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1

Exploring the Ubiquitin-Proteasome Protein Degradation Pathway in Yeast  

ERIC Educational Resources Information Center

This article describes an undergraduate biochemistry laboratory investigating the ubiquitin-proteasome pathway in yeast. In this exercise, the enzyme beta-galactosidase (beta-gal) is expressed in yeast under the control of a stress response promoter. Following exposure to heat stress to induce beta-gal expression, cycloheximide is added to halt…

Will, Tamara J.; McWatters, Melissa K.; McQuade, Kristi L.

2006-01-01

2

GSH-MODIFIED GAMMAC-CRYSTALLIN IS SELECTIVELY DEGRADED BY THE UBIQUITIN-PROTEASOME PATHWAY  

Technology Transfer Automated Retrieval System (TEKTRAN)

The purpose of this research was to explore the ubiquitin-proteasome pathway (UPP) as an important protein quality control process by which cells selectively degrade and remove damaged proteins. Gamma-crystallin is susceptible to formation of mixed disulfides with GSH upon oxidative stress. The aim ...

3

The significance of ubiquitin proteasome pathway in cancer development.  

PubMed

The ubiquitin proteasome pathway is the most significant intracellular proteolytic pathway. The target proteins are usually ubiquitinated prior to degradation by the proteasome; however, ubiquitin-independent targeting mechanisms have also been reported (e.g., the antizyme-mediated degradation of ornithine decarboxylase). Aberrations in the components of the ubiquitin proteasome pathway are commonly observed in many cancers, and uncontrolled growth of cancer cells can result either from stabilization of oncoproteins (e.g., c-jun) or increased degradation of tumor suppressor proteins (e.g., p53). In addition, due to the pleiotropic functions of the ubiquitin proteasome pathway in cells, there is great interest in developing inhibitors to specifically block this pathway for cancer treatment. This review summarizes the recent literature and several patented inventions on the ubiquitin proteasome pathway with respect to its role in cancer development and treatment. PMID:23061719

Yerlikaya, Azmi; Yöntem, Mustafa

2013-09-01

4

REG? regulates ER? degradation via ubiquitin-proteasome pathway in breast cancer.  

PubMed

REG? is a proteasome coactivator which regulates proteolytic activity in eukaryotic cells. Abundant lines of evidence have showed that REG? is over expressed in a number of human carcinomas. However, its precise role in the pathogenesis of cancer is still unclear. In this study, by examining 200 human breast cancer specimens, we demonstrated that REG? was highly expressed in breast cancers, and the expression of REG? was positively correlated with breast cancer patient estrogen receptor alpha (ER?) status. Moreover, the expression of REG? was found positively associated with poor clinical features and low survival rates in ER? positive breast cancer patients. Further cell culture studies using MCF7 and BT474 breast cancer cell lines showed that cell proliferation, motility, and invasion capacities were decreased significantly by REG? knockdown. Lastly, we demonstrated that REG? indirectly regulates the degradation of ER? protein via ubiquitin-proteasome pathway. In conclusion, our findings provide the evidence that REG? expression was positively correlated with ER? status and poor clinical prognosis in ER? positive breast cancer patients. As well, we disclose a new connection between the two molecules that are both highly expressed in most breast cancer cases. PMID:25490392

Chai, Fan; Liang, Yan; Bi, Jiong; Chen, Li; Zhang, Fan; Cui, Youhong; Jiang, Jun

2015-01-01

5

Glutathiolation Enhances the Degradation of ?C-crystallin in Lens and Reticulocyte Lysates, Partially via the Ubiquitin–Proteasome Pathway  

PubMed Central

PURPOSE S-glutathiolated proteins are formed in the lens during aging and cataractogenesis. The objective of this work was to explore the role of the ubiquitin–proteasome pathway in eliminating S-glutathiolated ?C-crystallin. METHODS Recombinant human ?C-crystallin was mixed with various concentrations of glutathione (GSH) and diamide at 25°C for 1 hour. The extent of glutathiolation of the ?C-crystallin was determined by mass spectrometry. Native and S-glutathiolated ?C-crystallins were labeled with 125I, and proteolytic degradation was determined using both lens fiber lysate and reticulocyte lysate as sources of ubiquitinating and proteolytic enzymes. Far UV circular dichroism, tryptophan fluorescence intensity, and binding to the hydrophobic fluorescence probe 4,4?-dianilino-1,1?-binaphthalene-5,5?-disulfonic acid (Bis-ANS), were used to characterize the native and glutathiolated ?C-crystallins. RESULTS On average, two and five of the eight cysteines in ?C-crystallin were glutathiolated when molar ratios of ?C-crystallin-GSH-diamide were 1:2:5 and 1:10:25, respectively. Native ?C-crystallin was resistant to degradation in both lens fiber lysate and reticulocyte lysate. However, glutathiolated ?C-crystallin showed a significant increase in proteolytic degradation in both lens fiber and reticulocyte lysates. Proteolysis was stimulated by addition of adenosine triphosphate (ATP) and Ubc4 and was substantially inhibited by the proteasome inhibitor MG132 and a dominant negative form of ubiquitin, indicating that at least part of the proteolysis was mediated by the ubiquitin-proteasome pathway. Spectroscopic analyses of glutathiolated ?C-crystallin revealed conformational changes and partial unfolding, which may provide a signal for the ubiquitin-dependent degradation. CONCLUSIONS The present data demonstrate that oxidative modification by glutathiolation can render lens proteins more susceptible to degradation by the ubiquitin–proteasome pathway. Together with previous results, these data support the concept that the ubiquitin–proteasome pathway serves as a general protein quality-control mechanism. PMID:16877417

Zetterberg, Madeleine; Zhang, Xinyu; Taylor, Allen; Liu, Bingfen; Liang, Jack J.; Shang, Fu

2007-01-01

6

Cancer cell death induced by novel small molecules degrading the TACC3 protein via the ubiquitin-proteasome pathway.  

PubMed

The selective degradation of target proteins with small molecules is a novel approach to the treatment of various diseases, including cancer. We have developed a protein knockdown system with a series of hybrid small compounds that induce the selective degradation of target proteins via the ubiquitin-proteasome pathway. In this study, we designed and synthesized novel small molecules called SNIPER(TACC3)s, which target the spindle regulatory protein transforming acidic coiled-coil-3 (TACC3). SNIPER(TACC3)s induce poly-ubiquitylation and proteasomal degradation of TACC3 and reduce the TACC3 protein level in cells. Mechanistic analysis indicated that the ubiquitin ligase APC/C(CDH1) mediates the SNIPER(TACC3)-induced degradation of TACC3. Intriguingly, SNIPER(TACC3) selectively induced cell death in cancer cells expressing a larger amount of TACC3 protein than normal cells. These results suggest that protein knockdown of TACC3 by SNIPER(TACC3) is a potential strategy for treating cancers overexpressing the TACC3 protein. PMID:25375378

Ohoka, N; Nagai, K; Hattori, T; Okuhira, K; Shibata, N; Cho, N; Naito, M

2014-01-01

7

A nuclear ubiquitin-proteasomal pathway targets inner nuclear membrane protein Asi2 for degradation  

PubMed Central

The nuclear envelope consists of inner and outer nuclear membranes. While the outer membrane is an extension of the endoplasmic reticulum, the inner nuclear membrane represents a unique membranous environment containing specific proteins. The mechanisms of integral inner nuclear membrane protein degradation are unknown. Here we investigated the turnover of Asi2, an integral INM protein in Saccharomyces cerevisiae. We report that Asi2 is degraded by the proteasome and independent of the vacuole exhibiting a half-life of ? 45 min. Asi2 exhibits enhanced stability in mutants lacking the E2 ubiquitin conjugating enzymes Ubc6 or Ubc7, or the E3 ubiquitin ligase Doa10. Consistently, Asi2 is post-translationally modified by poly-ubiquitylation in a Ubc7- and Doa10-dependent manner. Importantly Asi2 degradation is significantly reduced in a sts1-2 mutant that fails to accumulate proteasomes in the nucleus, indicating that Asi2 is degraded in the nucleus. Our results reveal a molecular pathway that affects the stability of integral proteins of the inner nuclear membrane and indicate that Asi2 is subject to protein quality control in the nucleus. PMID:24928896

Boban, Mirta; Pantazopoulou, Marina; Schick, Anna; Ljungdahl, Per O.; Foisner, Roland

2015-01-01

8

Genome-wide approaches to systematically identify substrates of the ubiquitin/proteasome pathway  

PubMed Central

The ubiquitin/proteasome system handles the majority of controlled proteolysis in eukaryotes. Defects in the ubiquitin/proteasome system have been implicated in diseases ranging from cancers to neurodegenerative disorders. However, the precise role of ubiquitin/proteasome-mediated degradation in health and disease is far from clear. A major challenge is to link specific substrates directly to a particular degradation pathway. Here, we review genome-wide approaches that have been developed in recent years to comprehensively identify ubiquitylated substrates of a particular pathway. The components of the ubiquitin/proteasome system are attractive drug targets, as illustrated by the efficacy of some proteasome inhibitors in the treatment of multiple myeloma. Information that has emerged from these studies could reveal novel drug targets and strategies for treating human diseases. PMID:20637515

Liu, Chang; Choe, Vitnary; Rao, Hai

2010-01-01

9

Selective degradation of aggregate-prone CryAB mutants by HSPB1 is mediated by ubiquitin-proteasome pathways.  

