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Effects of Inhibition of N-linked Glycosylation on Cellular Autophagy

Needs, Sarah; Alonzi, Dominic; Bootman, Martin and Allman, Sarah (2016). Effects of Inhibition of N-linked Glycosylation on Cellular Autophagy. In: Evolution brings Ca2+ and ATP together to control life and death, 16 - 17 March 2016, Kavli Royal Society Centre, Chicheley Hall, Newport Pagnell.

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Abstract

Accumulation of misfolded proteins can result in severe and varied pathologies. Peptide N-glycanase is an endoglycosidase that cleaves N-linked glycans from incorrectly folded glycoproteins prior to their proteasomal degradation and mutations in peptide N-glycanase can cause result in severe multi-system disorders.

Using z-VAD-fmk (50 µM) as a pharmacological inhibitor of peptide N-glycanase, and Thioflavin T as a fluorescent label for protein aggregates in living cells, we found a biphasic change in protein aggregate accumulation. Z-VAD-fmk induced an increase in protein aggregate accumulation up to 48 hours, which declined to near basal levels after 72 hours. Western blotting showed that BiP, a calcium binding ER stress marker, had a similar temporal expression profile.

Autophagy was increased after 72 hours of peptide N-glycanase inhibition, which corresponded with the decrease of protein aggregates and BiP expression. In ATG13-/- MEFs, which are autophagy deficient, inhibition of peptide N-glycanase by Z-VAD-fmk lead to a 45 % decrease in cell viability within 24 hours. There was no decline of cell viability in Z-VAD-fmk-treated wild type MEFs.

These data indicate that inhibition of peptide N-glycanase leads to protein aggregate accumulation and ER stress, and in cells where peptide N-glycanase function is ablated, autophagy is essential for protein aggregate removal and maintaining cell survival.

Item Type: Conference or Workshop Item
Copyright Holders: 2016 The Authors
Project Funding Details:
Funded Project NameProject IDFunding Body
Not SetNot SetThe Open University (OU)
Keywords: autophagy; glycosylation; protein aggregation; protein folding; quality control; glycoprotein synthesis
Academic Unit/School: Faculty of Science, Technology, Engineering and Mathematics (STEM) > Life, Health and Chemical Sciences
Faculty of Science, Technology, Engineering and Mathematics (STEM)
Other Departments > Research and Academic Strategy
Other Departments
Interdisciplinary Research Centre: Biomedical Research Network (BRN)
Health and Wellbeing PRA (Priority Research Area)
Related URLs:
Item ID: 45936
Depositing User: Sarah Allman
Date Deposited: 04 Apr 2016 09:15
Last Modified: 11 Sep 2017 15:32
URI: http://oro.open.ac.uk/id/eprint/45936
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