scholarly journals USP5 enhances SGTA mediated protein quality control

2021 ◽  
Author(s):  
Yvonne Nyathi ◽  
Jake Alfie Hill
Keyword(s):  
Quality Control ◽  
Steady State ◽  
Type Ii Diabetes ◽  
Protein Quality ◽  
Proteasomal Degradation ◽  
Aggregate Formation ◽  
Deubiquitinating Enzymes ◽  
Physiological Processes ◽  
Hydrophobic Amino Acids ◽  
Steady State Levels

Mislocalised membrane proteins (MLPs) present a risk to the cell due to exposed hydrophobic amino acids which cause MLPs to aggregate. Previous studies identified SGTA as a key component of the machinery that regulates the quality control of MLPs. Overexpression of SGTA promotes deubiqutination of MLPs resulting in their accumulation in cytosolic inclusions, suggesting SGTA acts in collaboration with deubiquitinating enzymes (DUBs) to exert these effects.  However, the DUBs that play a role in this process have not been identified.  In this study we have identified the ubiquitin specific peptidase 5 (USP5) as a DUB important in regulating the quality control of MLPs. We show that USP5 is in complex with SGTA, and this association is increased in the presence of an MLP. Overexpression of SGTA results in an increase in steady-state levels of MLPs suggesting a delay in proteasomal degradation of substrates. However, our results show that this effect is strongly dependent on the presence of USP5.  We find that in the absence of USP5, the ability of SGTA to increase the steady state levels of MLPs is compromised. Moreover, knockdown of USP5 results in a reduction in the steady state levels of MLPs, while overexpression of USP5 increases the steady state levels. Our findings suggest that the interaction of SGTA with USP5 enables specific MLPs to escape proteasomal degradation allowing selective modulation of MLP quality control. These findings progress our understanding of aggregate formation, a hallmark in a range of neurodegenerative diseases and type II diabetes, as well as physiological processes of aggregate clearance.

scholarly journals TheMycobacterium tuberculosisPup-proteasome system regulates nitrate metabolism through an essential protein quality control pathway

2019 ◽  
Vol 116 (8) ◽  
pp. 3202-3210 ◽  
Author(s):  
Samuel H. Becker ◽  
Jordan B. Jastrab ◽  
Avantika Dhabaria ◽  
Catherine T. Chaton ◽  
Jeffrey S. Rush ◽  
...  
Keyword(s):  
Quality Control ◽  
Steady State ◽  
Nitrate Reduction ◽  
Protein Quality ◽  
Nitrogen Sources ◽  
Sole Nitrogen Source ◽  
Human Pathogen ◽  
Bacterial Proteins ◽  
Nitrate Metabolism ◽  
Steady State Levels

The human pathogenMycobacterium tuberculosisencodes a proteasome that carries out regulated degradation of bacterial proteins. It has been proposed that the proteasome contributes to nitrogen metabolism inM. tuberculosis, although this hypothesis had not been tested. Upon assessingM. tuberculosisgrowth in several nitrogen sources, we found that a mutant strain lacking theMycobacteriumproteasomal activator Mpa was unable to use nitrate as a sole nitrogen source due to a specific failure in the pathway of nitrate reduction to ammonium. We found that the robust activity of the nitrite reductase complex NirBD depended on expression of thegroEL/groESchaperonin genes, which are regulated by the repressor HrcA. We identified HrcA as a likely proteasome substrate, and propose that the degradation of HrcA is required for the full expression of chaperonin genes. Furthermore, our data suggest that degradation of HrcA, along with numerous other proteasome substrates, is enhanced during growth in nitrate to facilitate the derepression of the chaperonin genes. Importantly, growth in nitrate is an example of a specific condition that reduces the steady-state levels of numerous proteasome substrates inM. tuberculosis.

scholarly journals Mapping the degradation pathway of a disease-linked aspartoacylase variant

PLoS Genetics ◽  
2021 ◽  
Vol 17 (4) ◽  
pp. e1009539
Author(s):  
Sarah K. Gersing ◽  
Yong Wang ◽  
Martin Grønbæk-Thygesen ◽  
Caroline Kampmeyer ◽  
Lene Clausen ◽  
...  
Keyword(s):  
Quality Control ◽  
Protein Quality ◽  
Protein Quality Control ◽  
Proteasomal Degradation ◽  
Degradation Pathway ◽  
Wild Type ◽  
Protein Variant ◽  
Stable Conformation ◽  
Spongy Degeneration ◽  
Brain White Matter

