A thiol probe for measuring unfolded protein load and proteostasis in cells
AuthorChen, MZ; Moily, NS; Bridgford, JL; Wood, RJ; Radwan, M; Smith, TA; Song, Z; Tang, BZ; Tilley, L; Xu, X; ...
Source TitleNATURE COMMUNICATIONS
PublisherNATURE PUBLISHING GROUP
University of Melbourne Author/sReid, Gavin; Wood, Rebecca; Tilley, Leann; Hatters, Daniel; Smith, Trevor; Hong, Yuning; Moily, Nagaraja Sundara; Bridgford, Jessica; Radwan Ahmed Elsayed, Mona; Moily, Nagaraj
AffiliationMedical Biology (W.E.H.I.)
Biochemistry and Molecular Biology
Microbiology and Immunology
School of Chemistry
Document TypeJournal Article
CitationsChen, MZ; Moily, NS; Bridgford, JL; Wood, RJ; Radwan, M; Smith, TA; Song, Z; Tang, BZ; Tilley, L; Xu, X; Reid, GE; Pouladi, MA; Hong, Y; Hatters, DM, A thiol probe for measuring unfolded protein load and proteostasis in cells, NATURE COMMUNICATIONS, 2017, 8
Access StatusOpen Access
Open Access at PMChttp://www.ncbi.nlm.nih.gov/pmc/articles/PMC5589734
When proteostasis becomes unbalanced, unfolded proteins can accumulate and aggregate. Here we report that the dye, tetraphenylethene maleimide (TPE-MI) can be used to measure cellular unfolded protein load. TPE-MI fluorescence is activated upon labelling free cysteine thiols, normally buried in the core of globular proteins that are exposed upon unfolding. Crucially TPE-MI does not become fluorescent when conjugated to soluble glutathione. We find that TPE-MI fluorescence is enhanced upon reaction with cellular proteomes under conditions promoting accumulation of unfolded proteins. TPE-MI reactivity can be used to track which proteins expose more cysteine residues under stress through proteomic analysis. We show that TPE-MI can report imbalances in proteostasis in induced pluripotent stem cell models of Huntington disease, as well as cells transfected with mutant Huntington exon 1 before the formation of visible aggregates. TPE-MI also detects protein damage following dihydroartemisinin treatment of the malaria parasites Plasmodium falciparum. TPE-MI therefore holds promise as a tool to probe proteostasis mechanisms in disease.Proteostasis is maintained through a number of molecular mechanisms, some of which function to protect the folded state of proteins. Here the authors demonstrate the use of TPE-MI in a fluorigenic dye assay for the quantitation of unfolded proteins that can be used to assess proteostasis on a cellular or proteome scale.
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