RESUMEN

Lewy body disease (LBD), one of the most common neurodegenerative diseases (NDDs), is characterized by excessive accumulation of α-synuclein (α-syn) in neurons. In recent years, environmental factors such as exposure to herbicides and pesticides have been attributed to the development of this condition. While majority of the studies on neurotoxic effects of paraquat (PQ) have focused on α-syn-mediated neuronal damage in the early stages of α-syn accumulation in neurons, efforts to explore the key target for α-syn degradation are limited. Recent research has suggested that histone deacetylase 6 (HDAC6) might possibly regulate amyloid clearance, and that the metabolism of compounds in neurons is also directly affected by axonal transport in neurons. Dynein predominantly mediates reverse transportation of metabolites and uptake of signal molecules and other compounds at the end of axons, which is conducive to the reuse of cell components. However, the role of interaction of dynein with HDAC6 in metabolites transport is still unclear. Therefore, this study aimed to investigate the role of HDAC6 in α-syn accumulation/clearance in neurons and the associated possible influencing factors. The results revealed that HDAC6 could transport ubiquitinated α-syn, bind to dynein, form an aggresome, and relocate to the center of the microtubule tissue, ultimately reducing abnormal accumulation of α-syn. However, PQ treatment resulted in HDAC6 upregulation, causing abnormal aggregation of α-syn. Taken together, these findings indicated that PQ exposure caused abnormal accumulation of α-syn and decreased effective degradation of α-syn by HDAC6-mediated aggresome-autophagy-lysosome pathway.

Asunto(s)

RESUMEN

We have reported that an environmental pollutant, cadmium, promotes cell death in the human renal tubular cells (RTCs) through hyperactivation of a serine/threonine kinase Akt. However, the molecular mechanisms downstream of Akt in this process have not been elucidated. Cadmium has a potential to accumulate misfolded proteins, and proteotoxicity is involved in cadmium toxicity. To clear the roles of Akt in cadmium exposure-induced RTCs death, we investigated the possibility that Akt could regulate proteotoxicity through autophagy in cadmium chloride (CdCl2)-exposed HK-2 human renal proximal tubular cells. CdCl2 exposure promoted the accumulation of misfolded or damaged proteins, the formation of aggresomes (pericentriolar cytoplasmic inclusions), and aggrephagy (selective autophagy to degrade aggresome). Pharmacological inhibition of Akt using MK2206 or Akti-1/2 enhanced aggrephagy by promoting dephosphorylation and nuclear translocation of transcription factor EB (TFEB)/transcription factor E3 (TFE3), lysosomal transcription factors. TFEB or TFE3 knockdown by siRNAs attenuated the protective effects of MK2206 against cadmium toxicity. These results suggested that aberrant activation of Akt attenuates aggrephagy via TFEB or TFE3 to facilitate CdCl2-induced cell death. Furthermore, these roles of Akt/TFEB/TFE3 were conserved in CdCl2-exposed primary human RTCs. The present study shows the molecular mechanisms underlying Akt activation that promotes cadmium-induced RTCs death.

Asunto(s)

RESUMEN

Protein homeostasis is fundamental to the development of tumors. Ribosome-associated quality-control (RQC) is able to add alanine and threonine to the stagnant polypeptide chain C-terminal (CAT-tail) when protein translation is hindered, while Ankyrin repeat and zinc-finger domain-containing-protein 1 (ANKZF1) can counteract the formation of the CAT-tail, preventing the aggregation of polypeptide chains. In particular, ANKZF1 plays an important role in maintaining mitochondrial protein homeostasis by mitochondrial RQC (mitoRQC) after translation stagnation of precursor proteins targeting mitochondria. However, the role of ANKZF1 in glioblastoma is unclear. Therefore, the current study was aimed to investigate the effects of ANKZF1 in glioblastoma cells and a nude mouse glioblastoma xenograft model. Here, we reported that knockdown of ANKZF1 in glioblastoma cells resulted in the accumulation of CAT-tail in mitochondria, leading to the activated mitochondrial unfolded protein response (UPRmt) and inhibits glioblastoma malignant progression. Excessive CAT-tail sequestered mitochondrial chaperones HSP60, mtHSP70 and proteases LONP1 as well as mitochondrial respiratory chain subunits ND1, Cytb, mtCO2 and ATP6, leading to mitochondrial oxidative phosphorylation dysfunction, membrane potential impairment, and mitochondrial apoptotic pathway activation. Our study highlights ANKZF1 as a valuable target for glioblastoma intervention and provides an innovative insight for the treatment of glioblastoma through the regulating of mitochondrial protein homeostasis.

