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Copyright of Journal of Hainan Medical University is the property of Journal of Hainan Medical College Editorial Office and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

Background: The ubiquitin proteasome has a major role in the development of many tumors. However, the prognostic importance of ubiquitin proteasome-system genes (UPSGs) in hepatocellular carcinoma (HCC) is not fully defined. Methods: The TCGA and ICGC datasets were utilized to obtain transcriptional profiling data as well as clinicopathological information about HCC. The 3-UPSGs signature for the TCGA cohort was developed via univariate and LASSO Cox regression analyses. Differential expression of genes was demonstrated by qRT-PCR and immunohistochemistry (IHC). Biological pathways were studied using GSVA and GSEA. Six algorithms were used to compare immune infiltration between the two risk groups. Furthermore, drug sensitivity was measured using the "pRRophetic" R package. The predictive capacity of the 3-UPSGs signature for sensitivity to immunotherapy was also explored. Moreover, we performed a pan-cancer analysis of the 3-UPSGs signature. Results: A risk model containing 3 UPSGs (DCAF13, CDC20 and PSMB5) was developed. IHC and qRT-PCR results showed that signature genes were significantly overexpressed in HCC tissues. The high-risk group had a worse prognosis, with a higher clinicopathological grade, higher levels of tumor mutation burden (TMB), elevated levels of immune checkpoint (IC) expression, as well as increased sensitivity to immunotherapy. The two risk groups also differ in their sensitivity to chemotherapeutic drugs. Furthermore, the three UPSGs may play crucial roles in the progression of multiple types of cancers. Conclusion: We created a 3-UPSGs signature to estimate the prognosis of HCC and to assist in individualized treatment. [ABSTRACT FROM AUTHOR]

Abstract Background The poor survival of pancreatic cancer patients is largely attributable to cachexia, a syndrome of severe weight and muscle loss. To investigate the aetiology of cancer cachexia, preclinical models that closely recapitulate the human disease process are essential. Patient derived tumour organoids are promising novel cancer models, but their ability to induce cachexia in mice has not been investigated. We developed two pancreatic tumour organoid‐based mouse models and demonstrate their potential for cancer cachexia research. Methods Two patients with pancreatic cancer, from whom tumour organoid cultures were previously established, were selected based on their cachexia phenotype. Patient 09 was characterized as cachectic according to the international consensus definition of cancer cachexia, whereas patient 12 was classified as non‐cachectic. Organoid cultures PANCO‐09b and PANCO‐12a in basement membrane extract (BME) were injected subcutaneously into the flanks of 9‐weeks old NMRI‐Foxn1nu mice (n = 8/group). A control group was injected with BME only (n = 4). Body weight was monitored every 2–3 days for 38 days. Hind limb muscle wet and dry weights were measured. Adipocyte size in inguinal white adipose tissue was measured using haematoxylin and eosin‐stained sections. Expression of genes associated with cancer cachexia in muscle and liver tissue was analysed using qPCR. Results Engraftment of tumour organoids was successful in 87.5% of PANCO‐09b implanted mice and in 50% of PANCO‐12a mice, with similar average tumour weights at endpoint (34.4 ± 25.1 mg vs. 32.8 ± 40.2 mg, respectively, P = 0.450). All groups initially gained weight, but PANCO‐12a implanted mice progressively lost an average body weight of 1.7 ± 0.8 g from day 28 onwards. PANCO‐12a‐implanted mice gained significantly less weight from baseline than controls (0.7 ± 0.6 g, P = 0.027). Overall body weight gain of PANCO‐09b mice was also lower but not significantly different from controls (2.0 ± 1.2 g vs. 2.9 ± 1.6 g, P = 0.961). Wet weights of hind leg muscles were negatively correlated with tumour weight but did not differ between groups. Adipocytes of PANCO‐12a implanted mice were smaller compared to SHAM as well as PANCO09b mice (P

Anabolic resistance to feeding in aged muscle is well-characterized; however, whether old skeletal muscle is intrinsically resistant to acute mechanical loading is less clear. The aim of this study was to determine the impact of aging on muscle protein synthesis (MPS), ribosome biogenesis, and protein breakdown in skeletal muscle following a single bout of resistance exercise. Adult male F344/BN rats aged 10 (Adult) and 30 (Old) months underwent unilateral maximal eccentric contractions of the hindlimb. Precursor rRNA increased early post-exercise (6-18 h), preceding elevations in ribosomal mass at 48 h in Adult and Old; there were no age-related differences in these responses. MPS increased early post-exercise in both Adult and Old; however, at 48 h of recovery, MPS returned to baseline in Old but not Adult. This abbreviated protein synthesis response in Old was associated with decreased levels of IRS1 protein and increased BiP, CHOP and eIF2α levels. Other than these responses, anabolic signaling was similar in Adult and Old muscle in the acute recovery phase. Basal proteasome activity was lower in Old, and resistance exercise did not increase the activity of either the ATP-dependent or independent proteasome, or autophagy (Cathepsin L activity) in either Adult or Old muscle. We conclude that MPS and ribosome biogenesis in response to maximal resistance exercise in old skeletal muscle are initially intact; however, the MPS response is abbreviated in Old, which may be the result of ER stress and/or blunted exercise-induced potentiation of the MPS response to feeding.

