-induced YAP activation was critical for accelerating glycolysis and reducing NIS expression in PTC cells. Inhibition of YAP had the opposite effects in vitro and tumorigenicity in vivo. Hypoxia inhibited the Hippo signaling pathway resulting in the inactivation of YAP phosphorylation, further promoting the nuclear localization of YAP in PTC cells. The mechanism is that hypoxic stress promoted YAP binding to HIF-1α in the nucleus and maintained HIF-1α protein stability. The YAP/HIF-1α complex bound and directly activated the GLUT1 transcription to accelerate glycolysis. Meanwhile, HIF-1α/YAP signaling might indirectly reduce the expression of NIS by promoting the output of MAPK signaling. In vivo studies confirmed the YAP-mediated reprogramming of glucose/iodine metabolism promoted PTC progression. Conclusions: Collectively, our data revealed a novel regulatory mechanism of the glucose/iodine metabolic program rewritten by HIF-1α/YAP signaling in PTC. Inhibition of HIF-1α/YAP signaling alone or in combination with other potential markers may effectively combat aggressive PTC.

Wolfberry enhanced the abundance of Akkermansia muciniphila by YAP1 in mice with acetaminophen-induced liver injury.

Drug-induced liver injury (DILI) by acetaminophen (APAP) was one of the most challenging liver diseases. Wolfberry (Lycium barbarum L.), a traditional Chinese medicinal material and food supplement, has a potential effect on increasing the abundance of Akkermansia muciniphila (A. muciniphila) in mice colons. However, the effect and mechanism of wolfberry remain unclear in APAP-induced DILI. In this study, wolfberry promoted the proliferation of activated-A. muciniphila in vitro and in vivo. For the first time, we detected that the activated-A. muciniphila but not the killed-A. muciniphila increased the expression level of Yes-associated protein 1 (YAP1) in the liver and alleviated liver injury in APAP-induced DILI mice. Mechanically, A. muciniphila improved the intestinal mucosal barrier and reduced lipopolysaccharide (LPS) content in the liver, leading to the increased expression level of YAP1. Furthermore, wolfberry increased the A. muciniphila abundance in the colon and YAP1 expression in the liver from APAP-induced DILI mice, which promoted the recovery of APAP-induced liver injury. Meanwhile, wolfberry combination with A. muciniphila synergistically increased AKK abundance and YAP1 expression in the liver. Our research provides an innovative strategy to improve DILI.

Changes of YAP activity at the early stage of nonalcoholic steatohepatitis and its spatiotemporal relationship with ductular reaction in mice

To explore the changes in Yes-associated protein (YAP) activity at the early stage of nonalcoholic steatohepatitis (NASH) and the spatiotemporal relationship between YAP and ductular reaction (DR). Male C57BL/6J mouse models of NASH were established by feeding with a methionine- and choline-deficient (MCD) diet or a thioacetamide (TAA) diet for 12 weeks. At different time points during the feeding, liver histology of the mice was observed with HE and Masson trichrome staining. The mRNA expressions of YAP and its target genes (Ctgf, Cyr61, Acta2) were determined by qPCR, and the total protein expression level of YAP was measured with immunoblotting. The expression and distribution of YAP and the markers of DR (K19 and Sox9) were observed with immunohistochemical staining. At the early stage of NASH induced by MCD diet (1 to 4 weeks), the mRNA expression of YAP and its target genes and the total protein expression of YAP increased significantly (P < 0.01). The number of YAP-positive hepatocytes reached the peak level of 90.8 (cells per ×400 field of view) at week 2 and then decreased to 30.8 at week 4 (P < 0.001); YAP-positive ductular cells appeared near the portal area, where DR began to occur. From 8 to 12 weeks, numerous K19/Sox9-positive DR cells were observed in the hepatic lobules around the central vein (P < 0.01), while only a few YAP-positive hepatocytes were present in the liver tissue (P > 0.05), and the number of YAP-positive ductular cells gradually increased with time (P < 0.001). At the early stage of NASH induced by TAA diet (3 days to 2 weeks), the mRNA expression of YAP and its target genes and the total protein expression of YAP increased significantly (P < 0.05), and the number of YAP-positive hepatocytes reached the peak of 69.2 at week 2 and then decreased to 55.2 at week 4 (P < 0.001); YAP-positive ductular cells first appeared at the initial location of DR near the central vein. From 6 to 12 weeks, numerous K19/Sox9-positive DR cells occurred in the hepatic lobules around the central vein (P < 0.01). While the number of YAP-positive hepatocytes decreased (P < 0.001), the number of YAP-positive ductular cells continued to increase (P < 0.001). During the development of NASH, YAP activation occurs earlier than DR but they are spatiotemporally correlated. YAP activation in hepatocytes may participate in DR by promoting hepatocyte dedifferentiation.