Asunto(s)

RESUMEN

Essential oils extracted from plant sources along with their biologically active components may have negative effects on insects. Diallyl trisulfide (DAT) is an active component of garlic essential oil, and it exhibits multi-targeted activity against many organisms. Previously we reported that DAT induces male infertility and leads to apyrene and eupyrene sperm dysfunction in Sitotroga cerealella. In this study, we conducted an analysis of testis-specific RNA-Seq data and identified 449 downregulated genes and 60 upregulated genes in the DAT group compared to the control group. The downregulated genes were significantly enriched in the ubiquitin-proteasome pathway. Furthermore, DAT caused a significant reduction in mRNA expression of proteasome regulatory subunit particles required for ATP-dependent degradation of ubiquitinated proteins as well as decreased the expression profile of proteasome core particles, including ß1, ß2, and ß5. Sperm physiological analysis showed that DAT decreased the chymotrypsin-like activity of the 20S proteasome and formed aggresomes in spermatozoa. Overall, our findings suggest that DAT impairs the testis proteasome, ultimately causing male infertility characterized by oligoasthenoteratospermia due to disruption in sperm proteasome assembly in S. cerealella.

Asunto(s)

RESUMEN

BACKGROUND: Ubiquitin-proteasome-system-mediated clearance of misfolded proteins is essential for cells to maintain proteostasis and reduce the proteotoxicity caused by these aberrant proteins. When proteasome activity is inadequate, ubiquitinated proteins are sorted into perinuclear aggresomes, which is a significant defense mechanism employed by cells to combat insufficient proteasome activity, hence mitigating the proteotoxic crisis. It has been demonstrated that phosphorylation of SQSTM1 is crucial in regulating misfolded protein aggregation and autophagic degradation. Although SQSTM1 S403 phosphorylation is essential for the autophagic degradation of ubiquitinated proteins, its significance in proteasome inhibition-induced aggresome formation is yet unknown. Herein, we investigated the influence of SQSTM1 S403 phosphorylation on the aggresome production of ubiquitinated proteins during proteasome suppression. METHODS: We examined the phosphorylation levels of SQSTM1 S403 or T269/S272 in cells after treated with proteasome inhibitors or/and autophagy inhibitors, by western blot and immunofluorescence. We detected the accumulation and aggresome formation of ubiquitinated misfolded proteins in cells treated with proteasome inhibition by western blot and immunofluorescence. Furthermore, we used SQSTM1 phosphorylation-associated kinase inhibitors and mutant constructs to confirm the regulation of different SQSTM1 phosphorylation in aggresome formation. We examined the cell viability using CCK-8 assay. RESULTS: Herein, we ascertained that phosphorylation of SQSTM1 S403 did not enhance the autophagic degradation of ubiquitinated proteins during proteasome inhibition. Proteasome inhibition suppresses the phosphorylation of SQSTM1 S403, which facilitated the aggresome production of polyubiquitinated proteins. Interestingly, we found proteasome inhibition-induced SQSTM1 T269/S272 phosphorylation inhibits the S403 phosphorylation. Suppressing S403 phosphorylation rescues the defective aggresome formation and protects cells from cell death caused by unphosphorylated SQSTM1 (T269/S272). CONCLUSIONS: This study shows that inhibition of SQSTM1 S403 phosphorylation facilitates the aggresome formation of ubiquitinated proteins during proteasome dysfunction. SQSTM1 T269/S272 phosphorylation inhibits the S403 phosphorylation, boosting the aggresome formation of ubiquitinated protein and shielding cells from proteotoxic crisis.

Asunto(s)

RESUMEN

A number of muscular disorders are hallmarked by the aggregation of misfolded proteins within muscle fibers. A specialized form of macroautophagy, termed aggrephagy, is designated to remove and degrade protein aggregates. This review aims to summarize what has been studied so far about the direct involvement of aggrephagy and the activation of the key players, among others, p62, NBR1, Alfy, Tollip, Optineurin, TAX1BP1 and CCT2 in muscular diseases. In the first part of the review, we describe the aggrephagy pathway with the involved proteins; then, we illustrate the muscular disorder histologically characterized by protein aggregates, highlighting the role of aggrephagy pathway abnormalities in these muscular disorders.

