Merkel, Chelsea
(2019)
The Adherens Junction Interactome and the Role of Vinculin in Cytoskeletal Integration at the Cardiomyocyte ICD.
Doctoral Dissertation, University of Pittsburgh.
(Unpublished)
Abstract
The primary function of the heart is to contract and pump oxygenated blood throughout the body. Coordinated contraction between individual cardiomyocytes requires mechanical and electrical coupling through a specialized adhesive structure called the intercalated disc (ICD). The ICD joins cardiomyocytes and is comprised of adherens junctions (AJ), desmosomes, and gap junctions. The core of the AJ is the cadherin-catenin complex and it links the myofibrils of neighboring cells. Much work has been performed in epithelial cells to study the role of force in regulating complex formation and ligand recruitment. However, the range and scale of forces experienced in epithelial cells is far less than what is produced in the heart, yet the same molecules are responsible to maintain adhesion and tissue integrity. Our studies sought to understand the specialization of the AJ in cardiomyocytes to uncover mechanisms of resilience in tissues. Our group used proximity based proteomics and mass spectrometry to investigate the N-cadherin interactome in primary neonatal mouse cardiomyocytes. We found that cardiomyocytes share core molecular components compared to epithelial cells, however, they recruit a host of unique adapter proteins. Additionally, we demonstrated cross-talk between the ICD and the Z-disk that was previously unreported. From our proximity proteomics data set, I chose to investigate two characterized AJ ligands previously studied in epithelial cells for their roles in mechanotransduction. I found that both vinculin and afadin are localized at the ICD and can be selectively enriched through the introduction of N-cadherin:E-catenin fusion constructs. With these constructs, I determined that vinculin enhances the stability of AJs and is necessary and sufficient to link the AJ to the myofibril network. Lastly, I demonstrated that vinculin recruitment is necessary to drive the maturation of the ICD by recruiting desmosomes. Our work demonstrates that tissue-specific differences in the AJ interactome give rise to its specialization and that the known E-catenin ligand, vinculin, plays an integral role in bridging the AJ-myofibril interface. Together, these data provide a repository for future work into novel ICD proteins and a novel function for vinculin, providing insight for declining cardiomyocyte function and remodeling in disease states.
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Details
| Item Type: |
University of Pittsburgh ETD
|
| Status: |
Unpublished |
| Creators/Authors: |
|
| ETD Committee: |
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| Date: |
19 September 2019 |
| Date Type: |
Publication |
| Defense Date: |
21 June 2019 |
| Approval Date: |
19 September 2019 |
| Submission Date: |
5 August 2019 |
| Access Restriction: |
No restriction; Release the ETD for access worldwide immediately. |
| Number of Pages: |
172 |
| Institution: |
University of Pittsburgh |
| Schools and Programs: |
School of Medicine > Cell Biology and Molecular Physiology |
| Degree: |
PhD - Doctor of Philosophy |
| Thesis Type: |
Doctoral Dissertation |
| Refereed: |
Yes |
| Uncontrolled Keywords: |
Cellular Biology |
| Date Deposited: |
19 Sep 2019 19:25 |
| Last Modified: |
19 Sep 2019 19:25 |
| URI: |
http://d-scholarship.pitt.edu/id/eprint/37285 |
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