Abstract 393: The Intercalated Disc Protein Myozap: A Novel Player in Cardiac Proteinopathy

2016 ◽  
Vol 119 (suppl_1) ◽  
Author(s):  
Ashraf Y Rangrez ◽  
Derk Frank ◽  
Liam Cassidy ◽  
Lynn Christen ◽  
Inka Geurink ◽  
...  
Keyword(s):  
Confocal Microscopy ◽  
Protein Aggregation ◽  
Protein Quality ◽  
Transcriptome Profiling ◽  
Protein Aggregates ◽  
Intercalated Disc ◽  
Protein Aggregate ◽  
General Role ◽  
Id Protein

Background: A growing number of cardiac muscle diseases are characterized by depositions of misfolded proteins, including cardiac amyloidosis and desmin-releated cardiomyopathy (DRM). The continued presence and chronic accumulation of misfolded or unfolded proteins can lead to aggregation and/or the formation of soluble peptides that are proteotoxic. This in turn leads to compromised protein quality control and precipitates a downward spiral of the cell’s ability to maintain homeostasis and may eventually result in cell death. We recently identified massive protein aggregates in the hearts of transgenic mice overexpressing the intercalated disc (ID) protein myozap (Myozap-tg). We now sought to investigate the precise composition of these aggregates and the role of Myozap in other proteinopathies such as DRM. Methods and Results: We employed multi-dimensional proteomics, transcriptomics, confocal microscopy, and molecular biology approaches to decipher the underlying causes and consequences of protein aggregate formation in Myozap-tg mice. Transcriptome profiling of these mice revealed striking upregulation of autophagy, protein synthesis, and pro-inflammatory pathways, whereas protein degradation pathways were down-regulated. Surprisingly, proteomics analyses revealed Desmin and α-crystallin B (CryAB) as the major constituents of the aggregates, which was further validated by confocal microscopy. Moreover, we identified the presence of toxic preamyloid oligomers in Myozap-tg mouse hearts, a hallmark in many protein aggregation-based diseases including DRM. Most interestingly, we also observed co-localization of Myozap with protein aggregates observed in both transgenic mouse hearts overexpressing mutant Desmin (D7) and mutant CryAB (R120G), as well as in human DRM patients. Conclusion: The present study implies a new role for Myozap, which was previously reported to affect cardiac SRF signaling: (1) Myozap accumulates in various forms of experimental and human protein aggregation cardiomyopathy, suggesting involvement in protein homoestasis. (2) The fact that Myozap is now the third ID protein (after desmin and CryAB) to cause cardiac proteinopathy points to a general role of the ID in its molecular pathogenesis.

scholarly journals Nα-terminal acetylation of proteins by NatA and NatB serves distinct physiological roles in Saccharomyces cerevisiae

2019 ◽  
Author(s):  
Ulrike A. Friedrich ◽  
Mostafa Zedan ◽  
Bernd Hessling ◽  
Kai Fenzl ◽  
Ludovic Gillet ◽  
...  
Keyword(s):  
Stress Responses ◽  
Ribosomal Proteins ◽  
Protein Modification ◽  
Protein Quality ◽  
Biological Significance ◽  
General Role ◽  
Altered Expression ◽  
Phenotypic Differences ◽  
Genes Encoding

SummaryN-terminal (Nt)-acetylation is a highly prevalent co-translational protein modification in eukaryotes, catalyzed by at least five Nt-acetyltransferases (Nat) with differing specificities. Nt-acetylation has been implicated in protein quality control but its broad biological significance remains elusive. We investigated the roles of the two major Nats of S. cerevisiae, NatA and NatB, by performing transcriptome, translatome and proteome profiling of natAΔ and natBΔ mutants. Our results do not support a general role of Nt-acetylation in protein degradation but reveal an unexpected range of Nat-specific phenotypes. NatA is implicated in systemic adaptation control, as natAΔ mutants display altered expression of transposons, sub-telomeric genes, pheromone response genes and nuclear genes encoding mitochondrial ribosomal proteins. NatB predominantly affects protein folding, as natBΔ mutants accumulate protein aggregates, induce stress responses and display reduced fitness in absence of the ribosome-associated chaperone Ssb. These phenotypic differences indicate that controlling Nat activities may serve to elicit distinct cellular responses.