INVESTIGATION OF THE POTENTIAL EFFECTS OF YAP-1 AND NESTIN ON RISK CLASSES AND PROGNOSIS IN NEUROBLASTOMA

Introduction: Neuroblastoma (NB) is the most common solid tumors in childhood. MYCN is the main prognostic factor for risk classification and staging in NB patients. YAP (Yes Associated Protein) plays a critical role in many types of cancer. In NB, YAP expression levels may be correlate with advanced tumor staging. Nestin is an important neuronal differentiation and cancer stem cell marker in NB. The aim of this study is to examine the potential roles of YAP-1 and Nestin expressions in terms of prognosis and risk classifications for neuroblastoma. Materials and Methods: Histological tumor sections of 55 cases were examined. YAP-1 and Nestin protein expression levels were evaluated by immunohistochemical staining on paraffin sections of different neuroblastoma risk groups. YAP-1 and Nestin expression levels were evaluated with clinical findings of NB patients. P value was &lt;0.05 accepted as a statistically significant. Results: 55 patient tissues were included in this study, of which 24 were male and 21 were female. The ages of the patients are between 1 month to 11 years. YAP-1 protein was expressed in 36.4% in MYCN(+)and 14.6% in MYCN(-) patients. Nestin protein was expressed 36.4% in MYCN(+), and 20.6% in MYCN(-) patients. YAP-1 was expressed 18% in all tissues. Nestin expressions were determined 20.4% in all tissues. YAP-1 was expressed in 26.9% in high-risk and 11.5% in low-risk groups. Nestin was expressed 24.4% in high-risk and 33.3% in low-risk groups. Conclusion: YAP-1 and Nestin might be a clinically risk markers for neuroblastoma patients.

FRK inhibits glioblastoma progression via phosphorylating YAP and inducing its ubiquitylation and degradation by Siah1.

We previously report that yes-associated protein (YAP), the core downstream effector of Hippo pathway, promotes the malignant progression of glioblastoma (GBM). However, although classical regulatory mechanisms of YAP are well explored, how YAP is modulated by the Hippo-independent manner remains poorly understood. Meanwhile, the nonreceptor tyrosine kinase Fyn-related kinase (FRK), which exhibits low expression and possesses tumor suppressor effects in GBM, is reported to be involved in regulation of protein phosphorylation. Here, we examined whether FRK could impede tumor progression by modulating YAP activities. Human GBM cells and intracranial GBM model were used to assess the effects of FRK and YAP on the malignant biological behaviors of GBM. Immunoblotting and immunohistochemistry were used to detect the expression of core proteins in GBM tissues. Co-immunoprecipitation, proximity ligation assay, luciferase assay and ubiquitination assay were utilized to determine the protein-protein interactions and related molecular mechanisms. The expression levels of FRK and YAP were inversely correlated with each other in glioma tissues. In addition, FRK promoted the ubiquitination and degradation of YAP, leading to tumor suppression in vitro and in vivo. Mechanistically, FRK interacted with and phosphorylated YAP on Tyr391/407/444, which recruited the classical E3 ubiquitin ligase Siah1 to catalyze ubiquitination and eventually degradation of YAP. Siah1 is required for YAP destabilization initiated by FRK. We identify a novel mechanism by which FRK orchestrates tumor-suppression effect through phosphorylating YAP and inducing its ubiquitination by Siah1. FRK-Siah1-YAP signaling axis may serve as a potential therapeutic target for GBM treatment.