Asunto(s)

RESUMEN

Physalis plants are commonly used traditional medicinal herbs, and most of their extracts containing withanolides show anticancer effects. Physapruin A (PHA), a withanolide isolated from P. peruviana, shows antiproliferative effects on breast cancer cells involving oxidative stress, apoptosis, and autophagy. However, the other oxidative stress-associated response, such as endoplasmic reticulum (ER) stress, and its participation in regulating apoptosis in PHA-treated breast cancer cells remain unclear. This study aims to explore the function of oxidative stress and ER stress in modulating the proliferation and apoptosis of breast cancer cells treated with PHA. PHA induced a more significant ER expansion and aggresome formation of breast cancer cells (MCF7 and MDA-MB-231). The mRNA and protein levels of ER stress-responsive genes (IRE1α and BIP) were upregulated by PHA in breast cancer cells. The co-treatment of PHA with the ER stress-inducer (thapsigargin, TG), i.e., TG/PHA, demonstrated synergistic antiproliferation, reactive oxygen species generation, subG1 accumulation, and apoptosis (annexin V and caspases 3/8 activation) as examined by ATP assay, flow cytometry, and western blotting. These ER stress responses, their associated antiproliferation, and apoptosis changes were partly alleviated by the N-acetylcysteine, an oxidative stress inhibitor. Taken together, PHA exhibits ER stress-inducing function to promote antiproliferation and apoptosis of breast cancer cells involving oxidative stress.

Asunto(s)

RESUMEN

BACKGROUND: Aggresomes are collections of intracellular protein aggregates. In liver cells of patients with alcoholic hepatitis, aggresomes appear histologically as cellular inclusions known as Mallory-Denk (M-D) bodies. The proteasome is a multicatalytic intracellular protease that catalyzes the degradation of both normal (native) and abnormal (misfolded and/or damaged) proteins. The enzyme minimizes intracellular protein aggregate formation by rapidly degrading abnormal proteins before they form aggregates. When proteasome activity is blocked, either by specific inhibitors or by intracellular oxidants (e.g., peroxynitrite, acetaldehyde), aggresome formation is enhanced. Here, we sought to verify whether inhibition of proteasome activity by ethanol exposure enhances protein aggregate formation in VL-17A cells, which are recombinant, ethanol-oxidizing HepG2 cells that express both alcohol dehydrogenase (ADH) and cytochrome P450 2E1 (CYP2E1). METHODS: We exposed ethanol-non-oxidizing HepG2 cells (ADH-/CYP2E1-) or ethanol-oxidizing VL-17A (ADH+/CYP2E1+) to varying levels of ethanol for 24 h or 72 h. After these treatments, we stained cells for aggresomes (detected microscopically) and quantified their numbers and sizes. We also conducted flow cytometric analyses to confirm our microscopic findings. Additionally, aggresome content in liver cells of patients with alcohol-induced hepatitis was quantified. RESULTS: After we exposed VL-17A cells to increasing doses of ethanol for 24 h or 72 h, 20S proteasome activity declined in response to rising ethanol concentrations. After 24 h of ethanol exposure, aggresome numbers in VL-17A cells were 1.8-fold higher than their untreated controls at all ethanol concentrations employed. After 72 h of ethanol exposure, mean aggresome numbers were 2.5-fold higher than unexposed control cells. The mean aggregate size in all ethanol-exposed VL-17A cells was significantly higher than in unexposed control cells but was unaffected by the duration of ethanol exposure. Co-exposure of cells to EtOH and rapamycin, the latter an autophagy activator, completely prevented EtOH-induced aggresome formation. In the livers of patients with alcohol-induced hepatitis (AH), the staining intensity of aggresomes was 2.2-fold higher than in the livers of patients without alcohol use disorder (AUD). CONCLUSIONS: We conclude that ethanol-induced proteasome inhibition in ethanol-metabolizing VL-17A hepatoma cells causes accumulation of protein aggregates. Notably, autophagy activation removes such aggregates. The significance of these findings is discussed.