PubMed

Disease-causing mutations of genes encoding small MW heat shock proteins (sHSPs) constitute a growing family of inherited myofibrillar disorders. In the present work, we found that three structurally-distinct CryAB mutants R120G, 450delA and 464delCT are mostly present in the detergent-insoluble fractions when overexpressed in H9c2 rat heart cells. We found that either over-expression or knockdown of HSPB1, a related sHSP, affects the solubility, stability, and degradation of aggregation-prone CryAB mutants. HSPB1 overexpression has negligible effects on the solubility and protein aggregates of either R120G and/or 450delA but increased the solubility and prevented formation of 464delCT aggregates. HSPB1 knockdown decreased solubility and increased protein aggregates of all CryAB mutants, indicating a key role for HSPB1 in clearance of CryAB mutants under basal conditions. We provide four lines of evidence that such selective clearance of R120G, 450delA and 464delCT mutants by HSPB1 is mediated by the ubiquitin-proteasome system (UPS). First, we found that treatment with the proteasome inhibitors increased the levels of all CryAB mutants. Second, R120G and 450delA overexpression corresponded to the accumulation of their specific ubiquitin conjugates in H9c2 cells. Third, HSPB1 knockdown directly increased the levels of all polyubiquitin conjugates. And fourth, the selective attenuation of 464delCT expression by HSPB1 over-expression was abrogated by the proteasome inhibition. We conclude that such selective interactions between CryAB mutants and HSPB1 overexpression might have important implications for the clinical manifestations and potential treatment. PMID:20863832

Zhang, Huali; Rajasekaran, Namakkal S; Orosz, Andras; Xiao, Xianzhong; Rechsteiner, Martin; Benjamin, Ivor J

2010-12-01

10

Selective degradation of aggregate-prone CryAB mutants by HSPB1 is mediated by ubiquitin–proteasome pathways  

PubMed Central

Disease-causing mutations of genes encoding small MW heat shock proteins (sHSPs) constitute a growing family of inherited myofibrillar disorders. In the present work, we found that three structurally-distinct CryAB mutants R120G, 450delA and 464delCT are mostly present in the detergent-insoluble fractions when overexpressed in H9c2 rat heart cells. We found that either over-expression or knockdown of HSPB1, a related sHSP, affects the solubility, stability, and degradation of aggregation-prone CryAB mutants. HSPB1 overexpression has negligible effects on the solubility and protein aggregates of either R120G and/or 450delA but increased the solubility and prevented formation of 464delCT aggregates. HSPB1 knockdown decreased solubility and increased protein aggregates of all CryAB mutants, indicating a key role for HSPB1 in clearance of CryAB mutants under basal conditions. We provide four lines of evidence that such selective clearance of R120G, 450delA and 464delCT mutants by HSPB1 is mediated by the ubiquitin–proteasome system (UPS). First, we found that treatment with the proteasome inhibitors increased the levels of all CryAB mutants. Second, R120G and 450delA overexpression corresponded to the accumulation of their specific ubiquitin conjugates in H9c2 cells. Third, HSPB1 knockdown directly increased the levels of all polyubiquitin conjugates. And fourth, the selective attenuation of 464delCT expression by HSPB1 over-expression was abrogated by the proteasome inhibition. We conclude that such selective interactions between CryAB mutants and HSPB1 overexpression might have important implications for the clinical manifestations and potential treatment. PMID:20863832

Zhang, Huali; Rajasekaran, Namakkal S.; Orosz, Andras; Xiao, Xianzhong; Rechsteiner, Martin; Benjamin, Ivor J.

2010-01-01

11

The Ubiquitin-Proteasome Pathway and Synaptic Plasticity  

ERIC Educational Resources Information Center

Proteolysis by the ubiquitin-proteasome pathway (UPP) has emerged as a new molecular mechanism that controls wide-ranging functions in the nervous system, including fine-tuning of synaptic connections during development and synaptic plasticity in the adult organism. In the UPP, attachment of a small protein, ubiquitin, tags the substrates for…

Hegde, Ashok N.

2010-01-01

12

Expression of the ubiquitin-proteasome pathway and muscle loss in experimental cancer cachexia  

Microsoft Academic Search

Muscle protein degradation is thought to play a major role in muscle atrophy in cancer cachexia. To investigate the importance of the ubiquitin-proteasome pathway, which has been suggested to be the main degradative pathway mediating progressive protein loss in cachexia, the expression of mRNA for proteasome subunits C2 and C5 as well as the ubiquitin-conjugating enzyme, E214k, has been determined

J Khal; S M Wyke; S T Russell; A V Hine; M J Tisdale

2005-01-01

13

The ubiquitin ligase FBG1 promotes the degradation of the disease-linked protein torsinA through the ubiquitin proteasome pathway and macroautophagy  

PubMed Central

DYT1 dystonia is a dominantly inherited, disabling neurological disorder with low penetrance that is caused by the deletion of a glutamic acid (?E) in the protein torsinA. We previously showed that torsinA(wt) is degraded through macroautophagy while torsinA(?E) is targeted to the ubiquitin proteasome pathway (UPP). The different catabolism of torsinA(wt) and (?E) potentially modulates torsinA(wt):torsinA(?E) stoichiometry. Therefore, gaining a mechanistic understanding on how the protein quality control machinery clears torsinA(?E) in neurons may uncover important regulatory steps in disease pathogenesis. Here, we asked whether FBG1, a ubiquitin ligase known to degrade neuronal glycoproteins, is implicated in the degradation of torsinA(?E) by the UPP. In a first set of studies completed in cultured cells, we show that FBG1 interacts with and influences the steady-state levels of torsinA(wt) and (?E). Interestingly, FBG1 achieves this effect promoting the degradation of torsinA not only through the UPP, but also by macroautophagy. To determine the potential clinical significance of these findings, we asked if eliminating expression of Fbg1 triggers a motor phenotype in torsinA(?E) knock in mice, a model of non-manifesting DYT1 mutation carriers. We detected differences in spontaneous locomotion between aged torsinA(?E) knock in-Fbg1 knock out and control mice. Furthermore, neuronal levels of torsinA were unaltered in Fbg1 null mice, indicating that redundant systems likely compensate in vivo for the absence of this ubiquitin ligase. In summary, our studies support a non-essential role for FBG1 on the degradation of torsinA and uncover a novel link of FBG1 to the autophagy pathway. PMID:22917612

Gordon, Kara L; Glenn, Kevin A; Bode, Nicole; Wen, Hsiang M; Paulson, Henry L; Gonzalez-Alegre, Pedro

2012-01-01

14

Fucoidan inhibition of lung cancer in vivo and in vitro : role of the Smurf2-dependent ubiquitin proteasome pathway in TGF? receptor degradation.  

PubMed

Fucoidan, a polysaccharide extracted from brown seaweeds, reduces tumor cell proliferation. In this study, we demonstrate that fucoidan reduces tumor size in LLC1-xenograft male C57BL/6 mice. Moreover, we found that LLC1-bearing mice continuously fed fucoidan showed greater antitumor activity than mice with discontinuous feeding. Fucoidan inhibited the in vitro growth of lung cancer cells. Transforming growth factor ? (TGF?) receptors (TGFRs) play important roles in the regulation of proliferation and progression, and high TGFRI expression in lung cancer specimens is associated with a worse prognosis. Herein, using lung cancer cells, we found that fucoidan effectively reduces TGFRI and TGFRII protein levels in vivo and in vitro. Moreover, fucoidan reduces TGFR downstream signaling events, including those in Smad2/3 and non-Smad pathways: Akt, Erk1/2, and FAK phosphorylation. Furthermore, fucoidan suppresses lung cancer cell mobility upon TGF? stimulation. To elucidate how fucoidan decreases TGFR proteins in lung cancer cells, we found that fucoidan enhances the ubiquitination proteasome pathway (UPP)-mediated degradation of TGFRs in A549 and CL1-5 cells. Mechanistically, fucoidan promotes Smurf2 and Smad7 to conjugate TGFRs, resulting in TGF degradation; however, Smurf2-shRNA abolishes fucoidan-enhanced UPP-mediated TGFR degradation. Our study is the first to identify a novel mechanism for the antitumor activity of fucoidan, namely decreasing tumor growth by modulating the TGFR/Smad7/Smurf2-dependent axis, leading to TGFR protein degradation and inhibition of lung cancer cell progression in vitro and in vivo. Our current findings indicate that fucoidan is a potential therapeutic agent or dietary supplementation for lung cancer, acting via the Smurf2-dependent ubiquitin degradation of TGF? receptors. PMID:25149540

Hsu, Hsien-Yeh; Lin, Tung-Yi; Wu, Yu-Chung; Tsao, Shu-Ming; Hwang, Pai-An; Shih, Yu-Wei; Hsu, Jason

2014-09-15

15

The ubiquitin-proteasome pathway and synaptic plasticity  

PubMed Central

Proteolysis by the ubiquitin-proteasome pathway (UPP) has emerged as a new molecular mechanism that controls wide-ranging functions in the nervous system, including fine-tuning of synaptic connections during development and synaptic plasticity in the adult organism. In the UPP, attachment of a small protein, ubiquitin, tags the substrates for degradation by a multisubunit complex called the proteasome. Linkage of ubiquitin to protein substrates is highly specific and occurs through a series of well-orchestrated enzymatic steps. The UPP regulates neurotransmitter receptors, protein kinases, synaptic proteins, transcription factors, and other molecules critical for synaptic plasticity. Accumulating evidence indicates that the operation of the UPP in neurons is not homogeneous and is subject to tightly managed local regulation in different neuronal subcompartments. Investigations on both invertebrate and vertebrate model systems have revealed local roles for enzymes that attach ubiquitin to substrate proteins, as well as for enzymes that remove ubiquitin from substrates. The proteasome also has been shown to possess disparate functions in different parts of the neuron. Here I give a broad overview of the role of the UPP in synaptic plasticity and highlight the local roles and regulation of the proteolytic pathway in neurons. PMID:20566674

Hegde, Ashok N.

2010-01-01

16

Inhibition of PCSK9 Transcription by Berberine Involves Down-regulation of Hepatic HNF1? Protein Expression through the Ubiquitin-Proteasome Degradation Pathway.  

PubMed

Our previous in vitro studies have identified hepatocyte nuclear factor 1? (HNF1?) as an obligated trans-activator for PCSK9 gene expression and demonstrated its functional involvement in the suppression of PCSK9 expression by berberine (BBR), a natural cholesterol-lowering compound. In this study, we investigated the mechanism underlying the inhibitory effect of BBR on HNF1?-mediated PCSK9 transcription. Administration of BBR to hyperlipidemic mice and hamsters lowered circulating PCSK9 concentrations and hepatic PCSK9 mRNA levels without affecting the gene expression of HNF1?. However, hepatic HNF1? protein levels were markedly reduced in BBR-treated animals as compared with the control. Using HepG2 cells as a model system, we obtained evidence that BBR treatment let to accelerated degradation of HNF1? protein. By applying inhibitors to selectively block the ubiquitin proteasome system (UPS) and autophagy-lysosomal pathway, we show that HNF1? protein content in HepG2 cells was not affected by bafilomycin A1 treatment, but it was dose-dependently increased by UPS inhibitors bortezomib and MG132. Bortezomib treatment elevated HNF1? and PCSK9 cellular levels with concomitant reductions of LDL receptor protein. Moreover, HNF1? protein displayed a multiubiquitination ladder pattern in cells treated with BBR or overexpressing ubiquitin. By expressing GFP-HNF1? fusion protein in cells, we observed that blocking UPS resulted in accumulation of GFP-HNF1? in cytoplasm. Importantly, we show that the BBR reducing effects on HNF1? protein and PCSK9 gene transcription can be eradicated by proteasome inhibitors. Altogether, our studies using BBR as a probe uncovered a new aspect of PCSK9 regulation by ubiquitin-induced proteasomal degradation of HNF1?. PMID:25540198

Dong, Bin; Li, Hai; Singh, Amar Bahadur; Cao, Aiqin; Liu, Jingwen

2015-02-13

17

Regulating the ubiquitin/proteasome pathway via cAMP-signaling: neuroprotective potential  

PubMed Central

The cAMP-signaling pathway has been under intensive investigation for decades. It is a wonder that such a small simple molecule like cAMP can modulate a vast number of diverse processes in different types of cells. The ubiquitous involvement of cAMP-signaling in a variety of cellular events requires tight spatial and temporal control of its generation, propagation, compartmentalization, and elimination. Among the various steps of the cAMP-signaling pathway, G-protein coupled receptors, adenylate cyclases, phosphodiesterases, the two major cAMP targets, i.e. protein kinase A and exchange protein activated by cAMP, as well as the A-kinase anchoring proteins, are potential targets for drug development. Herein we review the recent progress on the regulation and manipulation of different steps of the cAMP-signaling pathway. We end by focusing on the emerging role of cAMP-signaling in modulating protein degradation via the ubiquitin/proteasome pathway. New discoveries on the regulation of the ubiquitin/proteasome pathway by cAMP-signaling support the development of new therapeutic approaches to prevent proteotoxicity in chronic neurodegenerative disorders and other human disease conditions associated with impaired protein turnover by the ubiquitin/proteasome pathway and the accumulation of ubiquitin-protein aggregates. PMID:23686612

Huang, He; Wang, Hu; Figueiredo-Pereira, Maria E.

