The Hippo pathway component WW domain-containing transcription regulator 1 (TAZ) is a transcriptional coactivator and an oncogene in breast and lung cancer. Transcriptional targets of TAZ that modulate immune cell function in the tumor microenvironment are poorly understood. Here, we perform a comprehensive screen for immune-related genes regulated by TAZ and its paralog YAP using NanoString gene expression profiling. We identify the immune checkpoint molecule PD-L1 as a target of Hippo signaling. The upstream kinases of the Hippo pathway, mammalian STE20-like kinase 1 and 2 (MST1/2), and large tumor suppressor 1 and 2 (LATS1/2), suppress PD-L1 expression while TAZ and YAP enhance PD-L1 levels in breast and lung cancer cell lines. PD-L1 expression in cancer cell lines is determined by TAZ activity and TAZ/YAP/TEAD increase PD-L1 promoter activity. Critically, TAZ-induced PD-L1 upregulation in human cancer cells is sufficient to inhibit T-cell function. The relationship between TAZ and PD-L1 is not conserved in multiple mouse cell lines, likely due to differences between the human and mouse PD-L1 promoters. To explore the extent of divergence in TAZ immune-related targets between human and mouse cells, we performed a second NanoString screen using mouse cell lines. We show that many targets of TAZ may be differentially regulated between these species. These findings highlight the role of Hippo signaling in modifying human/murine physiologic/pathologic immune responses and provide evidence implicating TAZ in human cancer immune evasion.Significance: Human-specific activation of PD-L1 by a novel Hippo signaling pathway in cancer immune evasion may have a significant impact on research in immunotherapy. Cancer Res; 78(6); 1457–70. ©2018 AACR.

The Hippo pathway is a central regulator of tissue development and homeostasis, and has been reported to have a role during vascular development. Here we develop a bioluminescence-based biosensor that monitors the activity of the Hippo core component LATS kinase. Using this biosensor and a library of small molecule kinase inhibitors, we perform a screen for kinases modulating LATS activity and identify VEGFR as an upstream regulator of the Hippo pathway. We find that VEGFR activation by VEGF triggers PI3K/MAPK signaling, which subsequently inhibits LATS and activates the Hippo effectors YAP and TAZ. We further show that the Hippo pathway is a critical mediator of VEGF-induced angiogenesis and tumor vasculogenic mimicry. Thus, our work offers a biosensor tool for the study of the Hippo pathway and suggests a role for Hippo signaling in regulating blood vessel formation in physiological and pathological settings.

Since its discovery, the Hippo pathway has emerged as a central signaling network in mammalian cells. Canonical signaling through the Hippo pathway core components (MST1/2, LATS1/2, YAP and TAZ) is important for development and tissue homeostasis while aberrant signaling through the Hippo pathway has been implicated in multiple pathologies, including cancer. Recent studies have uncovered new roles for the Hippo pathway in immunology. In this review, we summarize the mechanisms by which Hippo signaling in pathogen-infected or neoplastic cells affects the activities of immune cells that respond to these threats. We further discuss how Hippo signaling functions as part of an immune response. Finally, we review how immune cell-intrinsic Hippo signaling modulates the development/function of leukocytes and propose directions for future work.

The Hippo pathway has emerged as a key signaling pathway that regulates a broad range of biological functions, and dysregulation of the Hippo pathway is a feature of a variety of cancers. Given this, some have suggested that disrupting the interaction of the Hippo core component YAP and its paralog TAZ with transcriptional factor TEAD may be an effective strategy for cancer therapy. However, there are currently no clinically available drugs targeting the YAP/TAZ–TEAD interaction for cancer treatment. To facilitate screens for small molecule compounds that disrupt the YAP–TEAD interaction, we have developed the first ultra-bright NanoLuc biosensor to quantify YAP/TAZ–TEAD protein–protein interaction (PPI) both in living cells and also in vitro using biosensor fusion proteins purified from bacteria. Using this biosensor, we have performed an in vitro high throughput screen (HTS) of small molecule compounds and have identified and validated the drug Celastrol as a novel inhibitor of YAP/TAZ–TEAD interaction. We have also demonstrated that Celastrol can inhibit cancer cell proliferation, transformation, and cell migration. In this study, we describe a new inhibitor of the YAP/TAZ–TEAD interaction warranting further investigation and offer a novel biosensor tool for the discovery of other new Hippo-targeting drugs in future work.