Sarcopenia is a debilitating skeletal muscle disease that accelerates in the last decades of life and is characterized by marked deficits in muscle strength, mass, quality, and metabolic health. The multifactorial causes of sarcopenia have proven difficult to treat and involve a complex interplay between environmental factors and intrinsic age-associated changes. It is generally accepted that sarcopenia results in a progressive loss of skeletal muscle function that exceeds the loss of mass, indicating that while loss of muscle mass is important, loss of muscle quality is the primary defect with advanced age. Furthermore, preclinical models have suggested that aged skeletal muscle exhibits defects in cellular quality control such as the degradation of damaged mitochondria. Recent evidence suggests that a dysregulation of proteostasis, an important regulator of cellular quality control, is a significant contributor to the aging-associated declines in muscle quality, function, and mass. Although skeletal muscle mammalian target of rapamycin complex 1 (mTORC1) plays a critical role in cellular control, including skeletal muscle hypertrophy, paradoxically, sustained activation of mTORC1 recapitulates several characteristics of sarcopenia. Pharmaceutical inhibition of mTORC1 as well as caloric restriction significantly improves muscle quality in aged animals, however, the mechanisms controlling cellular proteostasis are not fully known. This information is important for developing effective therapeutic strategies that mitigate or prevent sarcopenia and associated disability. This review identifies recent and historical understanding of the molecular mechanisms of proteostasis driving age-associated muscle loss and suggests potential therapeutic interventions to slow or prevent sarcopenia. [ABSTRACT FROM AUTHOR]

Anabolic resistance to feeding in aged muscle is well-characterized; however, whether old skeletal muscle is intrinsically resistant to acute mechanical loading is less clear. The aim of this study was to determine the impact of aging on muscle protein synthesis (MPS), ribosome biogenesis, and protein breakdown in skeletal muscle following a single bout of resistance exercise. Adult male F344/BN rats aged 10 (Adult) and 30 (Old) months underwent unilateral maximal eccentric contractions of the hindlimb. Precursor rRNA increased early post-exercise (6-18 h), preceding elevations in ribosomal mass at 48 h in Adult and Old; there were no age-related differences in these responses. MPS increased early post-exercise in both Adult and Old; however, at 48 h of recovery, MPS returned to baseline in Old but not Adult. This abbreviated protein synthesis response in Old was associated with decreased levels of IRS1 protein and increased BiP, CHOP and eIF2α levels. Other than these responses, anabolic signaling was similar in Adult and Old muscle in the acute recovery phase. Basal proteasome activity was lower in Old, and resistance exercise did not increase the activity of either the ATP-dependent or independent proteasome, or autophagy (Cathepsin L activity) in either Adult or Old muscle. We conclude that MPS and ribosome biogenesis in response to maximal resistance exercise in old skeletal muscle are initially intact; however, the MPS response is abbreviated in Old, which may be the result of ER stress and/or blunted exercise-induced potentiation of the MPS response to feeding.

Objective: Skeletal muscle is a pivotal organ for the coordination of systemic metabolism, constituting one of the largest storage site for glucose, lipids and amino acids. Tight temporal orchestration of protein breakdown in times of fasting has to be balanced with preservation of muscle mass and function. However, the molecular mechanisms that control the fasting response in muscle are poorly understood. Methods: We now have identified a role for the peroxisome proliferator-activated receptor γ coactivator 1β (PGC-1β) in the regulation of catabolic pathways in this context in muscle-specific loss-of-function mouse models. Results: Muscle-specific knockouts for PGC-1β experience mitigated muscle atrophy in fasting, linked to reduced expression of myostatin, atrogenes, activation of AMP-dependent protein kinase (AMPK) and other energy deprivation signaling pathways. At least in part, the muscle fasting response is modulated by a negative effect of PGC-1β on the nuclear factor of activated T-cells 1 (NFATC1). Conclusions: Collectively, these data highlight the complex regulation of muscle metabolism and reveal a new role for muscle PGC-1β in the control of proteostasis in fasting.

Endothelial cAMP-specific phosphodiesterase PDE3A is one of the major negative regulators of the endothelial barrier function in acute lung injury models. However, the mechanisms underlying its regulation still need to be fully resolved. We show here that the PDE3A is a newly described client of the molecular chaperone heat shock protein 90 (hsp90). In endothelial cells (ECs), hsp90 inhibition by geldanamycin (GA) led to a disruption of the hsp90/PDE3A complex, followed by a significant decrease in PDE3A protein levels. The decrease in PDE3A protein levels was ubiquitin-proteasome-dependent and required the activity of the E3 ubiquitin ligase C terminus of Hsc70-interacting protein. GA treatment also enhanced the association of PDE3A with hsp70, which partially prevented PDE3A degradation. GA-induced decreases in PDE3A protein levels correlated with decreased PDE3 activity and increased cAMP levels in EC. We also demonstrated that protein kinase G-dependent phosphorylation of PDE3A at Ser 654 can signal the dissociation of PDE3A from hsp90 and PDE3A degradation. This was confirmed by endogenous PDE3A phosphorylation and degradation in 8-Br-cGMP- or 8-CPT-cGMP- and Bay 41-8543-stimulated EC and comparisons of WT- and phospho-mimic S 654 D mutant PDE3A protein stability in transiently transfected HEK293 cells. In conclusion, we have identified a new mechanism of PDE3A regulation mediated by the ubiquitin-proteasome system. Further, the degradation of PDE3A is controlled by the phosphorylation of S 654 and the interaction with hsp90. We speculate that targeting the PDE3A/hsp90 complex could be a therapeutic approach for acute lung injury., Competing Interests: Conflict of interest The authors declare they have no conflicts of interest with the contents of this article., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

