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Cytoplasmic protein misfolding titrates nuclear Hsp70 to unleash active Hsf1
Stockholm University, Faculty of Science, Department of Molecular Biosciences, The Wenner-Gren Institute.
Stockholm University, Faculty of Science, Department of Molecular Biosciences, The Wenner-Gren Institute. Stockholm University, Science for Life Laboratory (SciLifeLab).
Stockholm University, Faculty of Science, Department of Molecular Biosciences, The Wenner-Gren Institute. Stockholm University, Science for Life Laboratory (SciLifeLab).
Stockholm University, Faculty of Science, Department of Molecular Biosciences, The Wenner-Gren Institute.
(English)Manuscript (preprint) (Other academic)
National Category
Biological Sciences
Research subject
Molecular Bioscience
Identifiers
URN: urn:nbn:se:su:diva-167079OAI: oai:DiVA.org:su-167079DiVA, id: diva2:1296547
Available from: 2019-03-15 Created: 2019-03-15 Last updated: 2019-03-25Bibliographically approved
In thesis
1. Controlling protein homeostasis through regulation of Heat shock factor 1
Open this publication in new window or tab >>Controlling protein homeostasis through regulation of Heat shock factor 1
2019 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

In order to thrive in a changing environment all organisms need to ensure protein homeostasis (proteostasis). Proteostasis is ensured by the proteostasis system that monitors the folding status of the proteome and regulates cell physiology and gene expression to counteract any perturbations. An increased burden on the proteostasis system activates Heat shock factor 1 (Hsf1) to induce transcription of the heat shock response (HSR), a transiently induced transcriptional program including core proteostasis genes, importantly those encoding the Hsp70 class of molecular chaperones. The HSR assists cells in counteracting the harmful effects of protein folding stress and restoring proteostasis. The work presented in this thesis is based on experiments with the Saccharomyces cerevisiae (yeast) model with the overall goal of deciphering how Hsp70 detects and impacts on perturbations of cellular proteostasis and controls Hsf1 activity.

In Study I we describe the fundamental mechanism by which Hsp70 maintains Hsf1 in its latent state by controlling its ability to bind DNA. We found that Hsf1 and unfolded proteins directly compete for binding to the Hsp70 substrate-binding domain. During heat shock the pool of unfolded proteins mainly consist of misfolded, newly synthesized proteins. Severe out-titration of Hsp70 by misfolded substrates resulted in unrestrained Hsf1 activity inducing a previously uncharacterized genetic hyper-stress program. More insight into regulation of Hsp70 availability was gained in Study II where the two splice isoforms of the Hsp70 nucleotide exchange factor Fes1 were characterized. We found that the cytosolic splice isoform Fes1S is crucial to release unfolded proteins from Hsp70 and that impaired release results in strong Hsf1 activation.

In Study III we developed methodology to easily measure the rapid changes in Hsf1 activity upon proteostatic perturbations and to monitor protein turnover using the novel bioluminescent reporter NanoLuc optimized for yeast expression (yNluc). In Study IV we report that yNluc also functions as an in vivo reporter that detects severe perturbations of de novo protein folding by its failure to fold to an active conformation under such conditions.

Finally, in Study V we investigated how organellar proteostasis impacts on the availability of cytosolic Hsp70. We found that a lowered mitochondrial proteostatic load as a result of high translation accuracy extended lifespan and improved cytosolic proteostasis capacity, evidenced by more rapid stress recovery and less sensitivity to toxic misfolded proteins. In contrast, lowered mitochondrial translation accuracy decreased lifespan and impaired management of cytosolic protein aggregates as well as elicited a general transcriptional stress response.

Taken together, the findings presented in this thesis advance our understanding of how the regulatory mechanisms of the proteostasis system function. Furthermore, they provide novel methodology that will facilitate future studies to improve our understanding how cells integrate internal and external stress cues to control proteostasis.

Place, publisher, year, edition, pages
Stockholm: Department of Molecular Biosciences, The Wenner-Gren Institute, Stockholm University, 2019. p. 60
Keywords
stress, protein homeostasis, molecular chaperones, heat shock response, Hsf1, Hsp70, Saccharomyces cerevisiae
National Category
Biochemistry and Molecular Biology Cell Biology
Research subject
Molecular Bioscience
Identifiers
urn:nbn:se:su:diva-167081 (URN)978-91-7797-688-2 (ISBN)978-91-7797-708-7 (ISBN)
Public defence
2019-05-21, Vivi Täckholmsalen (Q-salen), NPQ-huset, Svante Arrhenius väg 20, Stockholm, 10:00 (English)
Opponent
Supervisors
Note

At the time of the doctoral defense, the following papers were unpublished and had a status as follows: Paper 1: Manuscript. Paper 4: Manuscript.

Available from: 2019-04-25 Created: 2019-03-25 Last updated: 2019-04-11Bibliographically approved

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Masser, Anna E.Kang, WenjingFriedländer, Marc R.Andréasson, Claes
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