Functional characteristics of YAP-positive hepatocytes expression in an early stage of NASH with transcriptome sequence analysis

Objective: To analyze and compare the differentially expressed genes (DEGs) of Yes-associated protein (YAP)-positive and negative hepatocytes and further understand the preliminary functional characteristics of YAP-positive hepatocytes in an early mouse model of nonalcoholic steatohepatitis (NASH) with transcriptome sequence (RNA-Seq). Methods: C57BL/6 mice were fed with methionine-choline deficiency (MCD) diet for 2 weeks to establish an early NASH model, and the control group was fed with normal diet. Liver tissue was stained with hematoxylin-eosin (HE) and Sirius red, and the pathological score was recorded. The expression of YAP and P-YAP were determined by immunohistochemistry (IHC) in liver tissues. Primary hepatocytes with viability greater than 90% were isolated and purified by collagenase perfusion combined with Percoll density gradient centrifugation. YAP-positive and negative hepatocytes were assessed by YAP antibody, flow cytometry and RNA-Seq analyses. Sequencing results were screened by GO, KEGG and interaction network analysis methods. RT-PCR was used to verify the expression levels of YAP and some DEGs in liver tissue model group. Two samples mean was compared by independent samples t-test. Results: Compared with the control group, the HE-stained liver tissue of MCD-induced mice at 2 weeks showed steatosis (pathological score 1.07±0.21), accompanied by lobular inflammation (pathological score 1.13±0.32) and ballooned hepatocyte (pathological score 0.80) ±0.20). Sirius red staining showed non-significant liver fibrosis (pathological score 0.40±0.40). IHC showed partial YAP-positive hepatocytes expression in an early stage of NASH. RNA-Seq analysis showed that clean reads of YAP-positive and negative hepatocytes were 49 310 604 and 5 4820 036, respectively. Compared with YAP-negative hepatocytes, YAP-positive hepatocytes had differential expression of 5 565 genes, including 1 662 up-regulated genes and 3 903 down-regulated genes. GO analysis of up-regulated genes showed that the metabolic processes related to mitochondrial functions, such as purine nucleoside triphosphate and nucleoside triphosphate were significantly enriched in biological processes (BP), while down-regulated gene analysis showed that olfactory-related receptor were significantly enriched in BP. KEGG analysis showed that DEGs were enriched in 292 pathways, and oxidative phosphorylation (OXPHOS) pathway was significantly enriched in signaling pathway. RT-PCR validated that inflammatory factors (interleukin-1β, interleukin-6), YAP and its target genes (Cyr61, Ankrd1), and Cox5b and Sdhc genes were significantly up-regulated in the OXPHOS pathway, which was consistent with the sequencing results. In addition, eight key genes with interaction network analysis were predicted. Conclusion: Changes in hepatocyte metabolic levels may be associated with increased YAP activity in an early stage of NASH.

Yes-associated protein promotes endothelial-to-mesenchymal transition of endothelial cells in choroidal neovascularization fibrosis.