Abstract Background Ebstein's anomaly (EA) is a rare congenital heart disease of the tricuspid valve and right ventricle. Patients with EA often manifest with left ventricular noncompaction (LVNC), a cardiomyopathy. Despite implication of cardiac sarcomere genes in some cases, very little is understood regarding the genetic etiology of EA/LVNC. Our study describes a multigenerational family with at least 10 of 17 members affected by EA/LVNC. Methods We performed echocardiography on all family members and conducted exome sequencing of six individuals. After identifying candidate variants using two different bioinformatic strategies, we confirmed segregation with phenotype using Sanger sequencing. We investigated structural implications of candidate variants using protein prediction models. Results Exome sequencing analysis of four affected and two unaffected members identified a novel, rare, and damaging coding variant in the Kelch‐like family member 26 (KLHL26) gene located on chromosome 19 at position 237 of the protein (GRCh37). This variant region was confirmed by Sanger sequencing in the remaining family members. KLHL26 (c.709C > T p.R237C) segregates only with EA/LVNC‐affected individuals (FBAT p

Abstract Background This study aimed to screen sensitive biomarkers for the efficacy evaluation of neoadjuvant chemotherapy in breast cancer. Methods In this study, Illumina digital gene expression sequencing technology was applied and differentially expressed genes (DEGs) between patients presenting pathological complete response (pCR) and non-pathological complete response (NpCR) were identified. Further, gene ontology and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis were then performed. The genes in significant enriched pathways were finally quantified by quantitative real-time PCR (qRT-PCR) to confirm that they were differentially expressed. Additionally, GSE23988 from Gene Expression Omnibus database was used as the validation dataset to confirm the DEGs. Results After removing the low-quality reads, 715 DEGs were finally detected. After mapping to KEGG pathways, 10 DEGs belonging to the ubiquitin proteasome pathway (HECTD3, PSMB10, UBD, UBE2C, and UBE2S) and cytokine–cytokine receptor interactions (CCL2, CCR1, CXCL10, CXCL11, and IL2RG) were selected for further analysis. These 10 genes were finally quantified by qRT-PCR to confirm that they were differentially expressed (the log2 fold changes of selected genes were − 5.34, 7.81, 6.88, 5.74, 3.11, 19.58, 8.73, 8.88, 7.42, and 34.61 for HECTD3, PSMB10, UBD, UBE2C, UBE2S, CCL2, CCR1, CXCL10, CXCL11, and IL2RG, respectively). Moreover, 53 common genes were confirmed by the validation dataset, including downregulated UBE2C and UBE2S. Conclusion Our results suggested that these 10 genes belonging to these two pathways might be useful as sensitive biomarkers for the efficacy evaluation of neoadjuvant chemotherapy in breast cancer.

Despite the saturating concentrations of intracellular l -arginine, nitric oxide (NO) production in endothelial cells (EC) can be stimulated by exogenous arginine. This phenomenon, termed the "arginine paradox" led to the discovery of an arginine recycling pathway in which l -citrulline is recycled to l -arginine by utilizing two important urea cycle enzymes argininosuccinate synthetase (ASS) and argininosuccinate lyase (ASL). Prior work has shown that ASL is present in a NO synthetic complex containing hsp90 and endothelial NO synthase (eNOS). However, it is unclear whether hsp90 forms functional complexes with ASS and ASL and if it is involved regulating their activity. Thus, elucidating the role of hsp90 in the arginine recycling pathway was the goal of this study. Our data indicate that both ASS and ASL are chaperoned by hsp90. Inhibiting hsp90 activity with geldanamycin (GA), decreased the activity of both ASS and ASL and decreased cellular l -arginine levels in bovine aortic endothelial cells (BAEC). hsp90 inhibition led to a time-dependent decrease in ASS and ASL protein, despite no changes in mRNA levels. We further linked this protein loss to a proteasome dependent degradation of ASS and ASL via the E3 ubiquitin ligase, C-terminus of Hsc70-interacting protein (CHIP) and the heat shock protein, hsp70. Transient over-expression of CHIP was sufficient to stimulate ASS and ASL degradation while the over-expression of CHIP mutant proteins identified both TPR- and U-box-domain as essential for ASS and ASL degradation. This study provides a novel insight into the molecular regulation l -arginine recycling in EC and implicates the proteasome pathway as a possible therapeutic target to stimulate NO signaling. Image 1 • Arginosuccinate synthase (ASS) and arginosuccinate lyase (ASL) are chaperoned by Hsp90. • Hsp90 inhibition leads to decrease the activity of both ASL and ASS, and decrease of cellular l -arginine level. • ASS and ASL are degraded through a CHIP-mediated proteasome pathway. [ABSTRACT FROM AUTHOR]

The maintenance of skeletal muscle mass depends on the overall balance between the rates of protein synthesis and degradation. Thus, age-related muscle atrophy and function, commonly known as sarcopenia, may result from decreased protein synthesis, increased proteolysis, or simultaneous changes in both processes governed by complex multifactorial mechanisms. Growing evidence implicates oxidative stress and reactive oxygen species (ROS) as an essential regulator of proteolysis. Our previous studies have shown that genetic deletion of CuZn superoxide dismutase (CuZnSOD, Sod1) in mice leads to elevated oxidative stress, muscle atrophy and weakness, and an acceleration in age-related phenotypes associated with sarcopenia. The goal of this study is to determine whether oxidative stress directly influences the acceleration of proteolysis in skeletal muscle of Sod1−/− mice as a function of age. Compared to control, Sod1−/− muscle showed a significant elevation in protein carbonyls and 3-nitrotyrosine levels, suggesting high oxidative and nitrosative protein modifications were present. In addition, age-dependent muscle atrophy in Sod1−/− muscle was accompanied by an upregulation of the cysteine proteases, calpain, and caspase-3, which are known to play a key role in the initial breakdown of sarcomeres during atrophic conditions. Furthermore, an increase in oxidative stress-induced muscle atrophy was also strongly coupled with simultaneous activation of two major proteolytic systems, the ubiquitin-proteasome and lysosomal autophagy pathways. Collectively, our data suggest that chronic oxidative stress in Sod1−/− mice accelerates age-dependent muscle atrophy by enhancing coordinated activation of the proteolytic systems, thereby resulting in overall protein degradation. [ABSTRACT FROM AUTHOR]