scholarly journals FLNC-Associated Myofibrillar Myopathy

2021 ◽  
Vol 7 (3) ◽  
pp. e590
Author(s):  
Rudolf Andre Kley ◽  
Yvonne Leber ◽  
Bertold Schrank ◽  
Heidi Zhuge ◽  
Zacharias Orfanos ◽  
...  
Keyword(s):  
Protein Aggregation ◽  
Protein Quality ◽  
Mutational Analysis ◽  
Protein Aggregates ◽  
Ubiquitin Proteasome System ◽  
Protein Accumulation ◽  
Control Mechanisms ◽  
Myofibrillar Myopathy ◽  
Dimerization Domain ◽  
Indel Mutation

ObjectiveTo determine whether a new indel mutation in the dimerization domain of filamin C (FLNc) causes a hereditary myopathy with protein aggregation in muscle fibers, we clinically and molecularly studied a German family with autosomal dominant myofibrillar myopathy (MFM).MethodsWe performed mutational analysis in 3 generations, muscle histopathology, and proteomic studies of IM protein aggregates. Functional consequences of the FLNC mutation were investigated with interaction and transfection studies and biophysics molecular analysis.ResultsEight patients revealed clinical features of slowly progressive proximal weakness associated with a heterozygous c.8025_8030delCAAGACinsA (p.K2676Pfs*3) mutation in FLNC. Two patients exhibited a mild cardiomyopathy. MRI of skeletal muscle revealed lipomatous changes typical for MFM with FLNC mutations. Muscle biopsies showed characteristic MFM findings with protein aggregation and lesion formation. The proteomic profile of aggregates was specific for MFM-filaminopathy and indicated activation of the ubiquitin-proteasome system (UPS) and autophagic pathways. Functional studies revealed that mutant FLNc is misfolded, unstable, and incapable of forming homodimers and heterodimers with wild-type FLNc.ConclusionsThis new MFM-filaminopathy family confirms that expression of mutant FLNC leads to an adult-onset muscle phenotype with intracellular protein accumulation. Mutant FLNc protein is biochemically compromised and leads to dysregulation of protein quality control mechanisms. Proteomic analysis of MFM protein aggregates is a potent method to identify disease-relevant proteins, differentiate MFM subtypes, evaluate the relevance of gene variants, and identify novel MFM candidate genes.

scholarly journals Intercellular Spread of Protein Aggregates in Neurodegenerative Disease

2018 ◽  
Vol 34 (1) ◽  
pp. 545-568 ◽  
Author(s):  
Albert A. Davis ◽  
Cheryl E.G. Leyns ◽  
David M. Holtzman
Keyword(s):  
Neurodegenerative Diseases ◽  
Protein Aggregation ◽  
Neurodegenerative Disease ◽  
Extracellular Space ◽  
Protein Aggregates ◽  
Therapeutic Strategies ◽  
Disease Pathogenesis ◽  
Donor Cells ◽  
Intercellular Spread

Most neurodegenerative diseases are characterized by the accumulation of protein aggregates, some of which are toxic to cells. Mounting evidence demonstrates that in several diseases, protein aggregates can pass from neuron to neuron along connected networks, although the role of this spreading phenomenon in disease pathogenesis is not completely understood. Here we briefly review the molecular and histopathological features of protein aggregation in neurodegenerative disease, we summarize the evidence for release of proteins from donor cells into the extracellular space, and we highlight some other mechanisms by which protein aggregates might be transmitted to recipient cells. We also discuss the evidence that supports a role for spreading of protein aggregates in neurodegenerative disease pathogenesis and some limitations of this model. Finally, we consider potential therapeutic strategies to target spreading of protein aggregates in the treatment of neurodegenerative diseases.

scholarly journals Aggrephagy: Selective Disposal of Protein Aggregates by Macroautophagy

2012 ◽  
Vol 2012 ◽  
pp. 1-21 ◽  
Author(s):  
Trond Lamark ◽  
Terje Johansen
Keyword(s):  
Protein Aggregation ◽  
Continuous Process ◽  
Protein Aggregates ◽  
Aggregate Formation ◽  
Selective Degradation ◽  
Chaperone Mediated Autophagy ◽  
Single Peptide

Protein aggregation is a continuous process in our cells. Some proteins aggregate in a regulated manner required for different vital functional processes in the cells whereas other protein aggregates result from misfolding caused by various stressors. The decision to form an aggregate is largely made by chaperones and chaperone-assisted proteins. Proteins that are damaged beyond repair are degraded either by the proteasome or by the lysosome via autophagy. The aggregates can be degraded by the proteasome and by chaperone-mediated autophagy only after dissolution into soluble single peptide species. Hence, protein aggregates as such are degraded by macroautophagy. The selective degradation of protein aggregates by macroautophagy is called aggrephagy. Here we review the processes of aggregate formation, recognition, transport, and sequestration into autophagosomes by autophagy receptors and the role of aggrephagy in different protein aggregation diseases.

