Mukundan, Ananya
(2023)
Structural insights into mimicry of TGF-β signaling by the murine parasite H. polygyrus.
Doctoral Dissertation, University of Pittsburgh.
(Unpublished)
Abstract
TGF-β is a key regulator of the adaptive immune system, acting to convert naïve CD4+ T-cells into Foxp3+ Tregs. The mouse intestinal hookworm Heligmosomoides polygyrus takes advantage of this pathway, surviving in its host by using a five-domain secreted TGF-β mimic protein, TGM or TGM-1 to expand the population of Tregs in the host. TGM-1 binds to the TGF-beta receptors directly and is part of a larger family of 10 secreted parasitic proteins (TGM-1:TGM-10), all of which lack sequence and structural homology to TGF-β, and are instead composed of five CCP domains (D1-D5). In this dissertation, biophysical methods were used to study TGM-1 and TGM-4, how they bind the TGF-β receptors and how to leverage these structural details towards the development of TGF-β-targeting therapeutics. SPR, ITC, and NMR showed that binding of TGM-1 to the TGF-β receptors is modular, with D1 and D2 binding to TβRI and D3 binding to TβRII. The domains of TGM-1 compete with TGF-β for binding to TβRI and TβRII, showing that TGM-1 mimics TGF-β both functionally and structurally. The solution structure of TGM-1 D3 and the TGM-1 D3:TBRII complex show that the TGM-1 D3 CCP domain is adapted to allow for expansion of C-terminal segment, partially exposing several hydrophobic residues that interact with the edged β-strand that TβRII also uses to bind TGF-β. Further studies revealed that domains 4 and 5 of TGM-1 bind a co-receptor CD44 that potentiates TGM-1 signaling in vitro. This co-receptor binds to TGM-1 and TGM-4 but not TGM-6, indicating that different TGM isoforms bind to distinct co-receptors depending on the domain 4-5 composition. This led to the hypothesis that cell-targeted TGF-β inhibitors could be developed by fusing known TGF-β inhibitors to domains 4 and 5 of TGM-1. Initial tests into these fused proteins indicate inhibition of TGF-β signaling, with studies into cell-specific inhibition and potentiation to be performed in the future. This dissertation demonstrates how H. polygyrus has adapted a family of proteins to mimic TGF-β signaling and opens up unique avenues for inhibition of TGF-β signaling.
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Details
| Item Type: |
University of Pittsburgh ETD
|
| Status: |
Unpublished |
| Creators/Authors: |
|
| ETD Committee: |
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| Date: |
8 December 2023 |
| Date Type: |
Publication |
| Defense Date: |
1 June 2023 |
| Approval Date: |
8 December 2023 |
| Submission Date: |
23 June 2023 |
| Access Restriction: |
2 year -- Restrict access to University of Pittsburgh for a period of 2 years. |
| Number of Pages: |
312 |
| Institution: |
University of Pittsburgh |
| Schools and Programs: |
School of Medicine > Molecular Biophysics and Structural Biology |
| Degree: |
PhD - Doctor of Philosophy |
| Thesis Type: |
Doctoral Dissertation |
| Refereed: |
Yes |
| Uncontrolled Keywords: |
TGF-β, parasitic TGF-β mimic, TGF-β mimic, murine parasite, H. polygyrus, parasitic mimicry, immune suppression, nuclear magnetic resonance, isothermal titration calorimetry, surface plasmon resonance, TGM |
| Date Deposited: |
08 Dec 2023 20:33 |
| Last Modified: |
08 Dec 2023 20:33 |
| URI: |
http://d-scholarship.pitt.edu/id/eprint/44997 |
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