Copyright of Chinese Journal of Preventive Veterinary Medicine / Zhongguo Yufang Shouyi Xuebao is the property of Chinese Journal of Preventive Veterinary Medicine Editorial Office and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

• Diacetyl vapor exposure causes airway basal cell injury with keratin 5 ubiquitination and decreased ΔNp63 expression. • Airway epithelial repair after diacetyl vapor exposure occurs through ubiquitination and subsequent proteasome degradation of keratin 5 protein damage. • Inhibition of the ubiquitin-proteasome pathway after diacetyl vapor exposure results in impaired clearance of keratin 5 ubiquitinated proteins. Diacetyl (DA; 2,3-butanedione) is a chemical found commonly in foods and e-cigarettes. When inhaled, DA causes epithelial injury, though the mechanism of repair remain poorly understood. The objective of this study was to evaluate airway basal cell repair after DA vapor exposure. Primary human bronchial epithelial cells were exposed to DA or PBS for 1 h. Lactate dehydrogenase, cleaved caspase 3/7 and trans-epithelial electrical resistance were measured prior to and following exposure. Exposed cultures were analyzed for the airway basal cell markers keratin 5 and p63 as well as ubiquitin and proteasome activity. Cultures were also treated with a proteasome inhibitor (MG132). DA vapor exposure caused a transient decrease in trans-epithelial electrical resistance in all DA-exposed cultures. Supernatant lactate dehydrogenase and cleaved caspase 3/7 increased significantly at the highest DA concentration but not at lower DA concentrations. Increased keratin 5 ubiquitination occurred after DA exposure but resolved by day 3. Damage to airway basal cells persisted at day 3 in the presence of MG132. Diacetyl exposure results in airway basal cell injury with keratin 5 ubiquitination and decreased p63 expression. The ubiquitin-proteasome-pathway partially mediates airway basal cell repair after acute DA exposure. [ABSTRACT FROM AUTHOR]

Background: Ebstein's anomaly (EA) is a rare congenital heart disease of the tricuspid valve and right ventricle. Patients with EA often manifest with left ventricular noncompaction (LVNC), a cardiomyopathy. Despite implication of cardiac sarcomere genes in some cases, very little is understood regarding the genetic etiology of EA/LVNC. Our study describes a multigenerational family with at least 10 of 17 members affected by EA/LVNC. Methods: We performed echocardiography on all family members and conducted exome sequencing of six individuals. After identifying candidate variants using two different bioinformatic strategies, we confirmed segregation with phenotype using Sanger sequencing. We investigated structural implications of candidate variants using protein prediction models. Results: Exome sequencing analysis of four affected and two unaffected members identified a novel, rare, and damaging coding variant in the Kelch‐like family member 26 (KLHL26) gene located on chromosome 19 at position 237 of the protein (GRCh37). This variant region was confirmed by Sanger sequencing in the remaining family members. KLHL26 (c.709C > T p.R237C) segregates only with EA/LVNC‐affected individuals (FBAT p <.05). Investigating structural implications of the candidate variant using protein prediction models suggested that the KLHL26 variant disrupts electrostatic interactions when binding to part of the ubiquitin proteasome, specifically Cullin3 (CUL3), a component of E3 ubiquitin ligase. Conclusion: In this familial case of EA/LVNC, we have identified a candidate gene variant, KLHL26 (p.R237C), which may have an important role in ubiquitin‐mediated protein degradation during cardiac development. [ABSTRACT FROM AUTHOR]

Flaviviruses target their replication on membranous structures derived from the ER, where both viral and host proteins play crucial structural and functional roles. Here, we have characterized the involvement of the ER-associated degradation (ERAD) pathway core E3 ligase complex (SEL1L-HRD1) regulator proteins in the replication of Japanese encephalitis virus (JEV). Through high-resolution immunofluorescence imaging of JEV-infected HeLa cells, we observe that the virus replication complexes marked by NS1 strongly colocalize with the ERAD adapter SEL1L, lectin OS9, ER-membrane shuttle factor HERPUD1, E3 ubiquitin ligase HRD1 and rhomboid superfamily member DERLIN1. NS5 positive structures also show strong overlap with SEL1L. While these effectors show significant transcriptional upregulation, their protein levels remain largely stable in infected cells. siRNA mediated depletion of OS9, SEL1L, HERPUD1 and HRD1 significantly inhibit viral RNA replication and titres, with SEL1L depletion showing the maximum attenuation of replication. By performing protein translation arrest experiments, we show that SEL1L, and OS9 are stabilised upon JEV infection. Overall results from this study suggest that these ERAD effector proteins are crucial host-factors for JEV replication.

