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Monitoring Hippo Signaling Pathway Activity Using a Luciferase-based Large Tumor Suppressor LATS Biosensor
Published on: September 13, 2018
An improved TEAD dominant-negative protein inhibitor to study Hippo YAP1/TAZ-dependent transcription
Briana Branch1,2, Yao Yuan1, Mariastella Cascone3
1Laboratory of Cellular and Molecular Biology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, Maryland, United States.
Abstract:
Hippo signaling is one of the top pathways altered in human cancer, and intensive focus has been devoted to developing therapies targeting Hippo-dependent transcription mediated by YAP1 and TAZ interaction with TEAD proteins. However, a significant challenge in evaluating the efficacy of these approaches is the lack of models that can precisely characterize the consequences of TEAD inhibition. To address this gap, our laboratory developed a strategy that utilizes a fluorescently traceable, dominant-negative protein named TEADi. TEADi specifically blocks the nuclear interactions of TEAD with YAP1 and TAZ, enabling precise dissection of Hippo TEAD-dependent and independent effects on cell fate. In this study, we aimed to enhance TEADi effectiveness by altering post-transcriptional modification sites within its TEAD-binding domains (TBDs). We demonstrate that a D93E mutation in the YAP1 TBD significantly increases TEADi inhibitory capacity. Additionally, we find that TBDs derived from VGLL4 and YAP1 are insufficient to block TAZ-induced TEAD activity, revealing crucial differences in YAP1 and TAZ displacement mechanisms by dominant-negative TBDs. Structural differences in YAP1 and TAZ TBDs were also identified, which may contribute to the distinct binding of these proteins to TEAD. Our findings expand our understanding of TEAD regulation and highlight the potential of an optimized TEADi as a more potent, specific, and versatile tool for studying TEAD-transcriptional activity.
Insights
Researchers enhanced a tool called TEAD inhibitor (TEADi) to better study the Hippo signaling pathway in cancer. A specific mutation improved its ability to block cancer-driving proteins, offering a more precise way to investigate cancer therapies.
Area of Science:
- Molecular Biology
- Cancer Research
- Cell Biology
Background:
- Hippo signaling pathway is frequently altered in human cancers.
- YAP1 and TAZ proteins interact with TEAD proteins, driving transcription in cancer.
- Developing therapies targeting TEAD-YAP1/TAZ interaction is a major focus, but effective models for TEAD inhibition are lacking.
Purpose of the Study:
- To enhance the effectiveness of a dominant-negative TEAD inhibitor (TEADi) by modifying its TEAD-binding domains (TBDs).
- To investigate the impact of specific mutations and TBD origins on TEADi's inhibitory capacity.
- To elucidate the distinct mechanisms of YAP1 and TAZ displacement by dominant-negative TBDs.
Main Methods:
- Engineered a fluorescently traceable, dominant-negative protein (TEADi) to block TEAD-YAP1/TAZ nuclear interactions.
- Introduced mutations, including D93E, into TEADi's TBDs to assess their effect on inhibitory capacity.
- Utilized TEADi variants with TBDs from VGLL4 and YAP1 to study TAZ-induced TEAD activity.
- Analyzed structural differences in YAP1 and TAZ TBDs.
Main Results:
- A D93E mutation in the YAP1 TBD significantly enhanced TEADi's inhibitory capacity.
- TEAD-binding domains derived from VGLL4 and YAP1 were insufficient to block TAZ-induced TEAD activity.
- Identified distinct structural differences between YAP1 and TAZ TBDs, potentially explaining differential binding to TEAD.
- Demonstrated that modified TEADi allows precise dissection of Hippo TEAD-dependent and independent effects.
Conclusions:
- An optimized TEADi with a D93E mutation represents a more potent and specific tool for cancer research.
- Understanding the differential mechanisms of YAP1 and TAZ inhibition is crucial for developing targeted cancer therapies.
- This enhanced TEADi tool facilitates the study of TEAD-transcriptional activity and evaluation of novel cancer treatments.
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