2013-01-01

18

The cell senescence inducing gene product MORF4 is regulated by degradation via the ubiquitin/proteasome pathway  

PubMed Central

After undergoing several rounds of divisions normal human fibroblasts enter a terminally non-dividing state referred to as cellular or replicative senescence. We cloned MORF4 (mortality factor on human chromosome 4), as a cellular senescence inducing gene that caused immortal cells assigned to complementation group B for indefinite division to stop dividing. To facilitate analyses of this gene, which is toxic to cells at low levels, we obtained stable clones of HeLa cells expressing a tetracycline-induced MORF4 construct that could be induced by doxycycline in a dose-dependent manner. MORF4 induction resulted in reduced colony formation after 14 days of culture, as previously observed. We determined that MORF4 protein was unstable and that addition of the proteasome inhibitor MG132 resulted in accumulation of the protein. Following removal of MG132 the protein was rapidly degraded. Subcellular fractionation following MG132 treatment demonstrated that the protein accumulates primarily in the cytoplasm with some amounts present in the nucleus. It is therefore possible that MORF4 protein, which escapes degradation in the cytoplasm, is transported to the nucleus where it is functional. The results suggest that levels of MORF4 in cells must be tightly controlled and one mechanism involves stability of the protein. PMID:19769966

Tominaga, Kaoru; Tominaga, Emiko; Ausserlechner, Michael J.; Pereira-Smith, Olivia M.

2009-01-01

19

Role of the Ubiquitin-Proteasome Pathway in Regulating Abundance of the Cyclin-Dependent Kinase Inhibitor p27  

Microsoft Academic Search

The p27 mammalian cell cycle protein is an inhibitor of cyclin-dependent kinases. Both in vivo and in vitro, p27 was found to be degraded by the ubiquitin-proteasome pathway. The human ubiquitin-conjugating enzymes Ubc2 and Ubc3 were specifically involved in the ubiquitination of p27. Compared with proliferating cells, quiescent cells exhibited a smaller amount of p27 ubiquitinating activity, which accounted for

Michele Pagano; Sun W. Tam; Anne M. Theodoras; Peggy Beer-Romero; Giannino del Sal; Vincent Chau; P. Renee Yew; Giulio F. Draetta; Mark Rolfe

1995-01-01

20

Protein Degradation by Ubiquitin-Proteasome System in Formation and Labilization of Contextual Conditioning Memory  

ERIC Educational Resources Information Center

The ubiquitin-proteasome system (UPS) of protein degradation has been evaluated in different forms of neural plasticity and memory. The role of UPS in such processes is controversial. Several results support the idea that the activation of this system in memory consolidation is necessary to overcome negative constrains for plasticity. In this…

Fustiñana, María Sol; de la Fuente, Verónica; Federman, Noel; Freudenthal, Ramiro; Romano, Arturo

2014-01-01

21

The ubiquitin-proteasome pathway: review of a novel intracellular mechanism of muscle protein breakdown during sepsis and other catabolic conditions.  

PubMed Central

SUMMARY BACKGROUND DATA: Patients with sepsis and other catabolic conditions, such as severe trauma, cancer, and fasting, suffer significant loss of body protein, the majority of which originates from skeletal muscle. Recent evidence suggests that muscle protein breakdown during sepsis is caused by upregulated activity in the ubiquitin-proteasome pathway and is associated with increased expression of the ubiquitin gene. PURPOSE: The purpose of the study was to review the role of the ubiquitin-proteasome pathway in the regulation of muscle proteolysis during sepsis and other catabolic conditions. REVIEW: Proteins that are degraded by the ubiquitin-proteasome mechanism are first conjugated to ubiquitin, a 76-amino-acid, highly conserved residue. Ubiquitinated proteins are recognized by the 26S proteasome, which is a large proteolytic complex consisting of the 19S cap complex and the 20S proteasome. The 20S proteasome is a cylindrical particle composed of four stacked rings, making it look like a barrel. The rings form a "tunnel" in which the target proteins are hydrolyzed, after which ubiquitin is released to be reused in the proteolytic pathway. A unique feature of the ubiquitin-proteasome proteolytic pathway is its energy dependency. CONCLUSIONS: An understanding of the molecular regulation of protein metabolism in patients with sepsis and other catabolic conditions is important because it may form the basis for improved treatment in the future. Images Figure 2. Figure 5. Figure 6. Figure 7. PMID:9060588

Hasselgren, P O; Fischer, J E

1997-01-01

22

Identification of the ubiquitin-proteasome pathway in the regulation of the stability of eukaryotic elongation factor-2 kinase.  

PubMed

Eukaryotic elongation factor-2 kinase (eEF-2 kinase) is a highly conserved calcium/calmodulin-dependent enzyme involved in the regulation of protein translation and cell proliferation. Rapid changes in the activity and abundance of eEF-2 kinase have been observed on growth stimulation, and increased enzyme activity is characteristic of malignant cell growth. Yet the mechanism for controlling the turnover of this kinase is unknown. The ubiquitin-proteasome pathway regulates the degradation of many cellular proteins, including transcription factors, cell cycle regulators, and signal transduction proteins. Therefore, we determined whether the ubiquitin-proteasome pathway regulates the turnover of eEF-2 kinase. We found that eEF-2 kinase was a relatively short-lived protein with a half-life of less than 6 hours. eEF-2 kinase was ubiquitinated in vivo as determined by coimmunoprecipitation and polyubiquitin affinity matrix. Incubation of purified eEF-2 kinase with a source of ubiquitination enzymes (rabbit reticulocyte lysate), purified ubiquitin, and ATP revealed the presence of increasing molecular weight species of ubiquitinated eEF-2 kinase. Treatment of cells with MG132, a proteasome inhibitor, inhibited eEF-2 kinase degradation and induced the accumulation of polyubiquitinated forms of the enzyme, resulting in an increase in its half-life. These results suggest involvement of the proteasome in the turnover of the ubiquitinated kinase. Because eEF-2 kinase is chaperoned by heat shock protein 90 (Hsp90), we next determined if disruption of the Hsp90-eEF-2 kinase complex promoted degradation of the kinase. Treatment of cells with geldanamycin, an Hsp90 inhibitor, enhanced ubiquitination of eEF-2 kinase and decreased the half-life of the kinase to less than 2 hours. These results indicate that cellular levels of eEF-2 kinase are maintained by a balance between association with Hsp90 and degradation by the ubiquitin-proteasome pathway. In conclusion, these data show that the turnover of eEF-2 kinase is regulated by the ubiquitin-proteasome pathway and, therefore, modulating the ubiquitination of eEF-2 kinase might control the abundance of this enzyme and have implications in the treatment of certain forms of cancer. PMID:15867377

Arora, Sonia; Yang, Jin-Ming; Hait, William N

2005-05-01

23

PTEN Increases Autophagy and Inhibits the Ubiquitin-Proteasome Pathway in Glioma Cells Independently of its Lipid Phosphatase Activity  

PubMed Central

Two major mechanisms of intracellular protein degradation, autophagy and the ubiquitin-proteasome pathway, operate in mammalian cells. PTEN, which is frequently mutated in glioblastomas, is a tumor suppressor gene that encodes a dual specificity phosphatase that antagonizes the phosphatidylinositol 3-kinase class I/AKT/mTOR pathway, which is a key regulator of autophagy. Here, we investigated in U87MG human glioma cells the role of PTEN in the regulation of autophagy and the ubiquitin-proteasome pathway, because both are functionally linked and are relevant in cancer progression. Since U87MG glioma cells lack a functional PTEN, we used stable clones that express, under the control of a tetracycline-inducible system (Tet-on), wild-type PTEN and two of its mutants, G129E-PTEN and C124S-PTEN, which, respectively, lack the lipid phosphatase activity only and both the lipid and the protein phosphatase activities of this protein. Expression of PTEN in U87MG glioma cells decreased proteasome activity and also reduced protein ubiquitination. On the contrary, expression of PTEN increased the autophagic flux and the lysosomal mass. Interestingly, and although PTEN negatively regulates the phosphatidylinositol 3-kinase class I/AKT/mTOR signaling pathway by its lipid phosphatase activity, both effects in U87MG cells were independent of this activity. These results suggest a new mTOR-independent signaling pathway by which PTEN can regulate in opposite directions the main mechanisms of intracellular protein degradation. PMID:24349488

Errafiy, Rajaa; Aguado, Carmen; Ghislat, Ghita; Esteve, Juan M.; Gil, Anabel; Loutfi, Mohammed; Knecht, Erwin

2013-01-01

24

Contributions of the ubiquitin–proteasome pathway and apoptosis to human skeletal muscle wasting with age  

Microsoft Academic Search

The primary mechanism that contributes to decreasing skeletal muscle strength and size with healthy aging is not presently known. This study examined the contribution of the ubiquitin–proteasome pathway and apoptosis to skeletal muscle wasting in older adults (n = 21; mean age = 72.76 ± 8.31 years) and young controls (n = 21; mean age = 21.48 ± 2.93 years). Subjects underwent

Samantha A. Whitman; Michael J. Wacker; Scott R. Richmond; Michael P. Godard

2005-01-01

25

An optimal ubiquitin-proteasome pathway in the nervous system: the role of deubiquitinating enzymes  

PubMed Central

The Ubiquitin-Proteasome Pathway (UPP), which is critical for normal function in the nervous system and is implicated in various neurological diseases, requires the small modifier protein ubiquitin to accomplish its duty of selectively degrading short-lived, abnormal or misfolded proteins. Over the past decade, a large class of proteases collectively known as deubiquitinating enzymes (DUBs) has increasingly gained attention in all manners related to ubiquitin. By cleaving ubiquitin from another protein, DUBs ensure that the UPP functions properly. DUBs accomplish this task by processing newly translated ubiquitin so that it can be used for conjugation to substrate proteins, by regulating the “where, when, and why” of UPP substrate ubiquitination and subsequent degradation, and by recycling ubiquitin for re-use by the UPP. Because of the reliance of the UPP on DUB activities, it is not surprising that these proteases play important roles in the normal activities of the nervous system and in neurodegenerative diseases. In this review, we summarize recent advances in understanding the functions of DUBs in the nervous system. We focus on their role in the UPP, and make the argument that understanding the UPP from the perspective of DUBs can yield new insight into diseases that result from anomalous intra-cellular processes or inter-cellular networks. Lastly, we discuss the relevance of DUBs as therapeutic options for disorders of the nervous system. PMID:25191222

Ristic, Gorica; Tsou, Wei-Ling; Todi, Sokol V.