Anabolic resistance to feeding in aged muscle is well-characterized; however, whether old skeletal muscle is intrinsically resistant to acute mechanical loading is less clear. The aim of this study was to determine the impact of aging on muscle protein synthesis (MPS), ribosome biogenesis, and protein breakdown in skeletal muscle following a single bout of resistance exercise. Adult male F344/BN rats aged 10 (Adult) and 30 (Old) months underwent unilateral maximal eccentric contractions of the hindlimb. Precursor rRNA increased early post-exercise (6–18 h), preceding elevations in ribosomal mass at 48 h in Adult and Old; there were no age-related differences in these responses. MPS increased early post-exercise in both Adult and Old; however, at 48 h of recovery, MPS returned to baseline in Old but not Adult. This abbreviated protein synthesis response in Old was associated with decreased levels of IRS1 protein and increased BiP, CHOP and eIF2α levels. Other than these responses, anabolic signaling was similar in Adult and Old muscle in the acute recovery phase. Basal proteasome activity was lower in Old, and resistance exercise did not increase the activity of either the ATP-dependent or independent proteasome, or autophagy (Cathepsin L activity) in either Adult or Old muscle. We conclude that MPS and ribosome biogenesis in response to maximal resistance exercise in old skeletal muscle are initially intact; however, the MPS response is abbreviated in Old, which may be the result of ER stress and/or blunted exercise-induced potentiation of the MPS response to feeding.

Graphical abstract Highlights • The pathways to the development of Alzheimer's and Parkinson's disease share similarities, though the specific components or proteins involved may differ. • This review highlights, insights regarding ubiquitin-proteasome system, aberrant alternative splicing of exons, dysfunctional mitochondria role in neurodegeneration and improves the understanding of the common mechanism. • This discussion is particularly relevant, given the strong relationship between major pathways and neurodegeneration, and the pressing need to improve the mechanisms contributing to biomarker screening. Abstract Neurodegenerative diseases are usually sporadic in nature and commonly influenced by a wide range of genetic, life style and environmental factors. A unifying feature of Alzheimer's disease (AD) and Parkinson's disease (PD) is the abnormal accumulation and processing of mutant or damaged intra and extracellular proteins; this leads to neuronal vulnerability and dysfunction in the brain. Through a detailed review of ubiquitin proteasome, mRNA splicing, mitochondrial dysfunction, and oxidative stress pathway interrelation on neurodegeneration can improve the understanding of the disease mechanism. The identified pathways common to AD and PD nominate promising new targets for further studies, and as well as biomarkers. These insights suggested would likely provide major stimuli for developing unified treatment approaches to combat neurodegeneration. More broadly, pathways can serve as vehicles for integrating findings from diverse studies of neurodegeneration. The evidence examined in this review provides a brief overview of the current literature on significant pathways in promoting in AD, PD. Additionally, these insights suggest that biomarkers and treatment strategies may require simultaneous targeting of multiple components. [ABSTRACT FROM AUTHOR]

Vascular inflammation in atheroprone vessels propagates throughout the arterial tree in dyslipidemic patients, thereby accelerating atherosclerotic progression. To elucidate the mechanism of vascular inflammation, most previous studies have focused on inflammation-related signals that are sent in response to vasoactive stimuli. However, it is also important to understand how normal blood vessels become defective and start degenerating. Growing evidence suggests that major protein catabolism pathways, including the ubiquitin-proteasome, autophagy, and calpain systems, are disturbed in atheroprone vessels and contribute to the pathogenesis of atherosclerosis. Indeed, dysregulation of ubiquitin–proteasome pathways results in the accumulation of defective proteins in blood vessels, leading to vascular endothelial dysfunction and apoptosis in affected cells. Impaired autophagy-lysosomal degradation affects smooth muscle cell transformation and proliferation, as well as endothelial integrity and phagocytic clearance of cellular corpses. Dysregulation of the calpain system confers proatherogenic properties to endothelial cells, smooth muscle cells, and macrophages. In this review article, we will discuss the current information available on defective protein catabolism in atheroprone vessels and its potential interrelation with inflammation-related signals.

The mechanisms responsible for maintaining macromolecular protein complexes, with their proper localization and subunit stoichiometry, are incompletely understood. Here we studied the maintenance of the sarcomere, the basic contractile macromolecular complex of cardiomyocytes. We performed single-cell analysis of cardiomyocytes using imaging of mRNA and protein synthesis, and demonstrate that three distinct mechanisms are responsible for the maintenance of the sarcomere: mRNAs encoding for sarcomeric proteins are localized to the sarcomere, ribosomes are localized to the sarcomere with localized sarcomeric protein translation, and finally, a localized E3 ubiquitin ligase allow efficient degradation of excess unincorporated sarcomeric proteins. We show that these mechanisms are distinct, required, and work in unison, to ensure both spatial localization, and to overcome the large variability in transcription. Cardiomyocytes simultaneously maintain all their sarcomeres using localized translation and degradation processes where proteins are continuously and locally synthesized at high rates, and excess proteins are continuously degraded. [ABSTRACT FROM AUTHOR]

In this review, we describe research findings on the effects of alcohol exposure on two major catabolic systems in liver cells: the ubiquitin–proteasome system (UPS) and autophagy. These hydrolytic systems are not unique to liver cells; they exist in all eukaryotic tissues and cells. However, because the liver is the principal site of ethanol metabolism, it sustains the greatest damage from heavy drinking. Thus, the focus of this review is to specifically describe how ethanol oxidation modulates the activities of the UPS and autophagy and the mechanisms by which these changes contribute to the pathogenesis of alcohol-induced liver injury. Here, we describe the history and the importance of cellular hydrolytic systems, followed by a description of each catabolic pathway and the differential modulation of each by ethanol exposure. Overall, the evidence for an involvement of these catabolic systems in the pathogenesis of alcoholic liver disease is quite strong. It underscores their importance, not only as effective means of cellular recycling and eventual energy generation, but also as essential components of cellular defense.

