TEAD2 initiates ground-state pluripotency by mediating chromatin looping.

The transition of mouse embryonic stem cells (ESCs) between serum/LIF and 2i(MEK and GSK3 kinase inhibitor)/LIF culture conditions serves as a valuable model for exploring the mechanisms underlying ground and confused pluripotent states. Regulatory networks comprising core and ancillary pluripotency factors drive the gene expression programs defining stable naïve pluripotency. In our study, we systematically screened factors essential for ESC pluripotency, identifying TEAD2 as an ancillary factor maintaining ground-state pluripotency in 2i/LIF ESCs and facilitating the transition from serum/LIF to 2i/LIF ESCs. TEAD2 exhibits increased binding to chromatin in 2i/LIF ESCs, targeting active chromatin regions to regulate the expression of 2i-specific genes. In addition, TEAD2 facilitates the expression of 2i-specific genes by mediating enhancer-promoter interactions during the serum/LIF to 2i/LIF transition. Notably, deletion of Tead2 results in reduction of a specific set of enhancer-promoter interactions without significantly affecting binding of chromatin architecture proteins, CCCTC-binding factor (CTCF), and Yin Yang 1 (YY1). In summary, our findings highlight a novel prominent role of TEAD2 in orchestrating higher-order chromatin structures of 2i-specific genes to sustain ground-state pluripotency.

Therapeutic targeting of TEAD transcription factors in cancer.

The Hippo signaling pathway inhibits the activity of the oncogenic YAP (Yes-associated protein)/TAZ (transcriptional co-activator with PDZ-binding motif)-TEAD (TEA/ATTS domain) transcriptional complex. In cancers, inactivating mutations in upstream Hippo components and/or enhanced activity of YAP/TAZ and TEAD have been observed. The activity of this transcriptional complex can be effectively inhibited by targeting the TEAD family of transcription factors. The development of TEAD inhibitors has been driven by the discovery that TEAD has druggable hydrophobic pockets, and is currently at the clinical development stage. Three small molecule TEAD inhibitors are currently being tested in Phase I clinical trials. In this review, we highlight the role of TEADs in cancer, discuss various avenues through which TEAD activity can be inhibited, and outline the opportunities for the administration of TEAD inhibitors.

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.

YY1-induced DLEU1/miR-149-5p Promotes Malignant Biological Behavior of Cholangiocarcinoma through Upregulating YAP1/TEAD2/SOX2.

Cholangiocarcinoma is an extremely malignant cancer with poor prognosis. Finding efficient diagnosis and treatment is the indispensable way to improve the prognosis of CCA patients. Therefore, exploring molecular abnormalities in CCA development is urgently needed. DLEU1 is a potential tumor-related lncRNA and abnormally expressed in multiple cancers. In this study, TCGA data analysis showed upregulation of DLEU1 expression in CCA. Furthermore, we confirmed that DLEU1 expression was increased in CCA tissues and cells compared with corresponding controls. Upregulated DLEU1 was related to poor clinicopathological characteristics. Functionally, silencing DLEU1 inhibited CCA proliferation, invasion, stemness maintenance and chemo-resistance, whereas amplifying DLEU1 promoted malignant biological behavior of CCA cells. Mechanistically, DLEU1 expression was transcriptionally facilitated by transcription factor YY1. Moreover, DLEU1 promoted oncogene YAP1 expression by functioning as a sponge to competitively bind to miR-149-5p. YAP1 promoted CCA proliferation, invasion and stemness maintenance, whereas miR-149-5p inhibited malignant biological behavior of CCA. Rescue experiments confirmed that the cancer-promoting effect of DLEU1 was saved by interfering miR-149-5p or YAP1. Furthermore, YAP1 promoted tumor stemness maintenance partly by acting as a transcriptional coactivator to promote TEAD2-induced SOX2 expression. These findings indicated that YY1-induced DLEU1 played a crucial role in CCA progression via miR-149-5p/YAP1/TEAD2/SOX2 axis.

Discovering inhibitors of TEAD palmitate binding pocket through virtual screening and molecular dynamics simulation.

Transcriptional enhanced associate domain (TEAD) proteins bind to YAP/TAZ and mediate YAP/TAZ-induced gene expression. TEADs are not only the key transcription factors and final effector of the Hippo signaling pathway, but also the proteins that regulate cell proliferation and apoptosis. Disorders of Hippo signaling pathway occur in liver cancer, breast cancer, colon cancer and other cancers. S-palmitylation can stabilize the structure of TEADs and is also a necessary condition for the binding of TEADs to YAP/TAZ. The absence of TEAD palmitoylation prevents TEADs from binding to chromatin, thereby inhibiting the transcription and expression of downstream target genes in the Hippo pathway through a dominant-negative mechanism. Therefore, disrupting the S-palmitylation of TEADs has become an attractive and very feasible method in cancer treatment. The palmitate binding pockets of TEADs are conservative, and the crystal structures of TEAD2-palmitoylation inhibitor complexes and the potential TEAD2 inhibitors are more than other TEADs, TEAD2 can be selected to be the target receptor. In this study, structure-based and ligand-based virtual screening, molecular dynamics simulations, Molecular Mechanics Poisson-Boltzmann Surface Area (MM/PBSA) calculations, residue decomposition binding energy calculations, and ADME predictions have been performed to discover 11 potential TEAD2 S-palmitylation inhibitors. ChEBML196567 and ZINC000013942794 are the most recommended, because they formed strong binding energies and stable hydrogen bonds with TEAD2 and have good drugbility and high human oral absorption. We found that it was easier to find the targeting small molecules using a combination of structure-based and ligand-based virtual screening methods. Besides, a new core structure has been found in the selected small molecules. In addition, we analyzed the binding modes of these small molecules to TEAD2, and confirmed the hot spot residues Cys380, Ser345, Tyr426, Phe428, Ile408, and Met379. AVAILABILITY OF DATA AND MATERIAL: Supplementary materials are available online.