2014-01-01

26

Malformed selenoproteins are removed by the ubiquitin--proteasome pathway in Stanleya pinnata.  

PubMed

Despite the widely accepted belief that selenium toxicity in plants is manifested by the misincorporation of selenocysteine into selenoproteins, there is a lack of data suggesting that selenoproteins are malformed or misfolded. Plant mechanisms to prevent the formation of selenoproteins are associated with increased selenium tolerance, yet there is no evidence to suggest that selenoproteins are malformed or potentially misfolded. We reasoned that if selenoproteins are malformed, then they might be degraded by the ubiquitin-proteasome pathway. The data demonstrate that selenate treatment induced the accumulation of both oxidized and ubiquitinated proteins, thus implicating both the 20S and 26S proteasome of Stanleya pinnata, a selenium-hyperaccumulating plant, in a selenate response. Inhibition of the proteasome increases the amount of selenium incorporated into protein, but not other elements. Furthermore, a higher percentage of selenium was found in a ubiquitinated protein fraction compared with other elements, suggesting that malformed selenoproteins are preferentially ubiquitinated and removed by the proteasome. Additionally, levels of the 20S and 26S proteasome and two heat shock proteins increase upon selenate treatment. Arabidopsis mutants with defects in the 26S proteasome have decreased selenium tolerance, which further supports the hypothesis that the 26S proteasome probably prevents selenium toxicity by removing selenoproteins. PMID:22323770

Sabbagh, Melissa; Van Hoewyk, Doug

2012-03-01

27

Space shuttle flight (STS90) enhances degradation of rat myosin heavy chain in association with activation of ubiquitin-proteasome pathway  

Microsoft Academic Search

To elucidate the mechanisms of microgravity-induced muscle atrophy, we focused on fast-type myosin heavy chain (MHC) degradation and expression of proteases in atrophied gastrocnemius muscles of neonatal rats exposed to 16-d spaceflight (STS-90). The spaceflight stimulated ubiquitination of proteins, including a MHC molecule, and accumulation of MHC degradation fragments in the muscles. Semiquantitative reverse transcriptase-polymerase chain reaction revealed that the

Madoka Ikemoto; Takeshi Nikawa; Shin' ichi Takeda; Chiho Watanabe; Takako Kitano; Kenneth M. Baldwin; Ryutaro Izumi; Ikuya Nonaka; Takae Towatari; Shigetada Teshima; Kazuhito Rokutan; Kyoichi Kishi

2001-01-01

28

Enzymatic Blockade of the Ubiquitin-Proteasome Pathway  

E-print Network

Ubiquitin-dependent processes control much of cellular physiology. We show that expression of a highly active, Epstein-Barr virus-derived deubiquitylating enzyme (EBV-DUB) blocks proteasomal degradation of cytosolic and ...

Ernst, Robert

29

Involvement of the ubiquitin\\/proteasome pathway in the organisation and polarised growth of kiwifruit pollen tubes  

Microsoft Academic Search

We recently reported the involvement of the ubiquitin pathway in microgametophyte development, and a direct role for the 26S proteasome in regulating pollen tube emergence in kiwifruit. Here we show that the ubiquitin\\/proteasome proteolytic pathway is involved not only in early kiwifruit pollen tube organisation, but also in maintaining polarised growth of tubes. By immunofluorescence analysis we show that ubiquitin

V. Scoccianti; E. Ovidi; A. R. Taddei; A. Tiezzi; R. Crinelli; L. Gentilini; A. Speranza

2003-01-01

30

Targeting Tumor Ubiquitin-Proteasome Pathway with Polyphenols for Chemosensitization  

PubMed Central

The development of tumor drug resistance is one of the biggest obstacles on the way to achieve a favorable outcome of chemotherapy. Among various strategies that have been explored to overcome drug resistance, the combination of current chemotherapy with plant polyphenols as a chemosensitizer has emerged as a promising one. Plant polyphenols are a group of phytochemicals characterized by the presence of more than one phenolic group. Mechanistic studies suggest that polyphenols have multiple intracellular targets, one of which is the proteasome complex. The proteasome is a proteolytic enzyme complex responsible for intracellular protein degradation and has been shown to play an important role in tumor growth and the development of drug resistance. Therefore, proteasome inhibition by plant polyphenols could be one of the mechanisms contributing to their chemosensitizing effect. Plant polyphenols that have been identified to possess proteasome-inhibitory activity include (?)-epigallocatechins-3-gallate (EGCG), genistein, luteolin, apigenin, chrysin, quercetin, curcumin and tannic acid. These polyphenols have exhibited an appreciable effect on overcoming resistance to various chemotherapeutic drugs as well as multidrug resistance in a broad spectrum of tumors ranging from carcinoma and sarcoma to hematological malignances. The in vitro and in vivo studies on polyphenols with proteasome-inhibitory activity have built a solid foundation to support the idea that they could serve as a chemosensitizer for the treatment of cancer. In-depth mechanistic studies and identification of optimal regimen are needed in order to eventually translate this laboratory concept into clinical trials to actually benefit current chemotherapy. PMID:22292765

Shen, Min; Chan, Tak Hang; Dou, Q. Ping

2012-01-01

31

Ubiquitin-proteasome dependent degradation of GABAA?1 in autism spectrum disorder  

PubMed Central

Background Although the neurobiological basis of autism spectrum disorder (ASD) is not fully understood, recent studies have indicated the potential role of GABAA receptors in the pathophysiology of ASD. GABAA receptors play a crucial role in various neurodevelopmental processes and adult neuroplasticity. However, the mechanism(s) of regulation of GABAA receptors in ASD remains poorly understood. Methods Postmortem middle frontal gyrus tissues (13 ASD and 13 control subjects) were used. In vitro studies were performed in primary cortical neurons at days in vitro (DIV) 14. The protein levels were examined by western blotting. Immunofluorescence studies were employed for cellular localization. The gene expression was determined by RT-PCR array and qRT-PCR. Results A significant decrease in GABAA?1 protein, but not mRNA levels was found in the middle frontal gyrus of ASD subjects indicating a post-translational regulation of GABAA receptors in ASD. At the cellular level, treatment with proteasomal inhibitor, MG132, or lactacystin significantly increased GABAA?1 protein levels and Lys48-linked polyubiquitination of GABAA?1, but reduced proteasome activity in mouse primary cortical neurons (DIV 14 from E16 embryos). Moreover, treatment with betulinic acid, a proteasome activator significantly decreased GABAA?1 protein levels in cortical neurons indicating the role of polyubiquitination of GABAA?1 proteins with their subsequent proteasomal degradation in cortical neurons. Ubiquitination specific RT-PCR array followed by western blot analysis revealed a significant increase in SYVN1, an endoplasmic reticulum (ER)-associated degradation (ERAD) E3 ubiquitin ligase in the middle frontal gyrus of ASD subjects. In addition, the inhibition of proteasomal activity by MG132 increased the expression of GABAA?1 in the ER. The siRNA knockdown of SYVN1 significantly increased GABAA?1 protein levels in cortical neurons. Moreover, reduced association between SYVN1 and GABAA?1 was found in the middle frontal gyrus of ASD subjects. Conclusions SYVN1 plays a critical role as an E3 ligase in the ubiquitin proteasome system (UPS)-mediated GABAA?1 degradation. Thus, inhibition of the ubiquitin-proteasome-mediated GABAA?1 degradation may be an important mechanism for preventing GABAA?1 turnover to maintain GABAA?1 levels and GABA signaling in ASD. PMID:25392730

2014-01-01

32

Involvement of Protein Degradation by the Ubiquitin Proteasome System in Opiate Addictive Behaviors  

PubMed Central

Plastic changes in the nucleus accumbens (NAcc), a structure occupying a key position in the neural circuitry related to motivation, are among the critical cellular processes responsible for drug addiction. During the last decade, it has been shown that memory formation and related neuronal plasticity may rely not only on protein synthesis but also on protein degradation by the ubiquitin proteasome system (UPS). In this study, we assess the role of protein degradation in the NAcc in opiate-related behaviors. For this purpose, we coupled behavioral experiments to intra-accumbens injections of lactacystin, an inhibitor of the UPS. We show that protein degradation in the NAcc is mandatory for a full range of animal models of opiate addiction including morphine locomotor sensitization, morphine conditioned place preference, intra-ventral tegmental area morphine self-administration and intra-venous heroin self-administration but not for discrimination learning rewarded by highly palatable food. This study provides the first evidence of a specific role of protein degradation by the UPS in addiction. PMID:23169349

Massaly, Nicolas; Dahan, Lionel; Baudonnat, Mathieu; Hovnanian, Caroline; Rekik, Khaoula; Solinas, Marcello; David, Vincent; Pech, Stéphane; Zajac, Jean-Marie; Roullet, Pascal; Mouledous, Lionel; Frances, Bernard

2013-01-01

33

Involvement of protein degradation by the ubiquitin proteasome system in opiate addictive behaviors.  

