Allosteric Modulation of the YAP/TAZ-TEAD Interaction by Palmitoylation and Small-Molecule Inhibitors.

The Hippo signaling pathway is a highly conserved signaling network that plays a central role in regulating cellular growth, proliferation, and organ size. This pathway consists of a kinase cascade that integrates various upstream signals to control the activation or inactivation of YAP/TAZ proteins. Phosphorylated YAP/TAZ is sequestered in the cytoplasm; however, when the Hippo pathway is deactivated, it translocates into the nucleus, where it associates with TEAD transcription factors. This partnership is instrumental in regulating the transcription of progrowth and antiapoptotic genes. Thus, in many cancers, aberrantly hyperactivated YAP/TAZ promotes oncogenesis by contributing to cancer cell proliferation, metastasis, and therapy resistance. Because YAP and TAZ exert their oncogenic effects by binding with TEAD, it is critical to understand this key interaction to develop cancer therapeutics. Previous research has indicated that TEAD undergoes autopalmitoylation at a conserved cysteine, and small molecules that inhibit TEAD palmitoylation disrupt effective YAP/TAZ binding. However, how exactly palmitoylation contributes to YAP/TAZ-TEAD interactions and how the TEAD palmitoylation inhibitors disrupt this interaction remains unknown. Utilizing molecular dynamics simulations, our investigation not only provides detailed atomistic insight into the YAP/TAZ-TEAD dynamics but also unveils that the inhibitor studied influences the binding of YAP and TAZ to TEAD in distinct manners. This discovery has significant implications for the design and deployment of future molecular interventions targeting this interaction.

WW domains form a folded type of nuclear localization signal to guide YAP1 nuclear import.

The nuclear translocation of YAP1 is significantly implicated in the proliferation, stemness, and metastasis of cancer cells. Although the molecular basis underlying YAP1 subcellular distribution has been extensively explored, it remains to be elucidated how the nuclear localization signal guides YAP1 to pass through the nuclear pore complex. Here, we define a globular type of nuclear localization signal composed of folded WW domains, named as WW-NLS. It directs YAP1 nuclear import through the heterodimeric nuclear transport receptors KPNA-KPNB1, bypassing the canonical nuclear localization signal that has been well documented in KPNA/KPNB1-mediated nuclear import. Strikingly, competitive interference with the function of the WW-NLS significantly attenuates YAP1 nuclear translocation and damages stemness gene activation and sphere formation in malignant breast cancer cells. Our findings elucidate a novel globular type of nuclear localization signal to facilitate nuclear entry of WW-containing proteins including YAP1.

Mesenchymal stromal cells modulate YAP by verteporfin to mimic cartilage development and construct cartilage organoids based on decellularized matrix scaffolds.

The mechanical elasticity or stiffness of the ECM modulates YAP activity to regulate the differentiation of stem cells during the development and defect regeneration of cartilage tissue. However, the understanding of the scaffold-associated mechanobiology during the initiation of chondrogenesis and hyaline cartilaginous phenotype maintenance remains unclear. In order to elucidate such mechanisms to promote articular cartilage repair by producing more hyaline cartilage, we identify the relationship between YAP subcellular localization and variation of the cartilage structure and organization during the early postnatal explosive growth in incipient articular cartilage. Next, we prepared a decellularized cartilage scaffold with different stiffness (2-33 kPa) to investigate the effect of scaffold stiffness on the formation of hyaline cartilage by mesenchymal stem cells and the change of YAP activity. Furthermore, we simulated the decrease of cellular YAP activity during postnatal cartilage development by inhibiting YAP activity with verteporfin, and realized that the timing of drug incorporation was critical to regulate the differentiation of MSCs to hyaline chondrocytes and inhibit their hypertrophy and fibrosis. On this basis, we constructed hyaline cartilage organoids by decellularized matrix scaffolds. Collectively, the results herein demonstrate that YAP plays a critical role during in vitro chondrogenic differentiation which is tightly regulated by biochemical and mechanical regulation.

Oncogenic YAP mediates changes in chromatin accessibility and activity that drive cell cycle gene expression and cell migration.

YAP, the key protein effector of the Hippo pathway, is a transcriptional co-activator that controls the expression of cell cycle genes, promotes cell growth and proliferation and regulates organ size. YAP modulates gene transcription by binding to distal enhancers, but the mechanisms of gene regulation by YAP-bound enhancers remain poorly understood. Here we show that constitutive active YAP5SA leads to widespread changes in chromatin accessibility in untransformed MCF10A cells. Newly accessible regions include YAP-bound enhancers that mediate activation of cycle genes regulated by the Myb-MuvB (MMB) complex. By CRISPR-interference we identify a role for YAP-bound enhancers in phosphorylation of Pol II at Ser5 at MMB-regulated promoters, extending previously published studies that suggested YAP primarily regulates the pause-release step and transcriptional elongation. YAP5SA also leads to less accessible 'closed' chromatin regions, which are not directly YAP-bound but which contain binding motifs for the p53 family of transcription factors. Diminished accessibility at these regions is, at least in part, a consequence of reduced expression and chromatin-binding of the p53 family member ΔNp63 resulting in downregulation of ΔNp63-target genes and promoting YAP-mediated cell migration. In summary, our studies uncover changes in chromatin accessibility and activity that contribute to the oncogenic activities of YAP.

