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The activation of yes‐associated protein 1 (YAP1) and transcriptional co‐activator with PDZ‐binding motif (TAZ) has been implicated in both regeneration and tumorigenesis, thus representing a double‐edged sword in tissue homeostasis. However, how the activity of YAP1/TAZ is regulated or what leads to its dysregulation in these processes remains unknown. To explore the upstream stimuli modulating the cellular activity of YAP1/TAZ, we developed a highly sensitive YAP1/TAZ/TEAD‐responsive DNA element (YRE) and incorporated it into a lentivirus‐based reporter cell system to allow for sensitive and specific monitoring of the endogenous activity of YAP1/TAZ in terms of luciferase activity in vitro and Venus fluorescence in vivo. Furthermore, by replacing YRE with TCF‐ and NF‐κB‐binding DNA elements, we demonstrated the applicability of this reporter system to other pathways such as Wnt/β‐catenin/TCF‐ and IL‐1β/NF‐κB‐mediated signaling, respectively. The practicality of this system was evaluated by performing cell‐based reporter screening of a chemical compound library consisting of 364 known inhibitors, using reporter‐introduced cells capable of quantifying YAP1/TAZ‐ and β‐catenin‐mediated transcription activities, which led to the identification of multiple inhibitors, including previously known as well as novel modulators of these signaling pathways. We further confirmed that novel YAP1/TAZ modulators, such as potassium ionophores, Janus kinase inhibitors, platelet‐derived growth factor receptor inhibitors, and genotoxic stress inducers, alter the protein level or phosphorylation of endogenous YAP1/TAZ and the expression of their target genes. Thus, this reporter system provides a powerful tool to monitor endogenous signaling activities of interest (even in living cells) and search for modulators in various cellular contexts. [ABSTRACT FROM AUTHOR]

Objective: It is well known that reduction of the cyclin-dependent kinase inhibitor p27[sup Kip1] protein correlates with the malignant behavior of various cancers including oral squamous cell carcinoma (OSCC). The loss of p27[sup Kip1] protein is suggested to be due to the enhancement of its posttranslational degradation. In the present study, to evaluate the effects of p27[sup Kip1] transfection on the cell cycle, we transfected OSCC cell lines with a high activity of p27[sup Kip1] degradation with p27[sup Kip1] threonine 187-to-alanine (T187A) mutant gene, which is not influenced by ubiquitin-mediated degradation, as well as with wild type gene. Methods: We transfected p27[sup Kip1] T187A mutant and wild type gene into OSCC cell lines (HSC2 and HSC3) by using an ecdysone-inducible gene expression system. Results: After treatment with ponasterone A, we could find an induction of both p27[sup Kip1] wild type and T187A mutant protein. Both wild type and T187A mutant protein induced by 5 μM ponasterone A inhibited cell growth and increased cell number at the G1 phase. After treatment with 1 μM ponasterone A, ectopic p27[sup Kip1] protein was degraded in wild type clones, but not in T187A mutant clones. Moreover, transfection of the T187A mutant gene was more effective in inhibiting cell growth even by induction of a small amount of protein. Conclusion: We suggest that the transfection of the p27[sup Kip1] T187A mutant gene can be a modality of cancer gene therapy for OSCC.Copyright © 2002 S. Karger AG, Basel [ABSTRACT FROM AUTHOR]

Aldehyde dehydrogenases (ALDHs) are overexpressed in various types of cancers. One of the ALDH family genes, ALDH1A2, is aberrantly expressed in more than 50% of T-cell acute lymphoblastic leukemia (T-ALL) cases. However, its molecular function and role in T-ALL pathogenesis are largely unknown. ChIP-seq and RNA-seq analyses showed that the oncogenic transcription factor TAL1 and its regulatory partners bind to the intronic regulatory element of the ALDH1A2 gene, directly inducing a T-ALL-specific isoform with enzymatic activity. ALDH1A2 was preferentially expressed in the TAL1-positive T-ALL subgroup. In T-ALL cell lines, depletion of ALDH1A2 inhibited cell viability and induced apoptosis. Interestingly, gene expression and metabolomic profiling revealed that ALDH1A2 supported glycolysis and the TCA cycle, accompanied by NADH production, by affecting multiple metabolic enzymes to promote ATP production. Depletion of ALDH1A2 increased the levels of reactive oxygen species (ROS), while ROS levels were reduced by ALDH1A2 overexpression both in vitro and in vivo. Overexpression of ALDH1A2 accelerated tumor onset and increased tumor penetrance in a zebrafish T-ALL model. Taken together, our results indicate that ALDH1A2 protects against intracellular stress and promotes T-ALL cell metabolism and survival. ALDH1A2 overexpression enables leukemic clones to sustain a hyper-proliferative state driven by oncogenes.

The extent of lymph node metastasis is a major determinant in the prognosis of oral squamous cell carcinoma (OSCC). Abnormalities of cell adhesion molecules are known to play an important role in invasion and metastasis of cancer cells through the loss of cell-to-cell adhesion. In this study, we isolated highly invasive clones from an OSCC cell line established from a lymph node metastasis by using an in vitro invasion assay method and compared the abnormalities of cell adhesion molecule E-cadherin and beta-catenin in these cells. The isolated, highly invasive clones showed significant invasive capacity and reduction of E-cadherin and membranous beta-catenin protein in comparison with parent cells. We found that reduced expression of E-cadherin was due to methylation of its promoter region. In fact, most invasive and metastatic area of OSCCs showed reduced expression and methylation of E-cadherin. Moreover, we found that reduced expression of membranous beta-catenin was due to its protein degradation. Reduced expression of membranous beta-catenin was also found frequently in invasive and metastatic areas of OSCCs. In summary, invasion and metastasis of OSCC cells require methylation of E-cadherin and/or degradation of membranous beta-catenin. In addition, we suggest that the method of isolation of highly invasive clones may be useful for studies aimed at discovering novel genes involved in invasion and metastasis.