PubMed

Plastic changes in the nucleus accumbens (NAcc), a structure occupying a key position in the neural circuitry related to motivation, are among the critical cellular processes responsible for drug addiction. During the last decade, it has been shown that memory formation and related neuronal plasticity may rely not only on protein synthesis but also on protein degradation by the ubiquitin proteasome system (UPS). In this study, we assess the role of protein degradation in the NAcc in opiate-related behaviors. For this purpose, we coupled behavioral experiments to intra-accumbens injections of lactacystin, an inhibitor of the UPS. We show that protein degradation in the NAcc is mandatory for a full range of animal models of opiate addiction including morphine locomotor sensitization, morphine conditioned place preference, intra-ventral tegmental area morphine self-administration and intra-venous heroin self-administration but not for discrimination learning rewarded by highly palatable food. This study provides the first evidence of a specific role of protein degradation by the UPS in addiction. PMID:23169349

Massaly, Nicolas; Dahan, Lionel; Baudonnat, Mathieu; Hovnanian, Caroline; Rekik, Khaoula; Solinas, Marcello; David, Vincent; Pech, Stéphane; Zajac, Jean-Marie; Roullet, Pascal; Mouledous, Lionel; Frances, Bernard

2013-03-01

34

Muscle wasting in insulinopenic rats results from activation of the ATP-dependent, ubiquitin-proteasome proteolytic pathway by a mechanism including gene transcription.  

PubMed Central

In normal subjects and diabetic patients, insulin suppresses whole body proteolysis suggesting that the loss of lean body mass and muscle wasting in insulinopenia is related to increased muscle protein degradation. To document how insulinopenia affects organ weights and to identify the pathway for accelerated proteolysis in muscle, streptozotocin-treated and vehicle-injected, pair-fed control rats were studied. The weights of liver, adipose tissue, and muscle were decreased while muscle protein degradation was increased 75% by insulinopenia. This proteolytic response was not eliminated by blocking lysosomal function and calcium-dependent proteases at 7 or 3 d after streptozotocin. When ATP synthesis in muscle was inhibited, the rates of proteolysis were reduced to the same level in insulinopenic and control rats suggesting that the ATP-dependent, ubiquitin-proteasome pathway is activated. Additional evidence for activation of this pathway in muscle includes: (a) an inhibitor of proteasome activity eliminated the increased protein degradation; (b) mRNAs encoding ubiquitin and proteasome subunits were increased two- to threefold; and (c) there was increased transcription of the ubiquitin gene. We conclude that the mechanism for muscle protein wasting in insulinopenia includes activation of the ubiquitin-proteasome pathway with increased expression of the ubiquitin gene. PMID:8878419

Price, S R; Bailey, J L; Wang, X; Jurkovitz, C; England, B K; Ding, X; Phillips, L S; Mitch, W E

1996-01-01

35

Formation of distinct inclusion bodies by inhibition of ubiquitin-proteasome and autophagy-lysosome pathways  

SciTech Connect

Research highlights: {yields} Distinct inclusion bodies are developed by inhibition of UPP and ALP. {yields} The inclusion bodies differ in morphology, localization and formation process. {yields} The inclusion bodies are distinguishable by the localization of TSC2. {yields} Inhibition of both UPP and ALP simultaneously induces those inclusion bodies. -- Abstract: Accumulation of misfolded proteins is caused by the impairment of protein quality control systems, such as ubiquitin-proteasome pathway (UPP) and autophagy-lysosome pathway (ALP). In this study, the formation of inclusion bodies was examined after the blockade of UPP and/or ALP in A549 cells. UPP inhibition induced a single and large inclusion body localized in microtubule-organizing center. Interestingly, however, ALP inhibition generated dispersed small inclusion bodies in the cytoplasm. Tuberous sclerosis complex 2 was selectively accumulated in the inclusion bodies of UPP-inhibited cells, but not those of ALP-inhibited cells. Blockade of transcription and translation entirely inhibited the formation of inclusion body induced by UPP inhibition, but partially by ALP inhibition. Moreover, the simultaneous inhibition of two protein catabolic pathways independently developed two distinct inclusion bodies within a single cell. These findings clearly demonstrated that dysfunction of each catabolic pathway induced formation and accumulation of unique inclusion bodies on the basis of morphology, localization and formation process in A549 cells.

Lee, Junho; Yang, Kyu-Hwan; Joe, Cheol O. [Department of Biological Sciences, Korea Advanced Institute of Science and Technology, Daejon 305-701 (Korea, Republic of)] [Department of Biological Sciences, Korea Advanced Institute of Science and Technology, Daejon 305-701 (Korea, Republic of); Kang, Seok-Seong, E-mail: sskang@ivi.int [Laboratory Sciences Division, International Vaccine Institute, Seoul 151-919 (Korea, Republic of)] [Laboratory Sciences Division, International Vaccine Institute, Seoul 151-919 (Korea, Republic of)

2011-01-14

36

Activation of the ubiquitin proteasome pathway by silk fibroin modified chitosan nanoparticles in hepatic cancer cells.  

PubMed

Silk fibroin (SF) is a protein with bulky hydrophobic domains and can be easily purified as sericin-free silk-based biomaterial. Silk fibroin modified chitosan nanoparticle (SF-CSNP), a biocompatible material, has been widely used as a potential drug delivery system. Our current investigation studied the bio-effects of the SF-CSNP uptake by liver cells. In this experiment, the characterizations of SF-CSNPs were measured by particle size analysis and protein assay. The average size of the SF-CSNP was 311.9 ± 10.7 nm, and the average zeta potential was +13.33 ± 0.3 mV. The SF coating on the SF-CSNP was 6.27 ± 0.17 ?g/mL. Moreover, using proteomic approaches, several proteins involved in the ubiquitin proteasome pathway were identified by analysis of differential protein expressions of HepG2 cell uptake the SF-CSNP. Our experimental results have demonstrated that the SF-CSNP may be involved in liver cancer cell survival and proliferation. PMID:25588218

Yang, Ming-Hui; Chung, Tze-Wen; Lu, Yi-Shan; Chen, Yi-Ling; Tsai, Wan-Chi; Jong, Shiang-Bin; Yuan, Shyng-Shiou; Liao, Pao-Chi; Lin, Po-Chiao; Tyan, Yu-Chang

2014-01-01

37

Activation of the Ubiquitin Proteasome Pathway by Silk Fibroin Modified Chitosan Nanoparticles in Hepatic Cancer Cells  

PubMed Central

Silk fibroin (SF) is a protein with bulky hydrophobic domains and can be easily purified as sericin-free silk-based biomaterial. Silk fibroin modified chitosan nanoparticle (SF-CSNP), a biocompatible material, has been widely used as a potential drug delivery system. Our current investigation studied the bio-effects of the SF-CSNP uptake by liver cells. In this experiment, the characterizations of SF-CSNPs were measured by particle size analysis and protein assay. The average size of the SF-CSNP was 311.9 ± 10.7 nm, and the average zeta potential was +13.33 ± 0.3 mV. The SF coating on the SF-CSNP was 6.27 ± 0.17 ?g/mL. Moreover, using proteomic approaches, several proteins involved in the ubiquitin proteasome pathway were identified by analysis of differential protein expressions of HepG2 cell uptake the SF-CSNP. Our experimental results have demonstrated that the SF-CSNP may be involved in liver cancer cell survival and proliferation. PMID:25588218

Yang, Ming-Hui; Chung, Tze-Wen; Lu, Yi-Shan; Chen, Yi-Ling; Tsai, Wan-Chi; Jong, Shiang-Bin; Yuan, Shyng-Shiou; Liao, Pao-Chi; Lin, Po-Chiao; Tyan, Yu-Chang

2015-01-01

38

The acidosis of chronic renal failure activates muscle proteolysis in rats by augmenting transcription of genes encoding proteins of the ATP-dependent ubiquitin-proteasome pathway.  

PubMed Central

Chronic renal failure (CRF) is associated with negative nitrogen balance and loss of lean body mass. To identify specific proteolytic pathways activated by CRF, protein degradation was measured in incubated epitrochlearis muscles from CRF and sham-operated, pair-fed rats. CRF stimulated muscle proteolysis, and inhibition of lysosomal and calcium-activated proteases did not eliminate this increase. When ATP production was blocked, proteolysis in CRF muscles fell to the same level as that in control muscles. Increased proteolysis was also prevented by feeding CRF rats sodium bicarbonate, suggesting that activation depends on acidification. Evidence that the ATP-dependent ubiquitin-proteasome pathway is stimulated by the acidemia of CRF includes the following findings: (a) An inhibitor of the proteasome eliminated the increase in muscle proteolysis; and (b) there was an increase in mRNAs encoding ubiquitin (324%) and proteasome subunits C3 (137%) and C9 (251%) in muscle. This response involved gene activation since transcription of mRNAs for ubiquitin and the C3 subunit were selectively increased in muscle of CRF rats. We conclude that CRF stimulates muscle proteolysis by activating the ATP-ubiquitin-proteasome-dependent pathway. The mechanism depends on acidification and increased expression of genes encoding components of the system. These responses could contribute to the loss of muscle mass associated with CRF. PMID:8617877

Bailey, J L; Wang, X; England, B K; Price, S R; Ding, X; Mitch, W E

1996-01-01

39

Activation of the Ubiquitin Proteasome Pathway in a Mouse Model of Inflammatory Myopathy  

PubMed Central

Objective Myositis is characterized by severe muscle weakness. We and others have previously shown that endoplasmic reticulum (ER) stress plays a role in the pathogenesis of myositis. The present study was undertaken to identify perturbed pathways and assess their contribution to muscle disease in a mouse myositis model. Methods Stable isotope labeling with amino acids in cell culture (SILAC) was used to identify alterations in the skeletal muscle proteome of myositic mice in vivo. Differentially altered protein levels identified in the initial comparisons were validated using a liquid chromatography tandem mass spectrometry spike-in strategy and further confirmed by immunoblotting. In addition, we evaluated the effect of a proteasome inhibitor, bortezomib, on the disease phenotype, using well-standardized functional, histologic, and biochemical assessments. Results With the SILAC technique we identified significant alterations in levels of proteins belonging to the ER stress response, ubiquitin proteasome pathway (UPP), oxidative phosphorylation, glycolysis, cytoskeleton, and muscle contractile apparatus categories. We validated the myositis-related changes in the UPP and demonstrated a significant increase in the ubiquitination of muscle proteins as well as a specific increase in ubiquitin carboxyl-terminal hydrolase isozyme L1 (UCHL-1) in myositis, but not in muscle affected by other dystrophies or normal muscle. Inhibition of the UPP with bortezomib significantly improved muscle function and also significantly reduced tumor necrosis factor ? expression in the skeletal muscle of mice with myositis. Conclusion Our findings indicate that ER stress activates downstream UPPs and contributes to muscle degeneration and that UCHL-1 is a potential biomarker for disease progression. UPP inhibition offers a potential therapeutic strategy for myositis. PMID:24022788

Rayavarapu, Sree; Coley, William; der Meulen, Jack H. Van; Cakir, Erdinc; Tappeta, Kathyayini; Kinder, Travis B.; Dillingham, Blythe C.; Brown, Kristy J.; Hathout, Yetrib; Nagaraju, Kanneboyina

2014-01-01

40

Sepsis is associated with increased mRNAs of the ubiquitin-proteasome proteolytic pathway in human skeletal muscle.  