Asunto(s)

RESUMEN

Reactive sulfur species (RSS) have emerged as key regulators of protein quality control. However, the mechanisms by which RSS contribute to cellular processes are not fully understood. In this study, we identified a novel function of RSS in preventing parthanatos, a nonapoptotic form of cell death that is induced by poly (ADP-ribose) polymerase-1 and mediated by the aggresome-like induced structures (ALIS) composed of SQSTM1/p62. We found that sodium tetrasulfide (Na2S4), a donor of RSS, strongly suppressed oxidative stress-dependent ALIS formation and subsequent parthanatos. On the other hand, the inhibitors of the RSS-producing enzymes, such as 3-mercaptopyruvate sulfurtransferase and cystathionine γ-lyase, clearly enhanced ALIS formation and parthanatos. Interestingly, we found that Na2S4 activated heat shock factor 1 by promoting its dissociation from heat shock protein 90, leading to accelerated transcription of HSP70. Considering that the genetic deletion of HSP70 allowed the enhanced ALIS formation, these findings suggest that RSS prevent parthanatos by specifically suppressing ALIS formation through induction of HSP70. Taken together, our results demonstrate a novel mechanism by which RSS prevent cell death, as well as a novel physiological role of RSS in contributing to protein quality control through HSP70 induction, which may lead to better understanding of the bioactivity of RSS.

Asunto(s)

RESUMEN

Histone deacetylase 6 (HDAC6) is a class IIb histone deacetylase that contains two catalytic domains and a zinc-finger ubiquitin binding domain (ZnF-UBP) domain. The deacetylation function of HDAC6 has been extensively studied with common substrates such as α-tubulin, cortactin, and Hsp90. Apart from its deacetylase activity, HDAC6 ZnF-UBP binds to unanchored ubiquitin of specific sequences and serves as a carrier for transporting aggregated proteins. As a result, aggresomes are formed and protein degradation is facilitated by the autophagy-lysosome pathway. This HDAC6-dependent microtubule transport can be used by cells to assemble and activate inflammasomes, which play a critical role in immune regulation. Even viruses can benefit from the carrier of HDAC6 to assist in uncoating their surfaces during their infection cycle. However, HDAC6 is also capable of blocking virus invasion and replication in a non-enzymatic manner. Given these non-enzymatic functions, HDAC6 is closely associated with various diseases, including neurodegeneration, inflammasome-associated diseases, cancer, and viral infections. Small molecule inhibitors targeting the ubiquitin binding pocket of HDAC6 have been investigated. In this review, we focus on mechanisms in non-enzymatic functions of HDAC6 and discuss the rationality and prospects of therapeutic strategies by intervening the activation of HDAC6 ZnF-UBP in concrete diseases.

Asunto(s)

RESUMEN

The spatiotemporal sequestration of misfolded proteins is a mechanism by which cells counterbalance proteome homeostasis upon exposure to various stress stimuli. Chronic inhibition of proteasomes results in a large, juxtanuclear, membrane-less inclusion, known as the aggresome. Although the molecular mechanisms driving its formation, clearance, and pathophysiological implications are continuously being uncovered, the biophysical aspects of aggresomes remain largely uncharacterized. Using fluorescence recovery after photobleaching and liquid droplet disruption assays, we found that the aggresomes are a homogeneously blended condensates with liquid-like properties similar to droplets formed via liquid-liquid phase separation. However, unlike fluidic liquid droplets, aggresomes have more viscosity and hydrogel-like characteristics. We also observed that the inhibition of aggresome formation using microtubule-disrupting agents resulted in less soluble and smaller cytoplasmic speckles, which was associated with marked cytotoxicity. Therefore, the aggresome appears to be cytoprotective and serves as a temporal reservoir for dysfunctional proteasomes and substrates that need to be degraded. Our results suggest that the aggresome assembles through distinct and potentially sequential processes of energy-dependent retrograde transportation and spontaneous condensation into a hydrogel.