To reveal whether and how Yes-associated protein (YAP) promotes the occurrence of subretinal fibrosis in age-related macular degeneration (AMD). Cobalt chloride (CoCl2) was used in primary human umbilical vein endothelial cells (HUVECs) to induce hypoxia in vitro. Eight-week-old male C57BL/6J mice weighing 19-25 g were used for a choroidal neovascularization (CNV) model induced by laser photocoagulation in vivo. Expression levels of YAP, phosphorylated YAP, mesenchymal markers [α smooth muscle actin (α-SMA), vimentin, and Snail], and endothelial cell markers (CD31 and zonula occludens 1) were measured by Western blotting, quantitative real-time PCR, and immunofluorescence microscopy. Small molecules YC-1 (Lificiguat, a specific inhibitor of hypoxia-inducible factor 1α), CA3 (CIL56, an inhibitor of YAP), and XMU-MP-1 (an inhibitor of Hippo kinase MST1/2, which activates YAP) were used to explore the underlying mechanism. CoCl2 increased expression of mesenchymal markers, decreased expression of endothelial cell markers, and enhanced the ability of primary HUVECs to proliferate and migrate. YC-1 suppressed hypoxia-induced endothelial-to-mesenchymal transition (EndMT). Moreover, hypoxia promoted total expression, inhibited phosphorylation, and enhanced the transcriptional activity of YAP. XMU-MP-1 enhanced hypoxia-induced EndMT, whereas CA3 elicited the opposite effect. Expression of YAP, α-SMA, and vimentin were upregulated in the laser-induced CNV model. However, silencing of YAP by vitreous injection of small interfering RNA targeting YAP could reverse these changes. The findings reveal a critical role of the hypoxia-inducible factor-1α (HIF-1α)/YAP signaling axis in EndMT and provide a new therapeutic target for treatment of subretinal fibrosis in AMD.

Expression of yes-associated protein, β-catenin and smoothened, and their clinical significance in invasive breast cancer.

The expression profile and role of yes-associated protein (YAP) in occurrence and development of breast cancer is ambiguous. The present study aimed to explore the relationship among the YAP, β-catenin and smoothened (SMO) signaling pathways to provide a theoretical basis for the clinical diagnosis and treatment of invasive breast cancer. Immunohistochemistry was used to determine the protein expression levels of YAP, β-catenin and SMO in tumor, tumor-adjacent and normal breast tissue. The possible association between the expression levels of these three proteins and the clinicopathological features of patients with breast cancer was then analyzed by the χ2 test. The protein expression of YAP was found to be downregulated, whilst β-catenin and SMO expression were found to be upregulated in tumor tissues as compared with that in normal breast tissues. In addition, the expression of YAP in breast cancer tissues was found to be associated with that of human epidermal growth factor receptor 2 (HER2), progesterone and estrogen receptors. By contrast, the protein expression of β-catenin and SMO in breast cancer tissues was only associated with HER2. There was a negative correlation between the expression of YAP and SMO protein in breast cancer tissues. Compared with that in the changes in each of YAP, β-catenin and SMO protein expression levels individually, their combined changes in expression were demonstrated to associate significantly with the tumor histological grade. To conclude, data from the present study suggest that the combined protein expression of YAP, β-catenin and SMO can be used as a prognostic indicator for the treatment of invasive breast cancer.

The Hippo Signaling Core Components YAP and TAZ as New Prognostic Factors in Lung Cancer

BackgroundThe Hippo pathway is an essential signaling cascade that regulates cell and organ growth. However, there is no consensus about (i) the expression levels of the Hippo signaling core components yes-associated protein (YAP) and transcriptional co-activator with PDZ-binding motif (TAZ) in lung cancer, especially in small cell lung cancer (SCLC), or (ii) their association with the prognosis of patients with SCLC.MethodsWe screened relevant articles and identified eligible studies in the PubMed, EMBASE, COCHRANE, and WanFang databases. A combined analysis was performed to investigate (i) the expression levels of the major effectors, YAP and TAZ, in lung cancer and its subsets and (ii) their prognostic role in lung cancer, especially in SCLC.ResultsIn total, 6 studies related to TAZ and 13 studies concerning YAP were enrolled in this meta-analysis. We found that high TAZ expression was significantly associated with poor overall survival (OS) of patients with non-small cell lung cancer (NSCLC) in the overall population [Ph < 0.001, crude hazard ratio (HR) = 1.629, 95% CI = 1.199–2.214 for TAZ expression; Ph = 0.029, adjusted HR = 2.127, 95% CI = 1.307–3.460 for TAZ], the Caucasian population (Ph = 0.043, crude HR = 1.233, 95% CI = 1.030–1.477 for TAZ expression), and the Asian population (Ph = 0.551, adjusted HR = 2.676, 95% CI = 1.798–3.982 for TAZ). Moreover, there was a significant negative association between YAP expression and an unsatisfactory survival of patients with lung cancer (Ph = 0.327, crude HR = 1.652, 95% CI = 1.211–2.253 for YAP expression) and patients with NSCLC [disease-free survival (DFS): Ph = 0.693, crude HR = 2.562, 95% CI = 1.876–3.499 for YAP expression; Ph = 0.920, crude HR = 2.617, 95% CI = 1.690–4.052 for YAP-mRNA; OS: Ph = 0.878, crude HR = 1.777, 95% CI = 1.233–2.562 for YAP expression], especially in the Asian population (DFS: Ph = 0.414, crude HR = 2.515, 95% CI = 1.755–3.063; OS: Ph = 0.712, crude HR = 1.772, 95% CI = 1.214–2.587). However, no association was observed in the multivariate combined analysis. High YAP expression was significantly associated with short OS of patients with SCLC in our combined multivariate analysis in the Asian population (Ph = 0.289, crude HR = 4.482, 95% CI = 2.182–9.209), but not with crude data (Ph = 0.033, crude HR = 1.654, 95% CI = 0.434–6.300).ConclusionThe Hippo pathway is involved in carcinogenesis and progression of NSCLC and SCLC, and high expression levels of YAP and TAZ are independent and novel prognostic factors for lung cancer.

