An Investigation of TorsinA Interaction Partners
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hernandez-vjhernan-phd-bio-2024-thesis.pdf
Description
Thesis PDF
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4.17 MB
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e38dc20cea664ce67873a62380667052
Author(s)
Hernandez, Victoria J.
Advisor(s)
Schwartz, Thomas U.
Date Issued
May 2024
Publisher
Massachusetts Institute of Technology
Abstract
TorsinA is a AAA+ (ATPases associated with a variety of cellular activities) protein that is implicated in the neuromuscular disorder DYT-TOR1A early-onset isolated dystonia. DYT-TOR1A is a heritable form of dystonia characterized by involuntary twisting movements and postures that arise during adolescence. A glutamate deletion towards the C terminus of TorsinA leads to DYT-TOR1A by disrupting the ability for TorsinA to interact with its ATPase activators Lamina Associated Polypeptide 1 (LAP1) and Luminal Domain Like LAP1 (LULL1), rendering TorsinA catalytically inert. TorsinA’s precise role in DYT-TOR1A remains elusive in large part because its function is unknown; a major impediment in understanding TorsinA’s function is the lack of any identified substrate of TorsinA. Here, we performed a TorsinA pulldown from mammalian cells to re-examine TorsinA’s interaction partners. We identified Calnexin, a lectin chaperone in the endoplasmic reticulum (ER), as the most abundant protein associated with TorsinA. Prior studies had identified Calnexin as a binding partner of TorsinA, assuming Calnexin to interact as folding chaperone for TorsinA. We chose to investigate the interaction between TorsinA and Calnexin in further detail, by studying the elements of TorsinA that are necessary for Calnexin binding. We found that while TorsinA N-glycosylation is required for Calnexin binding, terminal mono-glucosylation is not. This finding deviates from Calnexin’s interactions with its canonical substrates, as Calnexin’s lectin domain specifically recognizes mono-glucosylated N-glycans. Furthermore, we found that TorsinA remains associated with Calnexin 3 hours following translation inhibition. This finding again deviates from the Calnexin substrate model, as Calnexin preferentially binds newly synthesized proteins. Therefore, we conclude that the interaction between TorsinA and Calnexin likely has functional significance unrelated to TorsinA biogenesis, and that the two proteins may function as co-chaperones. Understanding the function of TorsinA in the context of Calnexin could bring us closer to identifying a substrate, or substrates, of TorsinA, thereby illuminating TorsinA’s role in DYT-TOR1A.
MIT Department
Massachusetts Institute of Technology. Department of Biology
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