We present the hypothesis that an accumulation of dysfunctional mitochondria initiates a signaling cascade leading to motor neuron and muscle fiber death and culminating in sarcopenia. Interactions between neural and muscle cells that contain dysfunctional mitochondria exacerbate sarcopenia. Preventing sarcopenia will require identifying mitochondrial sources of dysfunction that are reversible. [ABSTRACT FROM AUTHOR]

Epithelial and neuronal cells are highly asymmetric, with discrete regions responsible for different roles that underlie the generation of specific compartments within cells that are distinct in biochemical composition, structure, and morphology that ultimately lead to distinct functions. Controlled and specific molecular targeting and sorting have been studied to understand the generation of asymmetric domains inside cells. Recently, a new and complementary explanation has emerged to account for the generation of domains that are enriched by a subset of proteins or polarization determinants: local proteolysis. In this review, we discuss the most conspicuous proteolytic systems that may contribute to the generation of cell polarity, namely the ubiquitin-proteosome and the calpain systems. Specifically, we focus this review on two cellular processes that depend on the acquisition of cell polarity; cell migration and the establishment of an axon in a neuronal cell.

Tribbles homolog 2 (TRIB2) is a member of the mammalian Tribbles family of serine/threonine pseudokinases (TRIB1-3). Studies of TRIB2 indicate that many of the molecular interactions between the single Drosophila Tribbles (Trbl) protein and interacting partners are evolutionary conserved. In this study, we examined the relationship between TRIB2 and cell division cycle 25 (CDC25) family of dual-specificity protein phosphatases (mammalian homologues of Drosophila String), which are key physiological cell cycle regulators. Using co-immunoprecipitation we demonstrate that TRIB2 interacts with CDC25B and CDC25C selectively. Forced overexpression of TRIB2 caused a marked decrease in total CDC25C protein levels. Following inhibition of the proteasome, CDC25C was stabilized in the nuclear compartment. This implicates TRIB2 as a regulator of nuclear CDC25C turnover. In complementary ubiquitination assays, we show that TRIB2-mediated degradation of CDC25C is associated with lysine-48-linked CDC25C polyubiquitination driven by the TRIB2 kinase-like domain. A cell cycle associated role for TRIB2 is further supported by the cell cycle regulated expression of TRIB2 protein levels. Our findings reveal mitotic CDC25C as a new target of TRIB2 that is degraded via the ubiquitin proteasome system. Inappropriate CDC25C regulation could mechanistically underlie TRIB2 mediated regulation of cellular proliferation in neoplastic cells. [ABSTRACT FROM AUTHOR]

Danon disease, a condition characterized by cardiomyopathy, myopathy, and intellectual disability, is caused by mutations in the LAMP-2 gene. Lamp-2A protein, generated by alternative splicing from the Lamp-2 pre-mRNA, is reported to be the lysosomal membrane receptor essential for the chaperone-mediated autophagic pathway (CMA) aimed to selective protein targeting and translocation into the lysosomal lumen for degradation. To study the relevance of Lamp-2 in protein degradation, a lymphoblastoid cell line was obtained by EBV transformation of B-cells from a Danon patient. The derived cell line showed no significant expression of Lamp-2 protein. The steady-state mRNA and protein levels of alpha-synuclein, IΚBα, Rcan1, and glyceraldehyde-3-phosphate dehydrogenase, four proteins reported to be selective substrates of the CMA pathway, were similar in control and Lamp-2-deficient cells. Inhibition of protein synthesis showed that the half-life of alpha-synuclein, IΚBα, and Rcan1 was similar in control and Lamp-2-deficient cells, and its degradation prevented by proteasome inhibitors. Both in control and Lamp-2-deficient cells, induction of CMA and macroautophagy by serum and aminoacid starvation of cells for 8 h produced a similar decrease in IΚBα and Rcan1 protein levels and was prevented by the addition of lysosome and autophagy inhibitors. In conclusion, the results presented here showed that Lamp-2 deficiency in human lymphoblastoid cells did not modify the steady-state levels or the degradation of several protein substrates reported as selective substrates of the CMA pathway. [ABSTRACT FROM AUTHOR]

Primary hyperoxaluria type-3 is characterized by increased oxalate production caused by mutations in the HOGA1 gene encoding 4-hydroxy-2-oxoglutarate aldolase (HOGA1). How the most commonly occurring mutations affect the cellular fates of the expressed HOGA1 mutants is still unknown. We show that two prevalent recombinant HOGA1 mutants are thermally unstable with evidence for chaperone-mediated degradation when expressed in E. coli. In stably transformed HEK-293 cells, protein expression of the Glu315 deletion mutant only becomes detectable during incubation with a 26S proteasome inhibitor. These findings suggest that failure of chaperone-assisted folding leads to targeted cellular degradation and an absolute absence of HOGA1 function. [ABSTRACT FROM AUTHOR]

Tribbles family of pseudokinase proteins are known to mediate the degradation of target proteins in Drosophila and mammalian systems. The main protein proteolysis pathway in eukaryotic cells is the ubiquitin proteasome system (UPS). The tribbles homolog 2 (TRIB2) mammalian family member has been well characterized for its role in murine and human leukaemia, lung and liver cancer. One of the most characterized substrates for TRIB2-mediated degradation is the myeloid transcription factor CCAAT enhancer binding protein a (C/EBPa). However, across a number of cancers, the molecular interactions that take place between TRIB2 and factors involved in the UPS are varied and have differential downstream effects. This review summarizes our current knowledge of these interactions and how this information is important for our understanding of TRIB2 in cancer. [ABSTRACT FROM AUTHOR]

