Investigating the biophysical mechanisms of substrate rigidity-induced hippo pathway regulation in human embryonic stem cells
Abstract/Contents
- Abstract
- Mechanobiology, the study of how cells sense and transduce mechanical information into chemical signals, has become an integral aspect of both basic and applied biological research over the past decade. Of particular interest to researchers is the Hippo signaling pathway, a mechanosensitive pathway that participates in diverse cellular processes ranging from embryonic development to cancer progression. The transcriptional coactivator YAP (yes-associated protein) is a central effector in the Hippo pathway. In embryonic stem cells, YAP activation promotes cell survival and pluripotency. YAP also directs cell fate in mouse embryos and mesenchymal stem cells in a manner that depends on both cell position within tissues and substrate rigidity. Despite YAP's central biological importance, the mechanisms underlying its regulation by mechanical inputs remain incompletely understood. We used human embryonic stem cells (hESCs) as a model system to determine how mechanical cues such as those that are present in mammalian embryos regulate YAP activity. To do so, we performed an unbiased proteomics screen to identify a Hippo pathway interactome, and found that the set of proteins interacting with YAP and its upstream kinase large tumor suppressor (Lats) was highly enriched for proteins associated with the actin cytoskeleton. Using engineered hydrogels with tunable stiffness, we observed that hESCs adopted drastically different colony morphologies, actin cytoskeleton organization, and YAP nuclear versus cytoplasmic localization on substrates of differing stiffnesses. Further investigation revealed that an intact actin cytoskeleton was required for the regulation of Hippo pathway activity as a function of cell position within hESC colonies, but that signaling downstream of RhoA and Rho-associated coiled-coil kinase (ROCK) was specifically required for Hippo pathway modulation in response to substrate stiffness. Contrary to results reported for other cell types, we found that both bundled F-actin and nonmuscle myosin II activity were dispensable for YAP mechanoregulation in hESCs. In addition, we found that YAP phosphorylation by Lats was required for the modulation of YAP activity in response to substrate stiffness. Lastly, we evaluated the biological relevance of YAP activation and found that YAP cytoplasmic sequestration in cells adhering to soft substrates did not induce changes in cell fate or cell proliferation. Instead, nuclear localization of YAP and its interaction with the transcription factor TEAD were crucial for hESC survival. A quantitative model relating substrate stiffness and ROCK activity to YAP phosphorylation and nuclear localization can account for our results and highlights the unique mechanisms that modulate Hippo pathway signaling in pluripotent cells.
Description
| Type of resource | text |
|---|---|
| Form | electronic resource; remote; computer; online resource |
| Extent | 1 online resource. |
| Place | California |
| Place | [Stanford, California] |
| Publisher | [Stanford University] |
| Copyright date | 2018; ©2018 |
| Publication date | 2018; 2018 |
| Issuance | monographic |
| Language | English |
Creators/Contributors
| Author | Huang, Eva Yi-Hsuan |
|---|---|
| Degree supervisor | Dunn, Alexander Robert |
| Thesis advisor | Dunn, Alexander Robert |
| Thesis advisor | Fuller, Margaret T, 1951- |
| Thesis advisor | Khosla, Chaitan, 1964- |
| Thesis advisor | Sebastiano, Vittorio |
| Degree committee member | Fuller, Margaret T, 1951- |
| Degree committee member | Khosla, Chaitan, 1964- |
| Degree committee member | Sebastiano, Vittorio |
| Associated with | Stanford University, Department of Chemical Engineering. |
Subjects
| Genre | Theses |
|---|---|
| Genre | Text |
Bibliographic information
| Statement of responsibility | Eva Yi-Hsuan Huang. |
|---|---|
| Note | Submitted to the Department of Chemical Engineering. |
| Thesis | Thesis Ph.D. Stanford University 2018. |
| Location | electronic resource |
Access conditions
- Copyright
- © 2018 by Eva Yi-Hsuan Huang
- License
- This work is licensed under a Creative Commons Attribution Non Commercial 3.0 Unported license (CC BY-NC).
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