INVESTIGATING THE ROLE OF SGTA IN PROTEIN QUALITY CONTROL

UoM administered thesis: Phd

  • Authors:
  • Silvia Dalba

Abstract

Mislocalised membrane and secretory proteins (MLPs) that fail to reach their final destination in the cell may undergo non-native interactions, resulting in cytoplasmic aggregates that are often toxic. In order to reduce the cellular damage caused by such aberrant polypeptides, mammalian cells have developed a specialised protein quality control network to deal with such substrates. This network of cytosolic chaperones and enzymes regulates the selective ubiquitination of hydrophobic MLPs in order to control their degradation. SGTA represents an important component of this quality control network since it promotes the deubiquitination of MLP substrates, thereby providing an extended opportunity for their successful protein folding and/or ER delivery. In order to study the role of SGTA during MLP quality control, I have refined model substrates derived from fragments of opsin and shown that SGTA can act on both canonically and non-canonically ubiquitinated substrates. In order to better define SGTA function, I have identified additional cytosolic quality control factors by performing a BIO-ID proximity assay. Amongst a number of candidates identified using this approach, I found that a number of molecular chaperones were associated with SGTA and I elected to investigate the significance of the Hsp90s in relation to MLP quality control. I found that, like SGTA, a specific isoform of Hsp90 (Hsp90β) regulates steady state MLP levels, most likely by protecting them from rapid proteasomal degradation. Furthermore, the stabilisation of MLPs that is observed following SGTA overexpression is selectively reversed by knocking down Hsp90β. We therefore conclude that SGTA can act synergistically with Hsp90β to regulate MLP quality control. To further define the functional interaction between SGTA and Hsp90β, I studied pVHL, a marginally stabile cytosolic protein known to utilise Hsp90 during its biogenesis and folding. My data suggest that cellular SGTA levels influence the ubiquitination and/or proteasomal degradation of pVHL, thereby expanding the range of clients that are dealt with via its protein quality control network.

Details

Original languageEnglish
Awarding Institution
Supervisors/Advisors
Award date31 Dec 2017