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Hsp70 nucleotide exchange factor Fes1 is essential for ubiquitin-dependent degradation of misfolded cytosolic proteins
Stockholm University, Faculty of Science, Department of Molecular Biosciences, The Wenner-Gren Institute.
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2013 (English)In: Proceedings of the National Academy of Sciences of the United States of America, ISSN 0027-8424, E-ISSN 1091-6490, Vol. 110, no 15, 5975-5980 p.Article in journal (Refereed) Published
Abstract [en]

Protein quality control systems protect cells against the accumulation of toxic misfolded proteins by promoting their selective degradation. Malfunctions of quality control systems are linked to aging and neurodegenerative disease. Folding of polypeptides is facilitated by the association of 70 kDa Heat shock protein (Hsp70) molecular chaperones. If folding cannot be achieved, Hsp70 interacts with ubiquitylation enzymes that promote the proteasomal degradation of the misfolded protein. However, the factors that direct Hsp70 substrates toward the degradation machinery have remained unknown. Here, we identify Fes1, an Hsp70 nucleotide exchange factor of hitherto unclear physiological function, as a cytosolic triaging factor that promotes proteasomal degradation of misfolded proteins. Fes1 selectively interacts with misfolded proteins bound by Hsp70 and triggers their release from the chaperone. In the absence of Fes1, misfolded proteins fail to undergo polyubiquitylation, aggregate, and induce a strong heat shock response. Our findings reveal that Hsp70 direct proteins toward either folding or degradation by using distinct nucleotide exchange factors.

Place, publisher, year, edition, pages
2013. Vol. 110, no 15, 5975-5980 p.
Keyword [en]
Hsp110, proteasome, triage, ubiquitin ligase, Ubr1
National Category
Cell Biology
Research subject
Cell Biology
URN: urn:nbn:se:su:diva-89195DOI: 10.1073/pnas.1216778110ISI: 000317537900050PubMedID: 23530227OAI: diva2:616361
Swedish Research Council, 621-2009-4421
Available from: 2013-04-16 Created: 2013-04-16 Last updated: 2016-04-22Bibliographically approved
In thesis
1. Regulation of Hsp70 function by nucleotide-exchange factors
Open this publication in new window or tab >>Regulation of Hsp70 function by nucleotide-exchange factors
2016 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Protein folding is the process in which polypeptides in their non-native states attain the unique folds of their native states. Adverse environmental conditions and genetic predisposition challenge the folding process and accelerate the production of proteotoxic misfolded proteins. Misfolded proteins are selectively recognized and removed from the cell by processes of protein quality control (PQC). In PQC molecular chaperones of the Heat shock protein 70 kDa (Hsp70) family play important roles by recognizing and facilitating the removal of misfolded proteins. Hsp70 function is dependent on cofactors that regulate the intrinsic ATPase activity of the chaperone. In this thesis I have used yeast genetic, cell biological and biochemical experiments to gain insight into the regulation of Hsp70 function in PQC by nucleotide-exchange factors (NEFs). Study I shows that the NEF Fes1 is a key factor essential for cytosolic PQC. A reverse genetics approach demonstrated that Fes1 NEF activity is required for the degradation of misfolded proteins associated with Hsp70 by the ubiquitin-proteasome system. Specifically, Fes1 association with Hsp70-substrate complexes promotes interaction of the substrate with downstream ubiquitin E3 ligase Ubr1. The consequences of genetic removal of FES1 (fes1Δ) are the failure to degrade misfolded proteins, the accumulation of protein aggregates and constitutive induction of the heat-shock response. Taken the experimental data together, Fes1 targets misfolded proteins for degradation by releasing them from Hsp70. Study II describes an unusual example of alternative splicing of FES1 transcripts that leads to the expression of the two alternative splice isoforms Fes1S and Fes1L. Both isoforms are functional NEFs but localize to different compartments. Fes1S is localized to the cytosol and is required for the efficient degradation of Hsp70-associated misfolded proteins. In contrast, Fes1L is targeted to the nucleus and represents the first identified nuclear NEF in yeast. The identification of distinctly localized Fes1 isoforms have implications for the understanding of the mechanisms underlying nucleo-cytoplasmic PQC. Study III reports on the mechanism that Fes1 employs to regulate Hsp70 function. Specifically Fes1 carries an N-terminal domain (NTD) that is conserved throughout the fungal kingdom. The NTD is flexible, modular and is required for the cellular function of Fes1. Importantly, the NTD forms ATP-sensitive complexes with Hsp70 suggesting that it competes substrates of the chaperone during Fes1-Hsp70 interactions. Study IV reports on methodological development for the efficient assembly of bacterial protein-expression plasmids using yeast homologous recombination cloning and the novel vector pSUMO-YHRC. The findings support the notion that Fes1 plays a key role in determining the fate of Hsp70-associated misfolded substrates and thereby target them for proteasomal degradation. From a broader perspective, the findings provide information essential to develop models that describe how Hsp70 function is regulated by different NEFs to participate in protein folding and degradation.

Place, publisher, year, edition, pages
Stockholm: Department of Molecular Biosciences, The Wenner-Gren Institute, Stockholm University, 2016. 73 p.
Heat shock proteins (Hsps), Hsp70, Molecular chaperones, Nucleotide-exchange factors (NEFs), Protein quality control (PQC), Proteostasis, Ubiquitin-proteasome system
National Category
Biological Sciences Cell Biology
Research subject
Cell Biology
urn:nbn:se:su:diva-129118 (URN)978-91-7649-376-2 (ISBN)
Public defence
2016-06-10, Högbomsalen, Geovetenskapens hus, Svante Arrhenius väg 12, Stockholm, 10:00 (English)

At the time of the doctoral defense, the following paper was unpublished and had a status as follows: Paper 3: Manuscript.

Available from: 2016-05-18 Created: 2016-04-14 Last updated: 2016-05-04Bibliographically approved

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Gowda, Naveen Kumar ChandappaAndréasson, Claes
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