PubMed Central

Previous studies provided evidence that sepsis-induced muscle proteolysis in experimental animals is caused by increased ubiquitin-proteasome-dependent protein breakdown. It is not known if a similar mechanism accounts for muscle proteolysis in patients with sepsis. We determined mRNA levels for ubiquitin and the 20 S proteasome subunit HC3 by Northern blot analysis in muscle tissue from septic (n = 7) and non-septic (n = 11) patients. Plasma and muscle amino acid concentrations and concentrations in urine of 3-methylhistidine (3-MH), creatinine, and cortisol were measured at the time of surgery to assess the catabolic state of the patients. A three- to fourfold increase in mRNA levels for ubiquitin and HC3 was noted in muscle tissue from the septic patients concomitant with increased muscle levels of phenylalanine and 3-MH and reduced levels of glutamine. Total plasma amino acids were decreased by approximately 30% in the septic patients. The 3-MH/creatinine ratio in urine was almost doubled in septic patients. The cortisol levels in urine were higher in septic than in control patients but this difference did not reach statistical significance. The results suggest that sepsis is associated with increased mRNAs of the ubiquitin-proteasome pathway in human skeletal muscle. PMID:9005983

Tiao, G; Hobler, S; Wang, J J; Meyer, T A; Luchette, F A; Fischer, J E; Hasselgren, P O

1997-01-01

41

Dystrophin involved in the susceptibility of slow muscles to hindlimb unloading via concomitant activation of TGF-?1/Smad3 signaling and ubiquitin-proteasome degradation in mice.  

PubMed

While it is well known that the slow-twitch muscles are vulnerable to microgravity conditions, the molecular and cellular mechanisms underlying this phenomenon remain unknown. Dystrophin, which constitutes an important link between the cytoskeleton and the extracellular matrix, is hypothesized to be involved in force generation and mechanical stabilization of the skeletal muscle. Here we have shown that after a 14-day hindlimb unloading (HU) of the C57BL/10 mice, the expression of dystrophin was significantly down-regulated in the fast-twitch myofibers, while in the slow-twitch myofibers, it was up-regulated. In order to investigate the role of dystrophin in HU-induced susceptibility to muscle atrophy, we compared the degradation signaling mechanisms of slow-twitch soleus muscle in dystrophin-deficient (mdx) and the wild-type (WT) mice. We found that mdx mice manifest less reduction of muscle mass and myofiber cross-sectional area than the control animals. Also, the expression of two ubiquitin ligases (MuRF1, Atrogin-1), which plays a crucial role in the ubiquitin-proteasome-mediated muscular degradation, was significantly down-regulated in soleus muscle of the hindlimb-unloaded mdx mice. In comparison, in the soleus muscle of unloaded WT mice, these ligases were significantly up-regulated. Whereas the hindlimb unloading reduced the expression of transforming growth factor ? (TGF-?1)/Smad3 in mdx mice, in WT mice, the expression of this growth factor was augmented in response to unloading. Correspondingly, as a result of HU of the mdx mice, the expression of four subtypes of the myosin heavy chain and troponin I was reduced or it exhibited a delayed slow-to-fast transition. In summary, our results suggest that dystrophin exerts an intermediary and positive role in the disuse atrophy of the slow-twitch muscles. This effect is mediated through the activation of TGF-?1/Smad3 signaling and downstream ubiquitin-proteasome pathway. PMID:24839113

Zhang, Peng; Li, Wenjiong; Liu, Hongju; Li, Jinglong; Wang, Jing; Li, Yanan; Chen, Xiaoping; Yang, Zhong; Fan, Ming

2014-11-01

42

CDK11{sup p58} represses vitamin D receptor-mediated transcriptional activation through promoting its ubiquitin-proteasome degradation  

SciTech Connect

Vitamin D receptor (VDR) is a member of the nuclear receptor superfamily and regulates transcription of target genes. In this study, we identified CDK11{sup p58} as a novel protein involved in the regulation of VDR. CDK11{sup p58}, a member of the large family of p34cdc2-related kinases, is associated with cell cycle progression, tumorigenesis, and apoptotic signaling. Our study demonstrated that CDK11{sup p58} interacted with VDR and repressed VDR-dependent transcriptional activation. Furthermore, overexpression of CDK11{sup p58} decreased the stability of VDR through promoting its ubiquitin-proteasome-mediated degradation. Taken together, these results suggest that CDK11{sup p58} is involved in the negative regulation of VDR.

Chi, Yayun; Hong, Yi; Zong, Hongliang; Wang, Yanlin; Zou, Weiying; Yang, Junwu; Kong, Xiangfei; Yun, Xiaojing [Gene Research Center, Shanghai Medical College and Institutes of Biomedical, Shanghai 200032 (China)] [Gene Research Center, Shanghai Medical College and Institutes of Biomedical, Shanghai 200032 (China); Gu, Jianxin, E-mail: jxgu@shmu.edu.cn [Gene Research Center, Shanghai Medical College and Institutes of Biomedical, Shanghai 200032 (China)] [Gene Research Center, Shanghai Medical College and Institutes of Biomedical, Shanghai 200032 (China)

2009-08-28

43

Docosahexaenoic acid induces the degradation of HPV E6/E7 oncoproteins by activating the ubiquitin-proteasome system.  

PubMed

The oncogenic human papillomavirus (HPV) E6/E7 proteins are essential for the onset and maintenance of HPV-associated malignancies. Here, we report that activation of the cellular ubiquitin-proteasome system (UPS) by the omega-3 fatty acid, docosahexaenoic acid (DHA), leads to proteasome-mediated degradation of E6/E7 viral proteins and the induction of apoptosis in HPV-infected cancer cells. The increases in UPS activity and degradation of E6/E7 oncoproteins were associated with DHA-induced overproduction of mitochondrial reactive oxygen species (ROS). Exogenous oxidative stress and pharmacological induction of mitochondrial ROS showed effects similar to those of DHA, and inhibition of ROS production abolished UPS activation, E6/E7 viral protein destabilization, and apoptosis. These findings identify a novel role for DHA in the regulation of UPS and viral proteins, and provide evidence for the use of DHA as a mechanistically unique anticancer agent for the chemoprevention and treatment of HPV-associated tumors. PMID:25393480

Jing, K; Shin, S; Jeong, S; Kim, S; Song, K-S; Park, J-H; Heo, J-Y; Seo, K-S; Park, S-K; Kweon, G-R; Wu, T; Park, J-I; Lim, K

2014-01-01

44

AGEs-RAGE System Down-Regulates Sirt1 Through the Ubiquitin-Proteasome Pathway to Promote FN and TGF-?1 Expression in Male Rat Glomerular Mesangial Cells.  

PubMed

We previously demonstrated that advanced glycation-end products (AGEs) promote the pathological progression of diabetic nephropathy by decreasing silent information regulator 2-related protein 1 (Sirt1) expression in glomerular mesangial cells (GMCs). Here, we investigated whether AGEs-receptor for AGEs (RAGE) system down-regulated Sirt1 expression through ubiquitin-proteasome pathway and whether Sirt1 ubiquitination affected fibronectin (FN) and TGF-?1, 2 fibrotic indicators in GMCs. Sirt1 was polyubiquitinated and subsequently degraded by proteasome. AGEs increased Sirt1 ubiquitination and proteasome-mediated degradation, shortened Sirt1 half-life, and promoted FN and TGF-?1 expression. Ubiquitin-specific protease 22 (USP22) reduced Sirt1 ubiquitination and degradation and decreased FN and TGF-?1 expression in GMCs under both basal and AGEs-treated conditions. USP22 depletion enhanced Sirt1 degradation and displayed combined effects with AGEs to further promote FN and TGF-?1 expression. RAGE functioned crucial mediating roles in these processes via its C-terminal cytosolic domain. Inhibiting Sirt1 by EX-527 substantially suppressed the down-regulation of FN and TGF-?1 resulting from USP22 overexpression under both normal and AGEs-treated conditions, eventually leading to their up-regulation in GMCs. These results indicated that the AGEs-RAGE system increased the ubiquitination and subsequent proteasome-mediated degradation of Sirt1 by reducing USP22 level, and AGEs-RAGE-USP22-Sirt1 formed a cascade pathway that regulated FN and TGF-?1 level, which participated in the pathological progression of diabetic nephropathy. PMID:25375034

Huang, Kai-Peng; Chen, Cheng; Hao, Jie; Huang, Jun-Ying; Liu, Pei-Qing; Huang, He-Qing

2015-01-01

45

Neuroinflammation and J2 prostaglandins: linking impairment of the ubiquitin-proteasome pathway and mitochondria to neurodegeneration  

PubMed Central

The immune response of the CNS is a defense mechanism activated upon injury to initiate repair mechanisms while chronic over-activation of the CNS immune system (termed neuroinflammation) may exacerbate injury. The latter is implicated in a variety of neurological and neurodegenerative disorders such as Alzheimer and Parkinson diseases, amyotrophic lateral sclerosis, multiple sclerosis, traumatic brain injury, HIV dementia, and prion diseases. Cyclooxygenases (COX-1 and COX-2), which are key enzymes in the conversion of arachidonic acid into bioactive prostanoids, play a central role in the inflammatory cascade. J2 prostaglandins are endogenous toxic products of cyclooxygenases, and because their levels are significantly increased upon brain injury, they are actively involved in neuronal dysfunction induced by pro-inflammatory stimuli. In this review, we highlight the mechanisms by which J2 prostaglandins (1) exert their actions, (2) potentially contribute to the transition from acute to chronic inflammation and to the spreading of neuropathology, (3) disturb the ubiquitin-proteasome pathway and mitochondrial function, and (4) contribute to neurodegenerative disorders such as Alzheimer and Parkinson diseases, and amyotrophic lateral sclerosis, as well as stroke, traumatic brain injury (TBI), and demyelination in Krabbe disease. We conclude by discussing the therapeutic potential of targeting the J2 prostaglandin pathway to prevent/delay neurodegeneration associated with neuroinflammation. In this context, we suggest a shift from the traditional view that cyclooxygenases are the most appropriate targets to treat neuroinflammation, to the notion that J2 prostaglandin pathways and other neurotoxic prostaglandins downstream from cyclooxygenases, would offer significant benefits as more effective therapeutic targets to treat chronic neurodegenerative diseases, while minimizing adverse side effects. PMID:25628533

Figueiredo-Pereira, Maria E.; Rockwell, Patricia; Schmidt-Glenewinkel, Thomas; Serrano, Peter

2015-01-01

46

Regulation of HTLV-1 tax stability, cellular trafficking and NF-?B activation by the ubiquitin-proteasome pathway.  

