Dissertations, Theses, and Capstone Projects

Date of Degree

9-2026

Document Type

Doctoral Dissertation

Degree Name

Doctor of Philosophy

Program

Biochemistry

Advisor

Zimei Bu

Committee Members

Ruth Stark

Reza Khayat

Francesca Vallese

Yin Liu

Subject Categories

Biochemistry | Biophysics | Structural Biology

Keywords

Talin, Vinculin, Focal Adhesion, Small Angle X-Ray Scattering, Small Angle Neutron Scattering, Cell-Matrix Adhesion

Abstract

Focal adhesions (FA) are integrin-based cell-matrix adhesion sites composed of adaptor proteins that facilitate the generation of traction forces important for biological functions such as cell migration, embryo development, wound healing, tissue organization and stability, and immune responses. Mature FA are observed when both autoinhibited multi-domain scaffolding protein talin and membrane-cytoskeletal protein vinculin are activated and interact thus promoting the assembly and function of FA. While research has resolved truncated sections of autoinhibited talin and recruitment of vinculin due to mechanotransduction that exposes cryptic vinculin binding sites (VBS), little is known about the global modifications of full length talin in solution when vinculin binds to the VBS without the use of mechanical tension due to intrinsic dynamics that cause limitations to resolve of the entire structure.

Our research determines that talin can bind to vinculin without the mechanical stretching of talin in contrast to the prevailing paradigm in the cell-matrix adhesion research field that says complexation only occurs during mechanical stress. We have determined the solution structure of full length talin by size-exclusion chromatography coupled with Small Angle X-Ray Scattering (SEC-SAXS) showing a dynamic ensemble of flexible conformations for the autoinhibited talin in solution. We have further determined the molecular conformations of talin and vinculin in the reconstituted talin•vinculin complex using SEC-SAXS and selective deuteration/Small Angle Neutron Scattering (SANS)as principal methods. We have found that both talin and vinculin undergo large conformational changes upon forming the talin•vinculin complex that identify the ii activation of VBS on talin and structural changes to vinculin that assemble FA. Our results provide a potentially new mechanism by which talin and vinculin regulate the assembly of focal adhesions in cell-extracellular matrix adhesion and may provide a synergistic pathway to complexation that extends and stabilizes talin in an active conformation.

My thesis aims to show 1) talin can bind to vinculin, forming a discrete complex through biochemical incubation without the need to apply mechanical tension and 2) determine the conformational changes to both talin and in vinculin upon complex formation. My studies provide new mechanisms by which talin engages with vinculin, and open new opportunities for future research to assemble larger FA assemblies for structural, dynamic and functional studies.

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