A novel high-throughput yeast genetic screen for factors modifying protein levels of the Early-Onset Torsion Dystonia-associated variant torsinAΔE
| Autor: | John C. Dittmar, Jeffrey L. Brodsky, Annette M. Shewan, Michael J. Mihalevic, Lucia F. Zacchi, Benjamin L. Schulz, Kara A. Bernstein |
|---|---|
| Rok vydání: | 2017 |
| Předmět: |
0301 basic medicine
Genetics Protein Disulfide-Isomerase Family biology Endoplasmic reticulum Saccharomyces cerevisiae Neuroscience (miscellaneous) Medicine (miscellaneous) Translocon biology.organism_classification General Biochemistry Genetics and Molecular Biology Cell biology 03 medical and health sciences 030104 developmental biology 0302 clinical medicine Immunology and Microbiology (miscellaneous) Chaperone (protein) biology.protein Protein disulfide-isomerase 030217 neurology & neurosurgery Biogenesis Genetic screen |
| Zdroj: | Disease Models & Mechanisms. |
| ISSN: | 1754-8411 1754-8403 |
| DOI: | 10.1242/dmm.029926 |
| Popis: | Dystonia is the third most common movement disorder, but its diagnosis and treatment remain challenging. One of the most severe types of Dystonia is Early-Onset Torsion Dystonia (EOTD). The best studied and validated EOTD-associated mutation, torsinAΔE, is a deletion of a C-terminal glutamate residue in the AAA+ ATPase, torsinA. TorsinA appears to be an Endoplasmic Reticulum (ER)/Nuclear Envelope chaperone with multiple roles in the secretory pathway and in determining subcellular architecture. Many functions are disabled in the torsinAΔE variant, and torsinAΔE is also less stable than wild-type torsinA and is a substrate for ER-associated degradation. Nevertheless, the molecular factors involved in torsinA and torsinAΔE's biogenesis and degradation have not been fully explored. To identify conserved cellular factors that can alter torsinAΔE protein levels, we designed a new high-throughput, automated, genome-wide screen utilizing our validated Saccharomyces cerevisiae torsinA expression system. By analyzing the non-essential yeast deletion collection, we identified 365 deletion strains with altered torsinAΔE steady-state levels. One notable hit was EUG1, which encodes a member of the protein disulfide isomerase family (PDIs). PDIs reside in the ER and catalyze the formation of disulfide bonds, mediate protein quality control, and aid in nascent protein folding. We validated the role of select human PDIs in torsinA biogenesis in mammalian cells and found that overexpression of PDIs reduced the levels of torsinA and torsinAΔE. Together, our data report the first genome-wide screen to identify cellular factors that reduce expression levels of the EOTD-associated protein torsinAΔE. More generally, the identified hits contribute to dissect the cellular machinery involved in folding and degrading a torsinA variant, and constitute potential therapeutic factors for EOTD. This screen can also be readily adapted to identify factors impacting the levels of any protein of interest, considerably expanding the applicability of yeast in both basic and applied research. |
| Databáze: | OpenAIRE |
| Externí odkaz: |
|