PubMed

Human T-cell leukemia virus type 1 (HTLV-1) is a complex retrovirus that infects CD4+ T cells and causes adult T-cell leukemia/lymphoma (ATLL) in 3%-5% of infected individuals after a long latent period. HTLV-1 Tax is a trans-activating protein that regulates viral gene expression and also modulates cellular signaling pathways to enhance T-cell proliferation and cell survival. The Tax oncoprotein promotes T-cell transformation, in part via constitutive activation of the NF-?B transcription factor; however, the underlying mechanisms remain unknown. Ubiquitination is a type of post-translational modification that occurs in a three-step enzymatic cascade mediated by E1, E2 and E3 enzymes and regulates protein stability as well as signal transduction, protein trafficking and the DNA damage response. Emerging studies indicate that Tax hijacks the ubiquitin machinery to activate ubiquitin-dependent kinases and downstream NF-?B signaling. Tax interacts with the E2 conjugating enzyme Ubc13 and is conjugated on C-terminal lysine residues with lysine 63-linked polyubiquitin chains. Tax K63-linked polyubiquitination may serve as a platform for signaling complexes since this modification is critical for interactions with NEMO and IKK. In addition to NF-?B signaling, mono- and polyubiquitination of Tax also regulate its subcellular trafficking and stability. Here, we review recent advances in the diverse roles of ubiquitin in Tax function and how Tax usurps the ubiquitin-proteasome pathway to promote oncogenesis. PMID:25341660

Lavorgna, Alfonso; Harhaj, Edward William

2014-10-01

47

The Ubiquitin-Proteasome System Regulates Mitochondrial Intermembrane Space Proteins  

PubMed Central

Mitochondrial precursor proteins are synthesized in the cytosol and subsequently imported into mitochondria. The import of mitochondrial intermembrane space proteins is coupled with their oxidative folding and governed by the mitochondrial intermembrane space import and assembly (MIA) pathway. The cytosolic steps that precede mitochondrial import are not well understood. We identified a role for the ubiquitin-proteasome system in the biogenesis of intermembrane space proteins. Interestingly, the function of the ubiquitin-proteasome system is not restricted to conditions of mitochondrial protein import failure. The ubiquitin-proteasome system persistently removes a fraction of intermembrane space proteins under physiological conditions, acting as a negative regulator in the biogenesis of this class of proteins. Thus, the ubiquitin-proteasome system plays an important role in determining the levels of proteins targeted to the intermembrane space of mitochondria. PMID:23508107

Bragoszewski, Piotr; Gornicka, Agnieszka; Sztolsztener, Malgorzata E.

2013-01-01

48

Ubiquitin-Proteasome System as a Modulator of Cell Fate  

PubMed Central

Summary The ubiquitin/ proteasome system is the major non-lysosymal system for degrading proteins in the cell; the work leading to its discovery was awarded the Nobel Prize in Chemistry in 2004. In addition to small ubiquitin-like modifiers (e.g. Sumo and Nedd8), ubiquitin is involved in the complex regulation of the levels and function of many proteins and signaling pathways involved in determining cell fate. The cell death regulatory proteins, such as Bcl-2 family proteins and caspases are targeted for degradation by the ubiquitin proteasome system (UPS). In addition to mediating the degradation of proteins, the UPS regulates function and translocation of proteins, many of which play a role in the determination of cell fate. For example the UPS can regulate the activity of transcription factors, such as P53, NF-?B and HIF-1?, which control the expression of protein mediators of cell death. Aberrant UPS function has been reported in multiple neuropathologies including Parkinson's diseases and ischemia. With the number of ubiquitin conjugating and de-conjugating enzymes reaching close to the levels of protein kinases and phosphatases, it is clear that ubiquitination is an important biological regulatory step for proteins. PMID:17981502

Thompson, Simon; Loftus, Liam; Ashley, Michelle; Meller, Robert

2008-01-01

49

Pexophagy-linked degradation of the peroxisomal membrane protein Pex3p involves the ubiquitin-proteasome system.  

PubMed

Peroxisome autophagy, also known as pexophagy, describes the wholesale degradation of peroxisomes via the vacuole, when organelles become damaged or redundant. In the methylotrophic yeast Hansenula polymorpha, pexophagy is stimulated when cells growing on methanol are exposed to excess glucose. Degradation of the peroxisomal membrane protein Pex3p, a process that does not involve the vacuole, was shown to trigger pexophagy. In this contribution, we have characterised pexophagy-associated Pex3p degradation further. We show that Pex3p breakdown depends on ubiquitin and confirm that Pex3p is a target for ubiquitination. Furthermore, we identify a role for the peroxisomal E3 ligases Pex2p and Pex10p in Pex3p degradation, suggesting the existence of a ubiquitin-dependent pathway involved in removing proteins from the peroxisomal membrane. PMID:23899522

Williams, Chris; van der Klei, Ida J

2013-08-23

50

The malin-laforin complex suppresses the cellular toxicity of misfolded proteins by promoting their degradation through the ubiquitin-proteasome system.  

PubMed

Lafora disease (LD), a progressive form of inherited epilepsy, is associated with widespread neurodegeneration and the formation of polyglucosan bodies in the neurons. Laforin, a protein phosphatase, and malin, an E3 ubiquitin ligase, are two of the proteins that are defective in LD. We have shown recently that laforin and malin (referred together as LD proteins) are recruited to aggresome upon proteasomal blockade, possibly to clear misfolded proteins through the ubiquitin-proteasome system (UPS). Here we test this possibility using a variety of cytotoxic misfolded proteins, including the expanded polyglutamine protein, as potential substrates. Laforin and malin, together with Hsp70 as a functional complex, suppress the cellular toxicity of misfolded proteins, and all the three members of this complex are required for this function. Laforin and malin interact with misfolded proteins and promote their degradation through the UPS. LD proteins are recruited to the polyglutamine aggregates and reduce the frequency of aggregate-positive cells. Taken together, our results suggest that the malin-laforin complex is a novel player in the neuronal response to misfolded proteins and could be potential therapeutic targets for neurodegenerative disorders associated with cytotoxic proteins. PMID:19036738

Garyali, Punitee; Siwach, Pratibha; Singh, Pankaj Kumar; Puri, Rajat; Mittal, Shuchi; Sengupta, Sonali; Parihar, Rashmi; Ganesh, Subramaniam

2009-02-15

51

Phosphorylation of serine 188 protects RhoA from ubiquitin/proteasome-mediated degradation in vascular smooth muscle cells.  

PubMed

cAMP and cyclic GMP-dependent kinases (PKA and PKG) phosphorylate the small G protein RhoA on Ser188. We have previously demonstrated that phosphorylation of Ser188 inhibits RhoA-dependent functions and positively regulates RhoA expression, and that the nitric oxide (NO)/cGMP-dependent protein kinase pathway plays an essential role, both in vitro and in vivo, in the regulation of RhoA protein expression and functions in vascular smooth muscle cells. Here we analyze the consequences of Ser188 phosphorylation on RhoA protein degradation. By expressing Ser188 phosphomimetic wild-type (WT-RhoA-S188E) and active RhoA proteins (Q63L-RhoA-S188E), we show that phosphorylation of Ser188 of RhoA protects RhoA, particularly its active form, from ubiquitin-mediated proteasomal degradation. Coimmunoprecipitation experiments indicate that the resistance of the phosphorylated active form of RhoA to proteasome-mediated degradation is because of its cytoplasmic sequestration through enhanced RhoGDI interaction. In rat aortic smooth muscle cells, stimulation of PKG and inhibition of proteasome by lactacystin, induce nonadditive increases in RhoA protein expression. In addition, stimulation of PKG leads to the accumulation of GTP-bound RhoA in the cytoplasm. In vivo stimulation of the NO/PKG signaling by treating rats with sildenafil increased RhoA level and RhoA phosphorylation, and enhanced its association to RhoGDI in the pulmonary artery, whereas opposite effects are induced by chronic inhibition of NO synthesis in N-omega-nitro-L-arginine-treated rats. Our results thus suggest that Ser188 phosphorylation-mediated protection against degradation is a physiological process regulating the level of endogenous RhoA and define a novel function for RhoGDI, as an inhibitor of Rho protein degradation. PMID:15890975

Rolli-Derkinderen, Malvyne; Sauzeau, Vincent; Boyer, Laurent; Lemichez, Emmanuel; Baron, Céline; Henrion, Daniel; Loirand, Gervaise; Pacaud, Pierre

2005-06-10

52

Sensitivity and protein turnover response to glucocorticoids are different in skeletal muscle from adult and old rats. Lack of regulation of the ubiquitin-proteasome proteolytic pathway in aging.  

PubMed Central

We studied glucocorticoid-induced muscle wasting and subsequent recovery in adult (7-mo-old) and old (22-mo-old) rats, since the increased incidence of various disease states may result in glucocorticoids hypersecretion in aging. Adult and old rats received dexamethasone in their drinking water and were then allowed to recover. Muscle wasting occurred more rapidly in old rats and the recovery of muscle mass was impaired, suggesting that glucocorticoids may be involved in the emergence of muscle atrophy with advancing age. According to measurements in incubated epitrochlearis muscles, dexamethasone-induced muscle wasting mainly resulted from increased protein breakdown in the adult, but from depressed protein synthesis in the aged animal. Increased expression of cathepsin D, m-calpain, and ubiquitin was observed in the muscles from both dexamethasone-treated adult and old rats. By contrast, the disappearance of the stimulatory effect of glucocorticoids on protein break-down in aging occurred along with a loss of ability of steroids to enhance the expression of the 14-kD ubiquitin carrier protein E2, which is involved in protein substrates ubiquitinylation, and of subunits of the 20 S proteasome (the proteolytic core of the 26 S proteasome that degrades ubiquitin conjugates). Thus, if glucocorticoids play any role in the progressive muscle atrophy seen in aging, this is unlikely to result from an activation of the ubiquitin-proteasome proteolytic pathway. Images PMID:7593595

Dardevet, D; Sornet, C; Taillandier, D; Savary, I; Attaix, D; Grizard, J

1995-01-01

53

Activation of the ATP-ubiquitin-proteasome pathway in skeletal muscle of cachectic rats bearing a hepatoma  

NASA Technical Reports Server (NTRS)

Rats implanted with Yoshida ascites hepatoma (YAH) show a rapid and selective loss of muscle protein due mainly to a marked increase (63-95%) in the rate of protein degradation (compared with rates in muscles of pair-fed controls). To define which proteolytic pathways contribute to this increase, epitrochlearis muscles from YAH-bearing and control rats were incubated under conditions that modify different proteolytic systems. Overall proteolysis in either group of rats was not affected by removal of Ca2+ or by blocking the Ca(2+)-dependent proteolytic system. Inhibition of lysosomal function with methylamine reduced proteolysis (-12%) in muscles from YAH-bearing rats, but not in muscles of pair-fed rats. When ATP production was also inhibited, the remaining accelerated proteolysis in muscles of tumor-bearing rats fell to control levels. Muscles of YAH-bearing rats showed increased levels of ubiquitin-conjugated proteins and a 27-kDa proteasome subunit in Western blot analysis. Levels of mRNA encoding components of proteolytic systems were quantitated using Northern hybridization analysis. Although their total RNA content decreased 20-38%, pale muscles of YAH-bearing rats showed increased levels of ubiquitin mRNA (590-880%) and mRNA for multiple subunits of the proteasome (100-215%). Liver, kidney, heart, and brain showed no weight loss and no change in these mRNA species. Muscles of YAH-bearing rats also showed small increases (30-40%) in mRNA for cathepsins B and D, but not for calpain I or heat shock protein 70. Our findings suggest that accelerated muscle proteolysis and muscle wasting in tumor-bearing rats result primarily from activation of the ATP-dependent pathway involving ubiquitin and the proteasome.

