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Mitochondrial Translation Efficiency Controls Cytoplasmic Protein Homeostasis
Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics.
Stockholm University, Faculty of Science, Department of Molecular Biosciences, The Wenner-Gren Institute.
Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics.
Stockholm University, Faculty of Science, Department of Molecular Biosciences, The Wenner-Gren Institute.
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Number of Authors: 142018 (English)In: Cell Metabolism, ISSN 1550-4131, E-ISSN 1932-7420, Vol. 27, no 6, p. 1309-1322Article in journal (Refereed) Published
Abstract [en]

Cellular proteostasis ismaintained via the coordinated synthesis, maintenance, and breakdown of proteins in the cytosol and organelles. While biogenesis of the mitochondrial membrane complexes that execute oxidative phosphorylation depends on cytoplasmic translation, it is unknown how translation within mitochondria impacts cytoplasmic proteostasis and nuclear gene expression. Here we have analyzed the effects of mutations in the highly conserved accuracy center of the yeast mitoribosome. Decreased accuracy of mitochondrial translation shortened chronological lifespan, impaired management of cytosolic protein aggregates, and elicited a general transcriptional stress response. In striking contrast, increased accuracy extended lifespan, improved cytosolic aggregate clearance, and suppressed a normally stress-induced, Msn2/4-dependent interor-ganellar proteostasis transcription program (IPTP) that regulates genes important for mitochondrial proteostasis. Collectively, the data demonstrate that cytosolic protein homeostasis and nuclear stress signaling are controlled by mitochondrial translation efficiency in an inter-connected organelle quality control network that determines cellular lifespan.

Place, publisher, year, edition, pages
2018. Vol. 27, no 6, p. 1309-1322
National Category
Biological Sciences
Research subject
Biochemistry; Molecular Bioscience
Identifiers
URN: urn:nbn:se:su:diva-157770DOI: 10.1016/j.cmet.2018.04.011ISI: 000434480000016PubMedID: 29754951OAI: oai:DiVA.org:su-157770DiVA, id: diva2:1223316
Available from: 2018-06-25 Created: 2018-06-25 Last updated: 2019-03-29Bibliographically approved
In thesis
1. Mitochondrial translation and its impact on protein homeostasis and aging
Open this publication in new window or tab >>Mitochondrial translation and its impact on protein homeostasis and aging
2019 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Besides their famous role as powerhouse of the cell, mitochondria are also involved in many signaling processes and metabolism. Therefore, it is unsurprising that mitochondria are no isolated organelles but are in constant crosstalk with other parts of the cell. Due to the endosymbiotic origin of mitochondria, they still contain their own genome and gene expression machinery. The mitochondrial genome of yeast encodes eight proteins whereof seven are core subunits of the respiratory chain and ATP synthase. These subunits need to be assembled with subunits imported from the cytosol to ensure energy supply of the cell. Hence, coordination, timing and accuracy of mitochondrial gene expression is crucial for cellular energy production and homeostasis. Despite the central role of mitochondrial translation surprisingly little is known about the molecular mechanisms.

In this work, I used baker’s yeast Saccharomyces cerevisiae to study different aspects of mitochondrial translation. Exploiting the unique possibility to make directed modifications in the mitochondrial genome of yeast, I established a mitochondrial encoded GFP reporter. This reporter allows monitoring of mitochondrial translation with different detection methods and enables more detailed studies focusing on timing and regulation of mitochondrial translation. Furthermore, employing insights gained from bacterial translation, we showed that mitochondrial translation efficiency directly impacts on protein homeostasis of the cytoplasm and lifespan by affecting stress handling. Lastly, we provided first evidence that mitochondrial protein quality control happens at a very early stage directly after or during protein synthesis at the ribosome. Surveillance of protein synthesis and assembly into complexes is important to avoid accumulation of misfolded or unassembled respiratory chain subunits which would disturb mitochondrial function.

Place, publisher, year, edition, pages
Stockholm: Department of Biochemistry and Biophysics, Stockholm University, 2019. p. 76
Keywords
mitochondrial ribosome, mitochondrial translation accuracy, mitochondrial communication, interorganellar communication, stress signaling, proteostasis, aging, yeast genetics, mitochondrial protein quality control, mitochondrial membrane protein insertion
National Category
Biochemistry and Molecular Biology
Research subject
Biochemistry
Identifiers
urn:nbn:se:su:diva-163149 (URN)978-91-7797-542-7 (ISBN)978-91-7797-543-4 (ISBN)
Public defence
2019-02-15, Magnélisalen, Kemiska övningslaboratoriet, Svante Arrhenius väg 16 B, Stockholm, 09:00 (English)
Opponent
Supervisors
Note

At the time of the doctoral defense, the following paper was unpublished and had a status as follows: Paper 2: Manuscript.

Available from: 2019-01-23 Created: 2018-12-17 Last updated: 2019-01-22Bibliographically approved
2. 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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Suhm, TamaraKaimal, Jayasankar MohanakrishnanDawitz, HannahPeselj, CarlottaMasser, Anna E.Smialowska, AgataBjörck, Markus L.Brzezinski, PeterBüttner, SabrinaAndréasson, ClaesOtt, Martin
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