Asunto(s)

RESUMEN

Hematopoietic stem cells (HSCs) regenerate blood cells throughout life. To preserve their fitness, HSCs are particularly dependent on maintaining protein homeostasis (proteostasis). However, how HSCs purge misfolded proteins is unknown. Here, we show that in contrast to most cells that primarily utilize the proteasome to degrade misfolded proteins, HSCs preferentially traffic misfolded proteins to aggresomes in a Bag3-dependent manner and depend on aggrephagy, a selective form of autophagy, to maintain proteostasis in vivo. When autophagy is disabled, HSCs compensate by increasing proteasome activity, but proteostasis is ultimately disrupted as protein aggregates accumulate and HSC function is impaired. Bag3-deficiency blunts aggresome formation in HSCs, resulting in protein aggregate accumulation, myeloid-biased differentiation, and diminished self-renewal activity. Furthermore, HSC aging is associated with a severe loss of aggresomes and reduced autophagic flux. Protein degradation pathways are thus specifically configured in young adult HSCs to preserve proteostasis and fitness but become dysregulated during aging.

Asunto(s)

RESUMEN

Manoalide provides preferential antiproliferation of oral cancer but is non-cytotoxic to normal cells by modulating reactive oxygen species (ROS) and apoptosis. Although ROS interplays with endoplasmic reticulum (ER) stress and apoptosis, the influence of ER stress on manoalide-triggered apoptosis has not been reported. The role of ER stress in manoalide-induced preferential antiproliferation and apoptosis was assessed in this study. Manoalide induces a higher ER expansion and aggresome accumulation of oral cancer than normal cells. Generally, manoalide differentially influences higher mRNA and protein expressions of ER-stress-associated genes (PERK, IRE1α, ATF6, and BIP) in oral cancer cells than in normal cells. Subsequently, the contribution of ER stress on manoalide-treated oral cancer cells was further examined. ER stress inducer, thapsigargin, enhances the manoalide-induced antiproliferation, caspase 3/7 activation, and autophagy of oral cancer cells rather than normal cells. Moreover, N-acetylcysteine, an ROS inhibitor, reverses the responses of ER stress, aggresome formation, and the antiproliferation of oral cancer cells. Consequently, the preferential ER stress of manoalide-treated oral cancer cells is crucial for its antiproliferative effect.

Asunto(s)

RESUMEN

Each protein must be synthesized with the correct amino acid sequence, folded into its native structure, and transported to a relevant subcellular location and protein complex. If any of these steps fail, the cell has the capacity to break down aberrant proteins to maintain protein homeostasis (also called proteostasis). All cells possess a set of well-characterized protein quality control systems to minimize protein misfolding and the damage it might cause. Autophagy, a conserved pathway for the degradation of long-lived proteins, aggregates, and damaged organelles, was initially characterized as a bulk degradation pathway. However, it is now clear that autophagy also contributes to intracellular homeostasis by selectively degrading cargo material. One of the pathways involved in the selective removal of damaged and misfolded proteins is chaperone-assisted selective autophagy (CASA). The CASA complex is composed of three main proteins (HSPA, HSPB8 and BAG3), essential to maintain protein homeostasis in muscle and neuronal cells. A failure in the CASA complex, caused by mutations in the respective coding genes, can lead to (cardio)myopathies and neurodegenerative diseases. Here, we summarize our current understanding of the CASA complex and its dynamics. We also briefly discuss how CASA complex proteins are involved in disease and may represent an interesting therapeutic target.Abbreviation ALP: autophagy lysosomal pathway; ALS: amyotrophic lateral sclerosis; AMOTL1: angiomotin like 1; ARP2/3: actin related protein 2/3; BAG: BAG cochaperone; BAG3: BAG cochaperone 3; CASA: chaperone-assisted selective autophagy; CMA: chaperone-mediated autophagy; DNAJ/HSP40: DnaJ heat shock protein family (Hsp40); DRiPs: defective ribosomal products; EIF2A/eIF2α: eukaryotic translation initiation factor 2A; EIF2AK1/HRI: eukaryotic translation initiation factor 2 alpha kinase 1; GABARAP: GABA type A receptor-associated protein; HDAC6: histone deacetylase 6; HSP: heat shock protein; HSPA/HSP70: heat shock protein family A (Hsp70); HSP90: heat shock protein 90; HSPB8: heat shock protein family B (small) member 8; IPV: isoleucine-proline-valine; ISR: integrated stress response; KEAP1: kelch like ECH associated protein 1; LAMP2A: lysosomal associated membrane protein 2A; LATS1: large tumor suppressor kinase 1; LIR: LC3-interacting region; MAP1LC3/LC3: microtubule associated protein 1 light chain 3; MTOC: microtubule organizing center; MTOR: mechanistic target of rapamycin kinase; NFKB/NF-κB: nuclear factor kappa B; NFE2L2: NFE2 like bZIP transcription factor 2; PLCG/PLCγ: phospholipase C gamma; polyQ: polyglutamine; PQC: protein quality control; PxxP: proline-rich; RAN translation: repeat-associated non-AUG translation; SG: stress granule; SOD1: superoxide dismutase 1; SQSTM1/p62: sequestosome 1; STUB1/CHIP: STIP1 homology and U-box containing protein 1; STK: serine/threonine kinase; SYNPO: synaptopodin; TBP: TATA-box binding protein; TARDBP/TDP-43: TAR DNA binding protein; TFEB: transcription factor EB; TPR: tetratricopeptide repeats; TSC1: TSC complex subunit 1; UBA: ubiquitin associated; UPS: ubiquitin-proteasome system; WW: tryptophan-tryptophan; WWTR1: WW domain containing transcription regulator 1; YAP1: Yes1 associated transcriptional regulator.