scholarly journals Mitochondrial fission facilitates the selective mitophagy of protein aggregates

2017 ◽  
Vol 216 (10) ◽  
pp. 3231-3247 ◽  
Author(s):  
Jonathon L. Burman ◽  
Sarah Pickles ◽  
Chunxin Wang ◽  
Shiori Sekine ◽  
Jose Norberto S. Vargas ◽  
...  
Keyword(s):  
Protein Aggregation ◽  
Matrix Protein ◽  
Mitochondrial Fission ◽  
Protein Aggregates ◽  
Mitochondrial Matrix ◽  
Unfolded Protein ◽  
Protein Response ◽  
Mitochondrial Networks ◽  
Mitochondrial Unfolded Protein Response

Within the mitochondrial matrix, protein aggregation activates the mitochondrial unfolded protein response and PINK1–Parkin-mediated mitophagy to mitigate proteotoxicity. We explore how autophagy eliminates protein aggregates from within mitochondria and the role of mitochondrial fission in mitophagy. We show that PINK1 recruits Parkin onto mitochondrial subdomains after actinonin-induced mitochondrial proteotoxicity and that PINK1 recruits Parkin proximal to focal misfolded aggregates of the mitochondrial-localized mutant ornithine transcarbamylase (ΔOTC). Parkin colocalizes on polarized mitochondria harboring misfolded proteins in foci with ubiquitin, optineurin, and LC3. Although inhibiting Drp1-mediated mitochondrial fission suppresses the segregation of mitochondrial subdomains containing ΔOTC, it does not decrease the rate of ΔOTC clearance. Instead, loss of Drp1 enhances the recruitment of Parkin to fused mitochondrial networks and the rate of mitophagy as well as decreases the selectivity for ΔOTC during mitophagy. These results are consistent with a new model that, instead of promoting mitophagy, fission protects healthy mitochondrial domains from elimination by unchecked PINK1–Parkin activity.

scholarly journals A Novel Mechanism for NF-κB-Activation via IκB-Aggregation: Implications for Hepatic Mallory-denk-body Induced Inflammation

2019 ◽  
Author(s):  
Yi Liu ◽  
Michael J. Trnka ◽  
Shenheng Guan ◽  
Doyoung Kwon ◽  
Do-Hyung Kim ◽  
...  
Keyword(s):  
Neurodegenerative Diseases ◽  
Protein Aggregation ◽  
Therapeutic Intervention ◽  
Liver Diseases ◽  
Cell Model ◽  
Protein Aggregates ◽  
Protein Aggregate ◽  
Similar Protein ◽  
Novel Targets ◽  
Proteomic Analyses

ABSTRACTBackground & AimsMallory-Denk-bodies (MDBs) are hepatic protein aggregates associated with inflammation both clinically and in MDB-inducing models. Similar protein aggregation in neurodegenerative diseases also triggers inflammation and NF-κB activation. However, the precise mechanism that links protein aggregation to NFκB-activation and inflammatory response remains unclear.MethodsHerein, we find that treating primary hepatocytes with MDB-inducing agents (N-methylprotoporphyrin, protoporphyrin IX (PPIX), or ZnPPIX) elicited an IκBα-loss with consequent NF-κB activation. We characterized the underlying mechanism in detail using hepatocytes from various knockout mice and MEF cell lines and multiple approaches including immunoblotting, EMSA, RT-PCR, confocal immunofluorescence microscopy, affinity immunoprecipitation, and protein solubility assays. Additionally, we performed rigorous proteomic analyses to identify the proteins aggregating upon PPIX treatment and/or co-aggregating with IκBα.ResultsFour known mechanisms of IκBα-loss were probed and excluded. Immunofluorescence analyses of ZnPPIX-treated cells coupled with 8 M urea/CHAPS-extraction revealed that this IκBα-loss was due to its sequestration along with IκBβ into insoluble aggregates. Through proteomic analyses we identified 47 aggregation-prone proteins that co-aggregate with IκBα through direct interaction or proximity. Of these ZnPPIX-aggregation targets, the nucleoporins Nup153 and Nup358/RanBP2 were identified through RNA-interference, as likely mediators of IκBα-nuclear import.ConclusionWe discovered a novel mechanism of inflammatory NF-κB activation through IκB-sequestration into insoluble aggregates along with interacting aggregation-prone proteins. This mechanism may account for the protein aggregate-induced inflammation observed in MDB-associated liver diseases, thereby identifying novel targets for therapeutic intervention. Because of inherent commonalities this MDB cell model is abona fideprotoporphyric model, making these findings equally relevant to the liver inflammation associated with clinical protoporphyria.Lay SummaryMallory-Denk-bodies (MDBs) are hepatic protein aggregates commonly featured in many liver diseases. MDB-presence is associated with the induction of inflammatory responses both clinically and in all MDB-inducing models. Similar protein aggregation in neurodegenerative diseases is also known to trigger inflammation and NFκB pathway activation via an as yet to be characterized non-canonical mechanism. Herein using a MDB-inducing cell model, we uncovered a novel mechanism for NFκB activation via cytosolic IκB-sequestration into insoluble aggregates. Furthermore, using a proteomic approach, we identified 47 aggregation-prone proteins that interact and co-aggregate with IκBα. This novel mechanism may account for the protein aggregate-induced inflammation observed in liver diseases, thereby identifying novel targets for therapeutic intervention.