New Therapeutic Approach for Targeting Hippo Signalling Pathway.

Nuclear localization signals are short amino acid sequences that target proteins for nuclear import. In this manuscript, we have generated a chimeric tri-functional peptide composed of a cell penetrating peptide (CPP), a nuclear localization sequence and an interfering peptide blocking the interaction between TEAD and YAP, two transcription factors involved in the Hippo signalling pathway, whose deregulation is related to several types of cancer. We have validated the cell penetration and nuclear localization by flow cytometry and fluorescence microscopy and shown that the new generated peptide displays an apoptotic effect in tumor cell lines thanks to the specific nuclear delivery of the cargo, which targets a protein/protein interaction in the nucleus. In addition, the peptide has an anti-tumoral effect in vivo in xenograft models of breast cancer. The chimeric peptide designed in the current study shows encouraging prospects for developing nuclear anti- neoplastic drugs.

The Hippo Pathway and YAP/TAZ-TEAD Protein-Protein Interaction as Targets for Regenerative Medicine and Cancer Treatment.

The Hippo pathway is an important organ size control signaling network and the major regulatory mechanism of cell-contact inhibition. Yes associated protein (YAP) and transcriptional co-activator with PDZ-binding motif (TAZ) are its targets and terminal effectors: inhibition of the pathway promotes YAP/TAZ translocation to the nucleus, where they interact with transcriptional enhancer associate domain (TEAD) transcription factors and coactivate the expression of target genes, promoting cell proliferation. Defects in the pathway can result in overgrowth phenotypes due to deregulation of stem-cell proliferation and apoptosis; members of the pathway are directly involved in cancer development. The pharmacological regulation of the pathway might be useful in cancer prevention, treatment, and regenerative medicine applications; currently, a few compounds can selectively modulate the pathway. In this review, we present an overview of the Hippo pathway, the sequence and structural analysis of YAP/TAZ, the known pharmacological modulators of the pathway, especially those targeting YAP/TAZ-TEAD interaction.

Identification of transcriptional enhancer factor-4 as a transcriptional modulator of CTP:phosphocholine cytidylyltransferase alpha.

CTP:phosphocholine cytidylyltransferase (CCT) is the rate-limiting and regulated enzyme of mammalian phosphatidylcholine biosynthesis. There are three isoforms, CCTalpha, CCTbeta1, and CCTbeta2. The mouse CCTalpha gene promoter is regulated by an enhancer element (Eb) located between -103 and -82 base pairs (5'-GTTTTCAGGAATGCGGAGGTGG-3') upstream from the transcriptional start site (Bakovic, M., Waite, K., Tang, W., Tabas, I., and Vance, D. E. (1999) Biochim. Biophys. Acta 1436, 147-165). To identify the Eb-binding protein(s), we screened a mouse embryo cDNA library by the yeast one-hybrid system and obtained 19 positive clones. Ten cDNA clones were identified as transcriptional enhancer factor-4 (TEF-4). The TEF-binding consensus sequence, 5'-(A/T)(A/G)(A/G)(A/T)ATG(C/T)(G/A)-3', was identified within the Eb binding region. Gel-shift analysis using radiolabeled Eb fragment as a probe showed that cell extracts from yeast expressing hemagglutinin-tagged TEF-4 caused a marked band retardation that could be prevented with an anti-hemagglutinin antibody. When COS-7 cells were transfected with TEF-4, CCTalpha promoter-luciferase reporter activity and CCTalpha mRNA levels increased. A TEF-4 deletion mutant containing a DNA-binding domain, mTEA(+), stimulated the CCTalpha promoter activity, whereas protein lacking the DNA binding domain, mTEA(-), did not. Unexpectedly, when the ATG core of the TEF-4 binding consensus within the Eb region was mutated, promoter activity was enhanced rather than decreased. Thus, TEF-4 might act as a dual transcriptional modulator as follows: as a suppressor via its direct binding to the Eb element and as an activator via its interactions with the basal transcriptional machinery. These results provide the first evidence that TEF-4 is an important regulator of CCTalpha gene expression.

Transforming growth factor-beta response elements of the skeletal alpha-actin gene. Combinatorial action of serum response factor, YY1, and the SV40 enhancer-binding protein, TEF-1.

Skeletal alpha-actin (SkA) is representative of the cardiac genes that are expressed at high levels in embryonic myocardium, down-regulated after birth, and reactivated by trophic signals including type beta-transforming growth factors (TGF beta). To investigate the molecular basis for cardiac-restricted and TGF beta-induced SkA transcription, we have undertaken a mutational analysis of the SkA promoter in ventricular myocytes, with emphasis on the role of three nominal serum response elements. Serum response factor (SRF) and the bifunctional factor YY1 are the predominant cardiac proteins contacting the proximal SRE (SRE1). Mutations of SRE1 that prevent recognition by SRF and YY1, or SRF alone, virtually abolish SkA transcription in both TGF beta- and vehicle-treated cells; mutation of distal SREs was ineffective. A mutation which selectively abrogates YY1 binding increases both basal and TGF beta-dependent expression, substantiating the predicted role of YY1 as an inhibitor of SRF effects. However, efficient SkA transcription requires combinatorial action of SRE1 with consensus sites for Sp1 and the SV40 enhancer-binding protein, TEF-1. As isolated motifs, either SRE1- or TEF-1-binding sites function as TGF beta response elements. Induction of the SkA promoter by TGF beta required SRF and TEF-1 in concert, unlike other pathways for TGF beta-dependent gene expression.