Yes-associated protein gene overexpression regulated by β-catenin promotes gastric cancer cell tumorigenesis.

BACKGROUND: Yes-associated protein (YAP) has been reported to act as a candidate human oncogene and played a critical role in the development of multiple cancer types.OBJECTIVE: We aimed to investigate the expression, function, and underlying mechanisms of YAP in gastric cancer (GC).METHODS: Expression levels of YAP in gastric tissues were tested. CCK8 assay, clonogenic assay, apoptosis assay, transwell assay, cell scratch assay and animal study were conducted to explore the function of YAP. Chromatin immunoprecipitation (ChIP) assay and luciferase reporter assay were performed to explore the underlying mechanism. Survival analysis was carried out to reveal the relationship between YAP and clinical outcome.RESULTS: YAP was upregulated in gastric cancer tissues and correlates with poor prognosis. YAP could promote GC cells proliferation, metastatic capacity, inhibit GC cells apoptosis in vitro and in vivo. Both-catenin and YAP were mainly localized withi the tumor cell nuclei. -catenincould upregulate YAP expression by binding to the promotor region of YAP. Patients with both YAP and -catenin negetive expression had a better prognosis than others.CONCLUSIONS: YAP overexpression is driven by aberrant Wnt -catenin signalingand then contributed to the GC tumorigenesis and progression. Thus, YAP might be a potential target for GC treatment.

Silibinin inhibits the migration, invasion and epithelial-mesenchymal transition of prostate cancer by activating the autophagic degradation of YAP.

Silibinin (SB), a flavonoid extracted from milk thistle seeds, has been found to exert antitumor effects in numerous tumor types. Our previous study reported that SB had anti-metastatic effects in prostate cancer (PCa). However, the exact underlying molecular mechanisms remain to be determined. The present study aimed to investigate the effects of SB on the migration, invasion and epithelial-mesenchymal transition (EMT) of castration-resistant PCa (CRPC) cells using wound healing, Transwell assays, and western blotting. The results revealed that SB treatment significantly inhibited the migration and invasion of CRPC cell lines. Moreover, SB was confirmed to activate autophagy, as determined using LC3 conversion, LC3 turnover and LC3 puncta assays. Further mechanistic studies indicated that the expression levels of Yes-associated protein (YAP) were downregulated in an autophagy-dependent manner after SB treatment. In addition, the SB-induced autophagic degradation of YAP was associated with the anti-metastatic effects of SB in CRPC. In conclusion, the findings of the present study suggested that SB might inhibit the migration, invasion and EMT of PCa cells by regulating the autophagic degradation of YAP, thus representing a potential novel treatment strategy for metastatic CRPC.