Bowman, Ray Wesley
(2022)
a-Arrestin phospho-regulation and control of autophagosome formation.
Doctoral Dissertation, University of Pittsburgh.
(Unpublished)
This is the latest version of this item.
Abstract
Cells selectively reorganize their proteome in response to environmental stressors. For membrane proteins, this reshuffling is controlled by selective protein trafficking. Studies in yeast demonstrate that the a-arrestins, a highly conserved family of protein trafficking adapters, are critical players in the selective movement of membrane proteins. In response to cell signaling, a-arrestins are post-translationally modified and bind specific membrane proteins. In doing so, they recruit with them the ubiquitin ligase Rsp5, acting as a bridge between the ligase and the membrane protein. Ubiquitination of the membrane protein promotes sorting into endocytic vesicles. There are many outstanding questions in the field of a-arrestins, and my research focuses on defining a new role for a-arrestins in influencing autophagy and expanding our understanding of a-arrestin phospho-regulation. Autophagy is a highly conserved self-degradative process controlled by the TORC1 signaling complex that is utilized by cells to survive periods of nutrient limitation through the bulk-transport of cellular components to the vacuole for degradation. We employ a targeted genetic screen to uncover a series of mutations that alter a-arrestins’ ability to confer resistance to the TORC1-inhibiting drug rapamycin, revealing a complex genetic tie between the a-arrestins and autophagy. We further show that the absence of a-arrestins results in impaired autophagic flux to the vacuole, a defect in the ability of a-arrestin mutants to form autophagy-dedicated membrane-bound vesicles (autophagosomes), as well as an aberrant retention of autophagic machinery to the vacuolar membrane that coincides with defects in the localization of phospholipid species and their modifying enzymes. Furthermore, phosphorylation of a-arrestins alters their protein trafficking function. We therefore performed a second targeted screen to identify a-arrestin phospho-regulators, finding the TORC1-Sit4-Npr1 signaling network to possess a prominent role in controlling a-arrestin phosphorylation and stability, as well as establishing a novel link between a-arrestin stability and vacuolar degradation. Together, my work forges new connections between the a-arrestins and the maintenance of the vacuolar membrane’s composition, the ability of the cell to properly respond to TORC1 inhibition in their absence, as well as the post-translational regulation and stability of the a-arrestins themselves.
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Details
| Item Type: |
University of Pittsburgh ETD
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| Status: |
Unpublished |
| Creators/Authors: |
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| ETD Committee: |
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| Date: |
3 November 2022 |
| Defense Date: |
21 July 2022 |
| Approval Date: |
25 January 2023 |
| Submission Date: |
10 November 2022 |
| Access Restriction: |
No restriction; Release the ETD for access worldwide immediately. |
| Number of Pages: |
262 |
| Institution: |
University of Pittsburgh |
| Schools and Programs: |
Dietrich School of Arts and Sciences > Biological Sciences |
| Degree: |
PhD - Doctor of Philosophy |
| Thesis Type: |
Doctoral Dissertation |
| Refereed: |
Yes |
| Uncontrolled Keywords: |
Arrestin phosphorylation ubiquitination autophagy |
| Related URLs: |
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| Date Deposited: |
25 Jan 2023 14:36 |
| Last Modified: |
25 Jan 2023 14:36 |
| URI: |
http://d-scholarship.pitt.edu/id/eprint/43830 |
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