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YAP/TAZ are Activated by Mechanical and Hormonal Stimuli in Myometrium and Exhibit Increased Baseline Activation in Uterine Fibroids

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Abstract

Uterine fibroids (UFs) are benign myometrial neoplasms. The mechanical environment activates signaling through the Hippo pathway effectors Yes-associated protein (YAP) and transcriptional coactivator with PDZ-binding domain (TAZ) in other fibrotic disorders. Here, we assess the differences in YAP/TAZ responsiveness to signals in UF compared with myometrium (Myo). Matched samples of UF and Myo were collected. Atomic force microscopy (AFM) was used to determine in situ stiffness. Cells were plated sparsely on hydrogels or at confluence. Ten nanomolars of estradiol (E2) and 100 nM progesterone (P4) were used. Immunostaining for YAP/TAZ and extracellular matrix (ECM) proteins was performed. Cells were incubated with control or YAP1 (YAP)/WWTR1 (TAZ) small interfering RNA (siRNA). Real time qPCR was completed for connective tissue growth factor (CTGF). Cells were treated with verteporfin (a YAP inhibitor) or Y27632 (a ROCK inhibitor), and ECM gene expression was analyzed. Paired t test and Wilcoxon sign-rank test were used. AFM-measured tissue stiffness and YAP/TAZ nuclear localization in situ and in confluent cells were higher in UF compared with Myo (p < 0.05). Decreasing substrate stiffness reduced YAP/TAZ nuclear localization for both Myo and UF (p = 0.05). Stimulating cells with E2 or P4 increased YAP/TAZ nuclear localization, but only in Myo (p = 0.01). UFs had increased FN, COLI, and COLIII deposition. Following siRNA targeting, CTGF was found to be statistically decreased. Verteporfin treatment reduced cell survival and reduced FN deposition. Treatment with Y27632 demonstrated better cell tolerance and a reduction in ECM deposition. The mechanosensitive pathway may be linked to YAP/TAZ function and involved in transducing fibroid growth.

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Acknowledgments

We are highly indebted to those in the Daniel Tschumperlin PhD., lab that helped with this project.

Funding

We would like to express thanks to the Center for Biomedical Discovery-Department of Obstetrics Gynecology Team Science Pilot Award Program for its financial assistance. Supported by the Mayo Clinic Center for Biomedical Discovery-Department of Obstetrics Gynecology Team Science Pilot Award Program.

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Author notes

  1. MacKenzie P. Purdy

    Present address: Division of Reproductive Endocrinology and Infertility, Department of Obstetrics and Gynecology, Mayo Clinic, 200 1st Street SW, Rochester, MN, USA

Authors and Affiliations

  1. Division of Reproductive Endocrinology and Infertility, Department of Obstetrics and Gynecology, Mayo Clinic, 200 1st Street SW, Rochester, MN, USA

    Elizabeth A. Stewart

  2. Department of Physiology and Biomedical Engineering, Mayo Clinic, 200 1st Street SW, Rochester, MN, USA

    Merrick Ducharme, Andrew J. Haak, Jovanka Ravix, Qi Tan, Delphine Sicard, Y. S. Prakash & Daniel J. Tschumperlin

  3. Department of Anesthesiology and Perioperative Medicine, Mayo Clinic, 200 1st Street SW, Rochester, MN, USA

    Jovanka Ravix & Y. S. Prakash

Authors
  1. MacKenzie P. Purdy
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  2. Merrick Ducharme
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  8. Daniel J. Tschumperlin
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  9. Elizabeth A. Stewart
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Contributions

M.P.P., M.D., and A.J.H. designed and executed the experiments, analyzed and interpreted all the data, and drafted the manuscript. J.R., Q.T., D.S., executed and analyzed the experiments, drafted the manuscript, and had final approval. Y.S.P., D.J.T., and E.A.S. contributed to the experimental designs, aided in drafting the manuscript, and had final approval.

Corresponding author

Correspondence to MacKenzie P. Purdy.

Ethics declarations

The protocol has been approved by the Mayo Clinic IRB (17-003124). All patient information was de-identified.

Conflict of Interest

Dr. Stewart reports personal fees from Bayer, personal fees from AbbVie, personal fees from Myovant, personal fees from UpToDate, personal fees from Med Learning Group, outside the submitted work; in addition, Dr. Stewart has a patent Methods and Compounds for Treatment of Abnormal Uterine Bleeding 6440445 issued.

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Purdy, M.P., Ducharme, M., Haak, A.J. et al. YAP/TAZ are Activated by Mechanical and Hormonal Stimuli in Myometrium and Exhibit Increased Baseline Activation in Uterine Fibroids. Reprod. Sci. 27, 1074–1085 (2020). https://doi.org/10.1007/s43032-019-00106-4

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  • DOI: https://doi.org/10.1007/s43032-019-00106-4

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