scholarly journals Protein Aggregation after Focal Brain Ischemia and Reperfusion

2001 ◽  
Vol 21 (7) ◽  
pp. 865-875 ◽  
Author(s):  
Bing-Ren Hu ◽  
Shorena Janelidze ◽  
Myron D. Ginsberg ◽  
Raul Busto ◽  
Miguel Perez-Pinzon ◽  
...  
Keyword(s):  
Confocal Microscopy ◽  
Protein Aggregation ◽  
Brain Ischemia ◽  
Neuronal Death ◽  
Laser Scanning ◽  
Protein Aggregates ◽  
Laser Scanning Confocal Microscopy ◽  
Artery Occlusion ◽  
Western Blots ◽  
Focal Brain Ischemia

Two hours of transient focal brain ischemia causes acute neuronal death in the striatal core region and a somewhat more delayed type of neuronal death in neocortex. The objective of the current study was to investigate protein aggregation and neuronal death after focal brain ischemia in rats. Brain ischemia was induced by 2 hours of middle cerebral artery occlusion. Protein aggregation was analyzed by electron microscopy, laser-scanning confocal microscopy, and Western blotting. Two hours of focal brain ischemia induced protein aggregation in ischemic neocortical neurons at 1 hour of reperfusion, and protein aggregation persisted until neuronal death at 24 hours of reperfusion. Protein aggregates were found in the neuronal soma, dendrites, and axons, and they were associated with intracellular membranous structures during the postischemic phase. High-resolution confocal microscopy showed that clumped protein aggregates surrounding nuclei and along dendrites were formed after brain ischemia. On Western blots, ubiquitinated proteins (ubi-proteins) were dramatically increased in neocortical tissues in the postischemic phase. The ubi-proteins were Triton-insoluble, indicating that they might be irreversibly aggregated. The formation of ubi-protein aggregates after ischemia correlated well with the observed decrease in free ubiquitin and neuronal death. The authors concluded that proteins are severely damaged and aggregated in neurons after focal ischemia. The authors propose that protein damage or aggregation may contribute to ischemic neuronal death.

scholarly journals Reversion of protein aggregation mediated by Sso7d in cell extracts of Sulfolobus solfataricus

2004 ◽  
Vol 381 (1) ◽  
pp. 249-255 ◽  
Author(s):  
Annamaria GUAGLIARDI ◽  
Lucia MANCUSI ◽  
Mosè ROSSI
Keyword(s):  
Escherichia Coli ◽  
Protein Aggregation ◽  
Atp Hydrolysis ◽  
Sulfolobus Solfataricus ◽  
Protein Aggregates ◽  
Eukaryotic Cells ◽  
Cell Extracts ◽  
Thermophilic Archaeon