Asunto(s)

RESUMEN

In the adenovirus-infected cells, virus mRNAs are selectively exported to the cytoplasm by virus early gene products to facilitate virus replication. We previously showed AU-rich elements (AREs) containing mRNAs are exported to the cytoplasm and stabilized in infected cells. Here, we analyzed ribonucleoprotein (RNP) granules in the cytoplasm that are involved in mRNA degradation to elucidate the mechanism of ARE-mRNA stabilization in adenovirus infected cells. Our findings showed that processing bodies (PBs) aggregate, then almost all PBs are translocated to aggresomes formed by adenoviral gene products during the late phase of infection. Furthermore, E4orf3 was required for the PBs translocation, and the same domains of E4orf3-mutants required to change the form of promyelocytic leukemia bodies were also needed for PBs translocation. Luciferase activity showed that these domains were critical for miRNA- and ARE-mediated mRNA decay. These findings suggest that adenovirus changes the behavior of PBs to prevent ARE-mRNA downregulation.

Asunto(s)

RESUMEN

Helicobacter pylori-mediated gastric carcinogenesis involves upregulation of the E3 ubiquitin ligase Siah2 and its phosphorylation-mediated stabilization. This study elucidates a novel mechanism of oxidative stress regulation by phosphorylated Siah2 in H. pylori-infected gastric epithelial cancer cells (GECs). We identify that H. pylori-mediated Siah2 phosphorylation at the 6th serine residue (P-S6-Siah2) enhances proteasomal degradation of the 78-kDa glucose-regulated protein (GRP78) possessing antioxidant functions. S6 phosphorylation stabilizes Siah2 and P-S6-Siah2 potentiates H. pylori-mediated reactive oxygen species (ROS) generation. However, infected S6A phospho-null Siah2-expressing cells have decreased cellular GRP78 level as surprisingly these cells release GRP78 to a higher extent and accumulate significantly higher ROS than the wild type (WT) Siah2 construct-expressing cells. Ectopic expression of GRP78 prevents the loss of mitochondrial membrane potential and cellular ROS accumulation caused by H. pylori. H. pylori-induced mitochondrial damage and mitochondrial membrane potential loss are potentiated in Siah2-overexpressing cells but these effects are further enhanced in S6A-expressing cells. This study also confirms that while phosphorylation-mediated Siah2 stabilization optimally upregulates aggresome accumulation, it suppresses autophagosome formation, thus decreasing the dependency on the latter mechanism in regulating cellular protein abundance. Disruption of the phospho-Siah2-mediated aggresome formation impairs proliferation of infected GECs. Thus, Siah2 phosphorylation has diagnostic and therapeutic significance in H. pylori-mediated gastric cancer (GC).