Canavan disease is a severe progressive neurodegenerative disorder that is characterized by swelling and spongy degeneration of brain white matter. The disease is genetically linked to polymorphisms in the aspartoacylase (ASPA) gene, including the substitution C152W. ASPA C152W is associated with greatly reduced protein levels in cells, yet biophysical experiments suggest a wild-type like thermal stability. Here, we use ASPA C152W as a model to investigate the degradation pathway of a disease-causing protein variant. When we expressed ASPA C152W in Saccharomyces cerevisiae, we found a decreased steady state compared to wild-type ASPA as a result of increased proteasomal degradation. However, molecular dynamics simulations of ASPA C152W did not substantially deviate from wild-type ASPA, indicating that the native state is structurally preserved. Instead, we suggest that the C152W substitution interferes with the de novo folding pathway resulting in increased proteasomal degradation before reaching its stable conformation. Systematic mapping of the protein quality control components acting on misfolded and aggregation-prone species of C152W, revealed that the degradation is highly dependent on the molecular chaperone Hsp70, its co-chaperone Hsp110 as well as several quality control E3 ubiquitin-protein ligases, including Ubr1. In addition, the disaggregase Hsp104 facilitated refolding of aggregated ASPA C152W, while Cdc48 mediated degradation of insoluble ASPA protein. In human cells, ASPA C152W displayed increased proteasomal turnover that was similarly dependent on Hsp70 and Hsp110. Our findings underscore the use of yeast to determine the protein quality control components involved in the degradation of human pathogenic variants in order to identify potential therapeutic targets.

scholarly journals The nucleus is a quality control center for non-imported mitochondrial proteins

2020 ◽  
Author(s):  
Viplendra P.S. Shakya ◽  
William A. Barbeau ◽  
Tianyao Xiao ◽  
Christina S. Knutson ◽  
Adam L. Hughes
Keyword(s):  
Quality Control ◽  
Steady State ◽  
Nuclear Protein ◽  
Protein Quality ◽  
Mitochondrial Proteins ◽  
Mitochondrial Targeting ◽  
Mitochondrial Import ◽  
Control Center ◽  
Mitochondrial Impairment ◽  
Precursor Proteins

AbstractMitochondrial import deficiency causes cellular stress due to the accumulation of non-imported mitochondrial precursor proteins. Despite the burden mis-localized mitochondrial precursors place on cells, our understanding of the systems that dispose of these proteins is incomplete. Here, we catalog the location and steady-state abundance of mitochondrial precursor proteins during mitochondrial impairment in S. cerevisiae. We find that a number of non-imported mitochondrial proteins localize to the nucleus, where they are eliminated by proteasome-based nuclear protein quality control. Recognition of mitochondrial precursors by the nuclear quality control machinery requires the presence of an N-terminal mitochondrial targeting sequence (MTS), and impaired breakdown of precursors leads to their buildup in nuclear-associated foci. These results identify the nucleus as a key destination for the disposal of non-imported mitochondrial precursors.

scholarly journals Evolutionarily conserved chaperone-mediated proteasomal degradation of a disease-linked aspartoacylase variant

2020 ◽  
Author(s):  
Sarah K. Gersing ◽  
Yong Wang ◽  
Martin Grønbæk-Thygesen ◽  
Caroline Kampmeyer ◽  
Lene Clausen ◽  
...  
Keyword(s):  
Quality Control ◽  
Protein Quality ◽  
De Novo ◽  
Neurodegenerative Disorder ◽  
Canavan Disease ◽  
Proteasomal Degradation ◽  
Degradation Pathway ◽  
Wild Type ◽  
Spongy Degeneration ◽  
Brain White Matter

AbstractCanavan disease is a severe progressive neurodegenerative disorder that is characterized by swelling and spongy degeneration of brain white matter. The disease is genetically linked to polymorphisms in the aspartoacylase (ASPA) gene, including the substitution C152W. ASPA C152W is associated with greatly reduced protein levels in cells, yet biophysical experiments suggest a wild-type like thermal stability. Here, we examine the stability and degradation pathway of ASPA C152W. When we expressed ASPA C152W in Saccharomyces cerevisiae, we found a decreased steady state compared to wild-type ASPA as a result of increased proteasomal degradation. However, molecular dynamics simulations of ASPA C152W did not substantially deviate from wild-type ASPA, indicating that the native state is structurally preserved. Instead, we suggest that the C152W substitution prevents ASPA from reaching its stable native conformation, presumably by impacting on de novo folding. Systematic mapping of the protein quality control components acting on misfolded and aggregation-prone species of C152W, revealed that the degradation is highly dependent on the molecular chaperone Hsp70, its co-chaperone Hsp110 as well as several quality control E3 ubiquitin-protein ligases, including Ubr1. In human cells, ASPA C152W displayed increased proteasomal turnover that was similarly dependent on Hsp70 and Hsp110. We propose that Hsp110 is a potential therapeutic target for misfolding ASPA variants that trigger Canavan disease due to excessive degradation.

