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Kinetochores are required to fully activate secretory pathway in elevated ER stress

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dc.contributor.advisor Atkinson, Paul
dc.contributor.author Low, Yee Syuen
dc.date.accessioned 2013-12-11T22:19:24Z
dc.date.accessioned 2022-11-03T00:30:25Z
dc.date.available 2013-12-11T22:19:24Z
dc.date.available 2022-11-03T00:30:25Z
dc.date.copyright 2013
dc.date.issued 2013
dc.identifier.uri https://ir.wgtn.ac.nz/handle/123456789/29385
dc.description.abstract The study of protein misfolding has been a major focus in many ER stress-related diseases, including type I and type II diabetes, neurodegenerative diseases, atherosclerosis and some cancers. There is a basal level of naturally occurring misfolded proteins that are corrected by the unfolded protein response (UPR) system or are degraded by the ER-associated degradation system (ERAD). In pathological conditions where there are increased and enduring amounts of misfolded proteins in the ER, an elevated level of ER stress triggers the UPR signal further. This thesis is about determining whether the same set of genes are involved in basal and elevated levels of ER stress. We developed ER stress reporter assays to measure high levels of ER stress caused by chronic misfolding in a model system, namely a mutant of carboxy peptidase Y called CPY* which does not fold. In this dissertation, genes that are required in basal levels and elevated levels of ER stress are compared in a genome-wide manner using the yeast deletion mutant array (DMA) and modified synthetic genetic array (mSGA) analysis. ER stress reporters used in CPY* over-expression conditions included induction of a general UPR promoter element (UPRE-GFP), induction of two COPII-related proteins (Yip3p and Erv29p) and of a sphingolipid regulatory protein (Orm2p). A novel group of kinetochore genes are required for full activation of all four ER stress reporters when CPY* is over-expressed. Deletion of kinetochore gene in the presence of CPY* over-expression does not cause a general reduction in protein synthesis. Instead, it results in the down-regulation of specific groups of lipid-related and protein trafficking proteins. To explain our results, a checkpoint must be present that monitors the function of the kinetochore or cell division related to ER stress levels, resulting in down regulation of UPR and secretory pathway proteins when the checkpoint is activated. A genome-wide GFP-tagged protein screen revealed that Bfr1p is the possible checkpoint as it is genetically linked with IRE1, a central gene in the UPR, and the spindle pole body. In conditions of CPY* over-expression in the absence of the kinetochore gene CTF19, Bfr1p as well as Scp160p are down-regulated and possibly re-localised to the cytoplasm from the ER. Both of these are mRNA ribosome binding proteins and are specifically required for transporting secretory pathway gene mRNAs to the ER. The reduction in the secretory pathway proteins in the absence of kinetochore genes might be due to reduced levels of Bfr1p and Scp160p expression. Results reported here suggested that PKA activation is another possible pathway that down-regulates the UPR when CPY* is over-expressed in the absence of kinetochore genes. It is also proposed that Bfr1p is a checkpoint that monitors kinetochores and ER stress levels and induces a secondary pathway that is able to work in parallel to IRE-HAC1-UPR. An overall novel conclusion of this thesis is that cells in a chronic misfolding ER-stress situation require intact kinetochores to maintain an elevated UPR. en_NZ
dc.language.iso en_NZ
dc.publisher Te Herenga Waka—Victoria University of Wellington en_NZ
dc.rights Access is restricted to staff and students only until 12/12/2015. en_NZ
dc.subject ER stress en_NZ
dc.subject Unfolded protein response en_NZ
dc.subject Kinetochore en_NZ
dc.title Kinetochores are required to fully activate secretory pathway in elevated ER stress en_NZ
dc.type Text en_NZ
vuwschema.contributor.unit School of Biological Sciences en_NZ
vuwschema.subject.anzsrcfor 110106 Medical Biochemistry: Proteins and Peptides (incl. Medical Proteomics) en_NZ
vuwschema.subject.anzsrcseo 920199 Clinical Health (Organs, Diseases and Abnormal Conditions) not elsewhere classified en_NZ
vuwschema.subject.anzsrcseo 970111 Expanding Knowledge in the Medical and Health Sciences en_NZ
vuwschema.type.vuw Awarded Doctoral Thesis en_NZ
thesis.degree.discipline Biomedical Science en_NZ
thesis.degree.grantor Te Herenga Waka—Victoria University of Wellington en_NZ
thesis.degree.level Doctoral en_NZ
thesis.degree.name Doctor of Philosophy en_NZ
thesis.degree.name Doctor of Philosophy en_NZ

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