Asunto(s)

RESUMEN

Alzheimer's disease (AD) is one of the most prevailing neurodegenerative diseases in the world, which is characterized by memory dysfunction and the formation of tau and amyloid ß (Aß) aggregates in multiple brain regions, including the hippocampus and cortex. The formation of senile plaques involving tau hyperphosphorylation, fibrillar Aß, and neurofibrillary tangles (NFTs) is used as a pathological marker of AD and eventually produces aggregation or misfolded protein. Importantly, it has been found that the failure to degrade these aggregate-prone proteins leads to pathological consequences, such as synaptic impairment, cytotoxicity, neuronal atrophy, and memory deficits associated with AD. Recently, increasing evidence has suggested that the autophagy pathway plays a role as a central cellular protection system to prevent the toxicity induced by aggregation or misfolded proteins. Moreover, it has also been revealed that AD-related protein aggresomes could be selectively degraded by autophagosome and lysosomal fusion through the autophagy pathway, which is known as aggrephagy. Therefore, the regulation of autophagy serve as a useful approach to modulate the formation of aggresomes associated with AD. This review focuses on the recent improvements in the application of natural compounds and small molecules as a potential therapeutic approach for AD prevention and treatment via aggrephagy.

RESUMEN

The deacetylase HDAC6 has tandem catalytic domains and a zinc finger domain (ZnF) binding ubiquitin (Ub). While the catalytic domain has an antiviral effect, the ZnF facilitates influenza A virus (IAV) infection and cellular stress responses. By recruiting Ub via the ZnF, HDAC6 promotes the formation of aggresomes and stress granules (SGs), dynamic structures associated with pathologies such as neurodegeneration. IAV subverts the aggresome/HDAC6 pathway to facilitate capsid uncoating during early infection. To target this pathway, we generate designed ankyrin repeat proteins (DARPins) binding the ZnF; one of these prevents interaction with Ub in vitro and in cells. Crystallographic analysis shows that it blocks the ZnF pocket where Ub engages. Conditional expression of this DARPin reversibly impairs infection by IAV and Zika virus; moreover, SGs and aggresomes are downregulated. These results validate the HDAC6 ZnF as an attractive target for drug discovery.

Asunto(s)

RESUMEN

As central effectors of ubiquitin (Ub)-mediated proteolysis, proteasomes are regulated at multiple levels, including degradation of unwanted or dysfunctional particles via autophagy (termed proteaphagy). In yeast, inactive proteasomes are exported from the nucleus, sequestered into cytoplasmic aggresomes via the Hsp42 chaperone, extensively ubiquitylated, and then tethered to the expanding phagophore by the autophagy receptor Cue5. Here, we demonstrate the need for ubiquitylation driven by the trio of Ub ligases (E3s), San1, Rsp5, and Hul5, which together with their corresponding E2s work sequentially to promote nuclear export and Cue5 recognition. Whereas San1 functions prior to nuclear export, Rsp5 and Hul5 likely decorate aggresome-localized proteasomes in concert. Ultimately, topologically complex Ub chain(s) containing both K48 and K63 Ub-Ub linkages are assembled, mainly on the regulatory particle, to generate autophagy-competent substrates. Because San1, Rsp5, Hul5, Hsp42, and Cue5 also participate in general proteostasis, proteaphagy likely engages a fundamental mechanism for eliminating inactive/misfolded proteins.

Asunto(s)

RESUMEN

TAR DNA-binding protein 43 (TDP-43) is a causative factor of amyotrophic lateral sclerosis (ALS). Cytoplasmic TDP-43 aggregates in neurons are a hallmark pathology of ALS. Under various stress conditions, TDP-43 localizes sequentially to two cytoplasmic protein aggregates, namely, stress granules (SGs) first and then aggresomes. Accumulating evidence suggests that delayed clearance of TDP-43-positive SGs is associated with pathological TDP-43 aggregates in ALS. We found that ubiquitin-specific protease 10 (USP10) promotes the clearance of TDP-43-positive SGs in cells treated with proteasome inhibitor, thereby promoting the formation of TDP-43-positive aggresomes, and the depletion of USP10 increases the amount of insoluble TDP-35, a cleaved product of TDP-43, in the cytoplasm. TDP-35 interacted with USP10 in an RNA-binding-dependent manner; however, impaired RNA binding of TDP-35 reduced the localization in SGs and aggresomes and induced USP10-negative TDP-35 aggregates. Immunohistochemistry showed that most of the cytoplasmic TDP-43/TDP-35 aggregates in the neurons of ALS patients were USP10 negative. Our findings suggest that USP10 inhibits aberrant aggregation of TDP-43/TDP-35 in the cytoplasm of neuronal cells by promoting the clearance of TDP-43/TDP-35-positive SGs and facilitating the formation of TDP-43/TDP-35-positive aggresomes.

Asunto(s)