Skeletal muscle is a pivotal organ for the coordination of systemic metabolism, constituting one of the largest storage site for glucose, lipids and amino acids. Tight temporal orchestration of protein breakdown in times of fasting has to be balanced with preservation of muscle mass and function. However, the molecular mechanisms that control the fasting response in muscle are poorly understood. We now have identified a role for the peroxisome proliferator-activated receptor γ coactivator 1β (PGC-1β) in the regulation of catabolic pathways in this context in muscle-specific loss-of-function mouse models. Muscle-specific knockouts for PGC-1β experience mitigated muscle atrophy in fasting, linked to reduced expression of myostatin, atrogenes, activation of AMP-dependent protein kinase (AMPK) and other energy deprivation signaling pathways. At least in part, the muscle fasting response is modulated by a negative effect of PGC-1β on the nuclear factor of activated T-cells 1 (NFATC1). Collectively, these data highlight the complex regulation of muscle metabolism and reveal a new role for muscle PGC-1β in the control of proteostasis in fasting. • PGC-1β transcription in muscle is controlled by the feeding state. • Muscle-specific PGC-1β knockout mitigates fasting-induced muscle atrophy. • Muscle PGC-1β affects different signaling pathways linked to ubiquitination and proteostasis. • PGC-1β controls the balance between retaining muscle mass and catabolic processes. • These findings are important to understand contexts of muscle wasting as well as protection, e.g. in hibernation. [ABSTRACT FROM AUTHOR]

Skeletal muscle proteolysis is highly regulated, involving complex intramuscular proteolytic systems that recognize and degrade muscle proteins, and recycle free amino acid precursors for protein synthesis and energy production. Autophagy-lysosomal, calpain, and caspase systems are contributors to muscle proteolysis, although the ubiquitin proteasome system (UPS) is the primary mechanism by which actomyosin fragments are degraded in healthy muscle. The UPS is sensitive to mechanical force and nutritional deprivation, as recent reports have demonstrated increased proteolytic gene expression and activity of the UPS in response to resistance and endurance exercise, and short-term negative energy balance. However, consuming dietary protein alone (or free amino acids), or as a primary component of a mixed meal, may attenuate intramuscular protein loss by down-regulating proteolytic gene expression and the catabolic activity of the UPS. Although these studies provide novel insight regarding the intramuscular regulation of skeletal muscle mass, the role of proteolysis in the regulation of skeletal muscle protein turnover in healthy human muscle is not well described. This article provides a contemporary review of the intramuscular regulation of skeletal muscle proteolysis in healthy muscle, methodological approaches to assess proteolysis, and highlights the effects of nutrition and exercise on skeletal muscle proteolysis. © 2014 IUBMB Life, 66(7):478-484, 2014 [ABSTRACT FROM AUTHOR]

This study was undertaken to characterize the ubiquitin proteasome system (LIPS) response to varied dietary protein intake, energy deficit (ED), and consumption of a mixed meal. A randomized, controlled trial of 39 adults consuming protein at 0.8 (recommended dietary allowance [RDA]), 1.6 (2x-RDA), or 2.4 (3x-RDA) g ⋅ kg-1 ⋅ d-1 for 31 d. A 10-d weight maintenance (WM) period was followed by 21 d of 40% ED. Ubiquitin (Ub)-mediated proteolysis and associated gene expression were assessed in the post-absorptive (fasted) and postprandial (fed; 480 kcal, 20 g protein) states after WM and ED by using muscle biopsies, fluorescence-based assays, immunoblot analysis, and real-time qRT-PCR. In the assessment of LIPS responses to varied protein intakes, ED, and feeding, the RDA, WM, and fasted measures served as appropriate controls. ED resulted in the up-regulation of UPS-associated gene expression, as mRNA expression of the mrogenes muscle RING finger-1 (MuRF1) and atrogirvl were 1.2- and 1.3-fold higher (P<0.05) for ED than for WM. However, mixed-meal consumption attenuated UPS-mediated proteolysis, independent of energy status or dietary protein, as the activities of the 26S proteasome subunits β1, β2, and β5 were lower (P<0.05) for fed than for fasted. Muscle protein ubiquitylation was also 45% lower (P<0.05) for fed than for fasted, regardless of dietary protein and energy manipulations. Independent of habitual protein intake and despite increased MuRF1 and atrogin-1 mRNA expression during ED, consuming a protein-containing mixed meal attenuates Ub-mediated proteolysis. [ABSTRACT FROM AUTHOR]

he NAD-synthesizing enzyme TNMNAT2 is critical for axon survival in primary culture and its depletion may contribute to axon degeneration in a variety of neurodegenerative disorders. Here we discuss several recent reports from our laboratory that establish a critical role for NMNAT2 in axon growth in vivo in mice and shed light on the delivery and turnover of this survival factor in axons. In the absence of NMNAT2, axons fail to extend more than a short distance beyond the cell body during embryonic development, implying a requirement for NMNAT2 in axon maintenance even during development. Furthermore, we highlight findings regarding the bidirectional trafficking of NMNAT2 in axons on a vesicle population that undergoes fast axonal transport in primary culture neurites and in mouse sciatic nerve axons in vivo. Surprisingly, loss of vesicle association boosts the axon protective capacity of NMNAT2, an effect that is at least partially mediated by a longer protein half-life of cytosolic NMNAT2 variants. Analysis of wild-type and variant NMNAT2 in mouse sciatic nerves and Drosophila olfactory receptor neuron axons supports the existence of a similar mechanism in vivo, highlighting the potential for regulation of NMNAT2 stability and turnover as a mechanism to modulate axon degeneration in vivo. [ABSTRACT FROM AUTHOR]

