Journal
DISEASE MODELS & MECHANISMS
Volume 10, Issue 9, Pages 1129-1140Publisher
COMPANY BIOLOGISTS LTD
DOI: 10.1242/dmm.029926
Keywords
Yeast; Genetic screen; Protein levels; Protein disulfide isomerase; TorsinA; Dystonia
Categories
Funding
- Dystonia Medical Research Foundation
- CONICET (Consejo Nacional de Investigaciones Cientificas y Tecnicas
- Argentina)
- Endeavour (Department of Education and Training
- Australia)
- National Cancer Institute training grant, USA [T32 CA009503]
- National Breast Cancer Foundation Early Career Researcher Fellowship, Australia [ECF-12-12]
- Australian National Health and Medical Research Council RD Wright Biomedical [APP1087975]
- Pittsburgh Center for Kidney Research-Model Organisms Core grant (University of Pittsburgh) [DK79307]
- National Institutes of Health, USA [GM75061, ES024872]
- Parkinson's and Movement Disorder Foundation
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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, torsinA.E, 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 torsinA.E variant, and torsinA.E is also less stable than wild-type torsinA and is a substrate for ER-associated degradation. Nevertheless, the molecular factors involved in the biogenesis and degradation of torsinA and torsinA.E have not been fully explored. To identify conserved cellular factors that can alter torsinA.E protein levels, we designed a new high-throughput, automated, genome-wide screen utilizing our validated Saccharomyces cerevisiae torsinA expression system. By analyzing the yeast non-essential gene deletion collection, we identified 365 deletion strains with altered torsinA.E 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 torsinA.E. Together, our data report the first genome-wide screen to identify cellular factors that alter expression levels of the EOTD-associated protein torsinA.E. More generally, the identified hits help in dissecting 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.
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