scholarly journals Comparative analysis of follistatin-, activin beta A- and activin beta B-mRNA steady-state levels in diverse porcine tissues by multiplex S1 nuclease analysis

2000 ◽  
pp. 537-544 ◽  
Author(s):  
O Schneider ◽  
R Nau ◽  
U Michel
Keyword(s):  
Steady State ◽  
Adrenal Gland ◽  
Mrna Expression ◽  
Cell Types ◽  
Mrna Levels ◽  
S1 Nuclease ◽  
Physiological Processes ◽  
Mrna Tissue Distribution ◽  
Steady State Levels ◽  
Porcine Tissues

OBJECTIVE: The relation of activins (dimers of the beta-subunits of inhibin) and follistatin (FS) (their binding protein) affect the growth and differentiation of many cell types. Activin- and FS-mRNAs show a widespread co-expression throughout the organism, indicating an essential role for the FS/activin system in diverse physiological processes. The present study was performed to investigate FS-, activin betaA-, and activin beta B-mRNA expression in porcine tissues and to compare the relative mRNA tissue distribution by a newly developed multiplex S1 nuclease protection assay. METHODS: Twenty micrograms total RNA from different porcine tissues were subjected to multiplex S1 analysis. Specific mRNA expression was determined by measurements of optical densities on autoradiographs. RESULTS: Activin beta A-mRNA expression was abundant in the ovary, adrenal gland, fat, vein, artery and uterus, activin beta B-mRNA was highly expressed in the ovary, pituitary, uterus, placenta, aorta and cerebellum. FS-mRNA showed a widespread expression with high levels in ovary, uterus, cerebellum, placenta and fat. The comparison of relative activin beta A-, activin beta B- and FS-mRNA expression within a certain tissue showed a predominance of activin beta A-mRNA in the adrenal gland, fat, artery, spinal cord, cerebrum and colon and of activin beta B-m RNA in pituitary, testis and placenta, while FS-mRNA levels exceeded those of activin subunits in epididymis, liver, lymphoid tissue, muscle, intestine, cerebellum, ovary and uterus. CONCLUSIONS: The presented data provide an overview of FS-, activin beta A-, and activin beta B-mRNA steady state levels in porcine tissues.

scholarly journals The extent of Ssa1/Ssa2 Hsp70 chaperone involvement in nuclear protein quality control degradation varies with the substrate

2020 ◽  
Vol 31 (3) ◽  
pp. 221-233 ◽  
Author(s):  
Ramon D. Jones ◽  
Charisma Enam ◽  
Rebeca Ibarra ◽  
Heather R. Borror ◽  
Kaitlyn E. Mostoller ◽  
...  
Keyword(s):  
Quality Control ◽  
Nuclear Protein ◽  
Protein Quality ◽  
Protein Quality Control ◽  
Proteasomal Degradation ◽  
Misfolded Proteins ◽  
Hsp70 Chaperone

The ubiquitin-mediated proteasomal degradation of misfolded proteins is generally thought to require Hsp70 chaperones, particularly Ssa1 and Ssa2 in yeast. This study reveals that Ssa1/Ssa2 are involved in the degradation of misfolded proteins in the yeast nucleus, but their degree of involvement varies depending on the misfolded nuclear protein.

scholarly journals Deubiquitinases USP20/33 promote the biogenesis of tail-anchored membrane proteins

2021 ◽  
Vol 220 (5) ◽  
Author(s):  
Jacob A. Culver ◽  
Malaiyalam Mariappan
Keyword(s):  
Quality Control ◽  
Membrane Proteins ◽  
Protein Quality ◽  
Protein Quality Control ◽  
Proteasomal Degradation ◽  
Transmembrane Domains ◽  
Misfolded Proteins ◽  
Hydrophobic Membrane ◽  
Cytosolic Protein ◽  
Ta Proteins

Numerous proteins that have hydrophobic transmembrane domains (TMDs) traverse the cytosol and posttranslationally insert into cellular membranes. It is unclear how these hydrophobic membrane proteins evade recognition by the cytosolic protein quality control (PQC), which typically recognizes exposed hydrophobicity in misfolded proteins and marks them for proteasomal degradation by adding ubiquitin chains. Here, we find that tail-anchored (TA) proteins, a vital class of membrane proteins, are recognized by cytosolic PQC and are ubiquitinated as soon as they are synthesized in cells. Surprisingly, the ubiquitinated TA proteins are not routed for proteasomal degradation but instead are handed over to the targeting factor, TRC40, and delivered to the ER for insertion. The ER-associated deubiquitinases, USP20 and USP33, remove ubiquitin chains from TA proteins after their insertion into the ER. Thus, our data suggest that deubiquitinases rescue posttranslationally targeted membrane proteins that are inappropriately ubiquitinated by PQC in the cytosol.

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