Abstract: Parkinson disease (PD) is the most common movement disorder and is characterized by dopaminergic dysfunction. The majority of PD cases are sporadic; however, the discovery of genes linked to rare familial forms of the disease has provided crucial insight into the molecular mechanisms of disease pathogenesis. Multiple genes mediating familial forms of Parkinson’s disease (PD) have been identified, such as parkin (PARK2) and phosphatase and tensin homologue deleted on chromosome ten (PTEN)-induced putative kinase 1: PINK1 (PARK6). Here, we showed that Parkin directly interacts with PINK1, but did not bind to pathogenic PINK1 mutants. Parkin, but not its pathogenic mutants, stabilizes PINK1 by interfering with its degradation via the ubiquitin-mediated proteasomal pathway. In addition, the interaction between Parkin and PINK1 resulted in reciprocal reduction of their solubility. Our results indicate that Parkin regulates PINK1 stabilization via direct interaction with PINK1, and operates through a common pathway with PINK1 in the pathogenesis of early-onset PD. [Copyright &y& Elsevier]

Protein degradation in eukaryotic cells is mediated primarily by the ubiquitin-proteasome system and autophagy. Turnover of protein aggregates and other cytoplasmic components, including organelles, is another function attributed to autophagy. The ubiquitin-proteasome system and autophagy are essential for normal cell function but under certain pathological conditions can be overwhelmed, which can lead to adverse effects in cells. In this review we will focus primarily on the insulin-producing pancreatic β. Pancreatic βs respond to glucose levels by both producing and secreting insulin. The inability of βs to secrete sufficient insulin is a major contributory factor in the development of type 2 diabetes. The aim of this review is to examine some of the crucial roles of the ubiquitin-proteasome system and autophagy in normal pancreatic β function and how these pathways may become dysfunctional under pathological conditions associated with metabolic syndromes. Protein degradation in eukaryotic cells is mediated primarily by the ubiquitin-proteasome system and autophagy. Turnover of protein aggregates and other cytoplasmic components, including organelles, is another function attributed to autophagy. The ubiquitin-proteasome system and autophagy are essential for normal cell function but under certain pathological conditions can be overwhelmed, which can lead to adverse effects in cells. In this review we will focus primarily on the insulin-producing pancreatic β-cell. Pancreatic β-cells respond to glucose levels by both producing and secreting insulin. The inability of β-cells to secrete sufficient insulin is a major contributory factor in the development of type 2 diabetes. The aim of this review is to examine some of the crucial roles of the ubiquitin-proteasome system and autophagy in normal pancreatic β-cell function and how these pathways may become dysfunctional under pathological conditions associated with metabolic syndromes. [ABSTRACT FROM AUTHOR]

Abstract: NSAIDs cause severe gastrointestinal injury, in part by suppressing survivin, an inhibitor of apoptosis protein, both in cultured gastric epithelial cells and in human and rat gastric mucosa. The mechanism(s) of survivin down-regulation by NSAIDs is unclear. In this study, we examined whether NSAID treatment decreases survivin mRNA expression and/or enhances degradation of survivin protein via ubiquitin proteasome system in rat gastric mucosal, RGM-1 cells, and whether survivin overexpression prevents indomethacin-induced cell injury and apoptosis. Effects of indomethacin on survivin mRNA expression, survivin protein half-life and ubiquitination were examined in RGM-1 cells. Proteasome inhibitors were utilized to prevent indomethacin-induced survivin protein degradation in RGM-1 cells. The effects of stable overexpression of survivin on indomethacin-induced RGM-1 cell injury and apoptosis were examined. Results showed: (1) Indomethacin treatment did not alter survivin mRNA expression, but significantly reduced survivin protein half-life from 1.5h to approximately 1h and increased survivin ubiquitination. (2) Inhibition of ubiquitin proteasome prolonged survivin protein half-life to over 2h and inhibited indomethacin-induced survivin degradation. (3) Overexpression of survivin significantly reduced indomethacin-induced cell injury and apoptosis. In conclusion, indomethacin treatment enhances degradation of survivin via the ubiquitin proteasome machinery in RGM-1 cells, and maintenance of survivin levels is important for prevention of gastric epithelial cell injury and apoptosis. [Copyright &y& Elsevier]

Nanoscale materials are presently under development for diagnostic (nanomedicine) and electronic purposes. In contrast to the potential benefits of nanotechnology, the effects of nanomaterials on human health are poorly understood. Nanomaterials are known to translocate into the circulation and could thus directly affect vascular endothelial cells (ECs), causing vascular injury that might be responsible for the development of atherosclerosis. To explore the direct effects of nanomaterials on endothelial toxicity, human umbilical vein ECs were treated with 1–100 μg/ml hydroxyl fullerene [C6o(0H)24, mean diameter, 7.1 ± 2.4 nm] for 24 h. C60(OH)24 induced cytotoxic morphological changes such as cytosolic vacuole formation and decreased cell density in a dose-dependent manner. Lactate dehydrogenase assay revealed that a maximal dose of C60(OH)24 (100 μg/ml) induced cytotoxic injury. Proliferation assay also showed that a maximal dose of C60(OH)24 inhibited EC growth. C60(OH)24 did not seem to induce apoptosis but caused the accumulation of polyubiquitinated proteins and facilitated autophagic cell death. Formation of autophagosomes was confirmed on the basis of Western blot analysis using a specific marker, light chain 3 antibody, and electron microscopy. Chronic treatment with low-dose C60(OH)24 (10 μg/ml for 8 days) inhibited cell attachment and delayed EC growth. In the present study, we have examined, for the first time, the toxicity of water-soluble fullerenes to ECs. Although fullerenes changed morphology in a dose-dependent manner, only maximal doses of fullerenes caused cytotoxic injury and/or death and inhibited cell growth. EC death seemed to be caused by activation of ubiquitin-autophagy cell death pathways. Although exposure to nanomaterials appears to represent a risk for cardiovascular disorders, further in vivo validations are necessary. [ABSTRACT FROM AUTHOR]