Advances of targeting the YAP/TAZ-TEAD complex in the hippo pathway for the treatment of cancers.

The Hippo pathway is an evolutionarily conserved signaling pathway that plays critical roles in the tumorigenesis and progression of breast cancer, oral cancer, rectal cancer, colloid cancer, and so on. YAP/TAZ-TEAD complex is a key knot in the Hippo pathway regulating cell proliferation and stem cell functions. Activation or overexpression of this complex has been proved to lead to cell transformation, proliferation and eventually cancerization. In this review, the association between the alterations of hippo pathway and tumorigenesis of various cancer had been elucidated. The structural basis of YAP/TAZ-TEAD complex is analyzed, and the targeting inhibitors are summarized within the medicinal chemistry classification. Moreover, we have also discussed the clinical status and current challenges of these drug candidates, and provide guidance for the future development of inhibitors targeting this pathway, especially YAP/TAZ-TEAD complex.

YAP Promotes Cell Proliferation and Stemness Maintenance of Porcine Muscle Stem Cells under High-Density Condition.

Muscle stem cells (MuSCs) isolated ex vivo are essential original cells to produce cultured meat. Currently, one of the main obstacles for cultured meat production derives from the limited capacity of large-scale amplification of MuSCs, especially under high-density culture condition. Here, we show that at higher cell densities, proliferation and differentiation capacities of porcine MuSCs are impaired. We investigate the roles of Hippo-YAP signaling, which is important regulators in response to cell contact inhibition. Interestingly, abundant but not functional YAP proteins are accumulated in MuSCs seeded at high density. When treated with lysophosphatidic acid (LPA), the activator of YAP, porcine MuSCs exhibit increased proliferation and elevated differentiation potential compared with control cells. Moreover, constitutively active YAP with deactivated phosphorylation sites, but not intact YAP, promotes cell proliferation and stemness maintenance of MuSCs. Together, we reveal a potential molecular target that enables massive MuSCs expansion for large-scale cultured meat production under high-density condition.

Synthetic molecules targeting yes associated protein activity as chemotherapeutics against cancer.

The Hippo signaling pathway extorts several signals that concomitantly target the activity of transcriptional cofactor yes associated protein (YAP). YAP is a key regulator that elicits signature gene expression by coupling with transcriptional enhanced associate domain (TEAD) family of transcriptional factors. The YAP-TEAD complex via target gene expression gets associated with the development, proliferation, and progression of cancerous cells. Moreover, YAP adorns cells with several oncogenic traits such as inhibition of apoptosis, enhanced proliferation, drug resistance, and immune response suppression, which later became associated with various diseases, particularly cancer. Therefore, inhibition of the YAP activity is an appealing and viable therapeutic target for cancer treatment. This review highlights the recent advances in existing and novel synthetic therapeutics targeting YAP inhibition and regulation. The synthetically produced YAPD93A belonging to cyclic peptides and DC-TEADin02 and vinyl sulfonamide class of compounds are the most potent compounds to inhibit the YAP-TEAD expression by targeting protein-protein interaction (IC50  = 25 nM) and palmitate binding central pocket of TEAD (IC50  = 197 nM), respectively. On the other hand, Chlorpromazine belonging to phenothiazines class has the least potential to suppress YAP via proteasomal degradation (cell viability value of <20% at 40 µM).

miR­375/Yes­associated protein axis regulates IL­6 and TGF­ß expression, which is involved in the cisplatin­induced resistance of liver cancer cells.

Chemotherapy resistance is one of the major challenges in the treatment of liver cancer (LC). The present study aimed to investigate the potential roles of Yes­associated protein (YAP), the core component of the Hippo signaling pathway, in chemoresistance of LC. YAP expression and its function in chemoresistance of LC cells were investigated. It was revealed that the expression levels and nuclear localization of YAP were increased in cisplatin (CDDP)­resistant LC (LC/CDDP) cells. The targeted inhibition of YAP using small interfering RNA or an inhibitor restored the CDDP sensitivity of LC cells. YAP overexpression was discovered to be essential for the increase of IL­6 and TGF­ß expression levels in LC/CDDP cells. Furthermore, it was identified that increased mRNA stability was the primary reason for the upregulation of YAP expression in LC/CDDP cells, which was due to the downregulation of microRNA (miR)­375 expression in LC/CDDP cells. In conclusion, the findings of the present study suggested that the miR­375/YAP axis may regulate the expression levels of IL­6 and TGF­ß, which may subsequently be involved in the CDDP resistance of LC cells. The current results indicated that the targeted inhibition of this axis and signaling pathway may be helpful in overcoming CDDP resistance.