Mechanotransduction at focal adhesions: interplay among force, FAK, and YAP

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MARQUEZ-DISSERTATION-2024.pdf (3.5 MB)
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Marquez, Elijah Nathaniel
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School of Chemical and Biomolecular Engineering
School established in 1901 as the School of Chemical Engineering; in 2003, renamed School of Chemical and Biomolecular Engineering
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https://hdl.handle.net/1853/78582
Abstract
The body experiences a variety of mechanical cues in day-to-day function: compressive and tensile stresses in bone and cartilage to latent mechanical signals from the extracellular matrix (ECM). Mechanical cues drive processes such as cell migration and proliferation. Cells sense mechanical signals via cell-ECM interactions, which are primarily facilitated through focal adhesions (FAs). FAs have 100s of unique components: a few key proteins are vinculin, talin, and focal adhesion kinase (FAK). While FA turnover has been well recognized, how cells translate mechanical signals at FAs into biochemical changes is less understood. Previous work demonstrated that yes-associated protein (YAP), a transcriptional coactivator, responds to changes in adhesive area and substrate rigidity. Deleting essential FA proteins alters YAP activity, and YAP upregulates FA-related genes. Our objective is to further our understanding of cell adhesion by elucidating how cells utilize focal adhesions to translate mechanical cues into biochemical signals. I hypothesize that altering vinculin-talin-FAK interactions will reduce YAP nuclear accumulation and activity, whereas spatially directing adhesive ligand uncaging will promote vinculin recruitment and FA force. Therefore, as I alter vinculin-talin-FAK interactions, I expect a reduction in YAP activity. As I spatially uncage adhesive ligands, I expect to induce vinculin localization and FA force. In this thesis, I demonstrated that reducing micropillar area while keeping array stiffness constant, which increases force map resolution, alters cell behavior by reducing cell contractility and spread area. I evaluated the impact of removing talin-FAK and talin-vinculin interactions on YAP activity. Upon removing vinculin, FAK, or talin; impairing FAK or vinculin’s functionality, or abrogating talin-FAK or talin-vinculin interactions, there was a drop in YAP’s nuclear accumulation and transcriptional activity. Lastly, I developed a platform for spatially and temporally directing FAs to monitor FA distribution, number, and cell generated traction forces. This work advances our understanding of mechanotransduction by dissecting the relationship between FAs and YAP, which aids in rationally designing biomaterial therapies for modulating YAP expression to treat cancer and fibrosis.
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2024-07-26
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