In eukaryotic cells and in Escherichia coli, reversion of protein aggregation is mediated by the network of chaperones belonging to Hsp70 and Hsp100 families [Weibezahn, Bukau and Mogk (2004) Microb. Cell Fact. 3, 1–12]. The thermophilic prokaryotes of the archaea domain lack homologues of these chaperone families, and the mechanisms they use to rescue aggregated proteins are unknown [Macario, Malz and Conway de Macario (2004) Front. Biosci. 9, 1318–1332]. In the present study, we show that stable protein aggregates can be detected in extracts of starved cells of the thermophilic archaeon Sulfolobus solfataricus, and that the protein Sso7d interacts with the aggregates and mediates the disassembly of the aggregates and the re-activation of insolubilized β-glycosidase in the presence of ATP hydrolysis. Furthermore, we report that heat-induced protein aggregates in extracts of exponential cells of S. solfataricus contain Sso7d that rescues insolubilized proteins in the presence of ATP hydrolysis. Results of these experiments performed in cell extracts are consistent with an in vivo role of Sso7d in reverting protein aggregation.

scholarly journals Identification of subfunctionalized aggregate-remodeling J-domain proteins in plants

2020 ◽  
Author(s):  
Yogesh Tak ◽  
Silviya S. Lal ◽  
Shilpa Gopan ◽  
Madhumitha Balakrishnan ◽  
Amit K. Verma ◽  
...  
Keyword(s):  
Protein Quality ◽  
Expression Patterns ◽  
Protein Aggregates ◽  
Cellular Protein ◽  
Multiple Model ◽  
Protein Aggregate ◽  
Cytotoxic Protein ◽  
J Domain ◽  
Critical Components

AbstractHsp70s and J-domain proteins (JDPs) are among the most critical components of the cellular protein quality control machinery, playing crucial roles in preventing and solubilizing cytotoxic protein aggregates. Bacteria, yeast and plants additionally have large, multimeric Hsp100-class disaggregases which, allow the resolubilization of otherwise “dead-end” aggregates, including amyloids. JDPs interact with aggregated proteins and specify the aggregate remodeling activities of Hsp70s and Hsp100s. Plants have a complex network of cytosolic Hsp70s and JDPs, however the aggregate remodeling properties of plant JDPs are not well understood. Here we identify evolutionary-conserved Class II JDPs in the model plant Arabidopsis thaliana with distinct aggregate remodeling functionalities. We identify eight plant orthologs of the yeast protein, Sis1, the principal JDP responsible for directing the yeast chaperone machinery for remodeling protein aggregates. Expression patterns vary dramatically among the eight paralogous proteins under a variety of stress conditions, indicating their subfunctionalization to address distinct stressors. Consistent with a role in solubilizing cytotoxic protein aggregates, six of these plant JDPs associate with heat-induced protein aggregates in vivo as well as colocalize with plant Hsp101 to distinct heat-induced protein aggregate centers. Finally, we show that these six JDPs can differentially remodel multiple model protein aggregates in yeast confirming their involvement in aggregate resolubilization. These results demonstrate that compared to complex metazoans, plants have a robust network of JDPs involved in aggregate remodeling activities with the capacity to process a variety of protein aggregate conformers.

scholarly journals Protein aggregation in bacteria

2019 ◽  
Vol 44 (1) ◽  
pp. 54-72 ◽  
Author(s):  
Frederic D Schramm ◽  
Kristen Schroeder ◽  
Kristina Jonas
Keyword(s):  
Protein Aggregation ◽  
Stress Resistance ◽  
Protein Quality ◽  
Protein Aggregates ◽  
Protein Homeostasis ◽  
Bacterial Protein ◽  
Bacterial Populations ◽  
Regulate Gene Expression ◽  
Specific Proteins ◽  
Cellular Stresses

ABSTRACT Protein aggregation occurs as a consequence of perturbations in protein homeostasis that can be triggered by environmental and cellular stresses. The accumulation of protein aggregates has been associated with aging and other pathologies in eukaryotes, and in bacteria with changes in growth rate, stress resistance and virulence. Numerous past studies, mostly performed in Escherichia coli, have led to a detailed understanding of the functions of the bacterial protein quality control machinery in preventing and reversing protein aggregation. However, more recent research points toward unexpected diversity in how phylogenetically different bacteria utilize components of this machinery to cope with protein aggregation. Furthermore, how persistent protein aggregates localize and are passed on to progeny during cell division and how their presence impacts reproduction and the fitness of bacterial populations remains a controversial field of research. Finally, although protein aggregation is generally seen as a symptom of stress, recent work suggests that aggregation of specific proteins under certain conditions can regulate gene expression and cellular resource allocation. This review discusses recent advances in understanding the consequences of protein aggregation and how this process is dealt with in bacteria, with focus on highlighting the differences and similarities observed between phylogenetically different groups of bacteria.

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