Baracos, V. E.; DeVivo, C.; Hoyle, D. H.; Goldberg, A. L.

1995-01-01

54

Differential ubiquitin-proteasome and autophagy signaling following rotator cuff tears and suprascapular nerve injury  

PubMed Central

Previous studies have evaluated role of Akt/mTOR signaling in rotator cuff muscle atrophy and determined that there was differential in signaling following tendon transection (TT) and suprascapular nerve (SSN) denervation (DN), suggesting that atrophy following TT and DN was modulated by different protein degradation pathways. In this study, two muscle proteolytic systems that have been shown to be potent regulators of muscle atrophy in other injury models, the ubiquitin-proteasome pathway and autophagy, were evaluated following TT and DN. In addition to examining protein degradation, this study assessed protein synthesis rate following these two surgical models to understand how the balance between protein degradation and synthesis results in atrophy following rotator cuff injury. In contrast to the traditional theory that protein synthesis is decreased during muscle atrophy, this study suggests that protein synthesis is up-regulated in rotator cuff muscle atrophy following both surgical models. While the ubiquitin-proteasome pathway was a major contributor to the atrophy seen following DN, autophagy was a major contributor following TT. The findings of this study suggest that protein degradation is the primary factor contributing to atrophy following rotator cuff injury. However, different proteolytic pathways are activated if SSN injury is involved. PMID:24018537

Joshi, Sunil K.; Kim, Hubert T.; Feeley, Brian T.; Liu, Xuhui

2013-01-01

55

Apoptosis in Schwann Cell Cultures Is Closely Interrelated with the Activity of the Ubiquitin-Proteasome Proteolytic Pathway  

Microsoft Academic Search

Although the participation of the ubiquitin-dependent pathway and of the proteasome in apoptosis has been proposed, its role in this process is not yet clearly defined. In previous studies, we have shown that in the central nervous system of the rat, programmed cell death and the ubiquitin-dependent proteolytic pathway are closely related to each other and that different types of

L. A. Pasquini; M. Besio Moreno; M. Lopez Salon; E. F. Soto

2002-01-01

56

Substrate recognition in selective autophagy and the ubiquitin-proteasome system.  

PubMed

Dynamic protein turnover through regulated protein synthesis and degradation ensures cellular growth, proliferation, differentiation and adaptation. Eukaryotic cells utilize two mechanistically distinct but largely complementary systems - the 26S proteasome and the lysosome (or vacuole in yeast and plants) - to effectively target a wide range of proteins for degradation. The concerted action of the ubiquitination machinery and the 26S proteasome ensures the targeted and tightly regulated degradation of a subset of commonly short-lived cellular proteins. Autophagy is a distinct degradation pathway, which transports a highly heterogeneous set of cargos in dedicated vesicles, called autophagosomes, to the lysosome. There the cargo becomes degraded and its molecular building blocks are recycled. While general autophagy randomly engulfs portions of the cytosol, selective autophagy employs dedicated cargo adaptors to specifically enrich the forming autophagosomes for a certain type of cargo as a response to various intra- or extracellular signals. Selective autophagy targets a wide range of cargos including long-lived proteins and protein complexes, organelles, protein aggregates and even intracellular microbes. In this review we summarize available data on cargo recognition mechanisms operating in selective autophagy and the ubiquitin-proteasome system (UPS), and emphasize their differences and common themes. Moreover, we derive general regulatory principles underlying cargo recognition in selective autophagy, and describe the system-wide crosstalk between these two cellular protein degradation systems. This article is part of a Special Issue entitled: Ubiquitin-Proteasome System. Guest Editors: Thomas Sommer and Dieter H. Wolf. PMID:23545414

Schreiber, Anne; Peter, Matthias

2014-01-01

57

Impairment of the Ubiquitin-Proteasome Pathway by Methyl N-(6-Phenylsulfanyl-1H-benzimidazol-2-yl)carbamate Leads to a Potent Cytotoxic Effect in Tumor Cells  

PubMed Central

In recent years, there has been a great deal of interest in proteasome inhibitors as a novel class of anticancer drugs. We report that fenbendazole (FZ) (methyl N-(6-phenylsulfanyl-1H-benzimidazol-2-yl)carbamate) exhibits a potent growth-inhibitory activity against cancer cell lines but not normal cells. We show here, using fluorogenic substrates, that FZ treatment leads to the inhibition of proteasomal activity in the cells. Succinyl-Leu-Leu-Val-Tyr-methylcoumarinamide (MCA), benzyloxycarbonyl-Leu-Leu-Glu-7-amido-4-MCA, and t-butoxycarbonyl-Gln-Ala-Arg-7-amido-4-MCA fluorescent derivatives were used to assess chymotrypsin-like, post-glutamyl peptidyl-hydrolyzing, and trypsin-like protease activities, respectively. Non-small cell lung cancer cells transiently transfected with an expression plasmid encoding pd1EGFP and treated with FZ showed an accumulation of the green fluorescent protein in the cells due to an increase in its half-life. A number of apoptosis regulatory proteins that are normally degraded by the ubiquitin-proteasome pathway like cyclins, p53, and I?B? were found to be accumulated in FZ-treated cells. In addition, FZ induced distinct ER stress-associated genes like GRP78, GADD153, ATF3, IRE1?, and NOXA in these cells. Thus, treatment of human NSCLC cells with fenbendazole induced endoplasmic reticulum stress, reactive oxygen species production, decreased mitochondrial membrane potential, and cytochrome c release that eventually led to cancer cell death. This is the first report to demonstrate the inhibition of proteasome function and induction of endoplasmic reticulum stress/reactive oxygen species-dependent apoptosis in human lung cancer cell lines by fenbendazole, which may represent a new class of anticancer agents showing selective toxicity against cancer cells. PMID:22745125

Dogra, Nilambra; Mukhopadhyay, Tapas

2012-01-01

58

Impairment of the ubiquitin-proteasome pathway by methyl N-(6-phenylsulfanyl-1H-benzimidazol-2-yl)carbamate leads to a potent cytotoxic effect in tumor cells: a novel antiproliferative agent with a potential therapeutic implication.  

PubMed

In recent years, there has been a great deal of interest in proteasome inhibitors as a novel class of anticancer drugs. We report that fenbendazole (FZ) (methyl N-(6-phenylsulfanyl-1H-benzimidazol-2-yl)carbamate) exhibits a potent growth-inhibitory activity against cancer cell lines but not normal cells. We show here, using fluorogenic substrates, that FZ treatment leads to the inhibition of proteasomal activity in the cells. Succinyl-Leu-Leu-Val-Tyr-methylcoumarinamide (MCA), benzyloxycarbonyl-Leu-Leu-Glu-7-amido-4-MCA, and t-butoxycarbonyl-Gln-Ala-Arg-7-amido-4-MCA fluorescent derivatives were used to assess chymotrypsin-like, post-glutamyl peptidyl-hydrolyzing, and trypsin-like protease activities, respectively. Non-small cell lung cancer cells transiently transfected with an expression plasmid encoding pd1EGFP and treated with FZ showed an accumulation of the green fluorescent protein in the cells due to an increase in its half-life. A number of apoptosis regulatory proteins that are normally degraded by the ubiquitin-proteasome pathway like cyclins, p53, and I?B? were found to be accumulated in FZ-treated cells. In addition, FZ induced distinct ER stress-associated genes like GRP78, GADD153, ATF3, IRE1?, and NOXA in these cells. Thus, treatment of human NSCLC cells with fenbendazole induced endoplasmic reticulum stress, reactive oxygen species production, decreased mitochondrial membrane potential, and cytochrome c release that eventually led to cancer cell death. This is the first report to demonstrate the inhibition of proteasome function and induction of endoplasmic reticulum stress/reactive oxygen species-dependent apoptosis in human lung cancer cell lines by fenbendazole, which may represent a new class of anticancer agents showing selective toxicity against cancer cells. PMID:22745125

Dogra, Nilambra; Mukhopadhyay, Tapas

2012-08-31

59

Potent activity of carfilzomib, a novel, irreversible inhibitor of the ubiquitin-proteasome pathway, against preclinical models of multiple myeloma  

PubMed Central

The proteasome has emerged as an important target for cancer therapy with the approval of bortezomib, a first-in-class, reversible proteasome inhibitor, for relapsed/refractory multiple myeloma (MM). However, many patients have disease that does not respond to bortezomib, whereas others develop resistance, suggesting the need for other inhibitors with enhanced activity. We therefore evaluated a novel, irreversible, epoxomicin-related proteasome inhibitor, carfilzomib. In models of MM, this agent potently bound and specifically inhibited the chymotrypsin-like proteasome and immunoproteasome activities, resulting in accumulation of ubiquitinated substrates. Carfilzomib induced a dose- and time-dependent inhibition of proliferation, ultimately leading to apoptosis. Programmed cell death was associated with activation of c-Jun-N-terminal kinase, mitochondrial membrane depolarization, release of cytochrome c, and activation of both intrinsic and extrinsic caspase pathways. This agent also inhibited proliferation and activated apoptosis in patient-derived MM cells and neoplastic cells from patients with other hematologic malignancies. Importantly, carfilzomib showed increased efficacy compared with bortezomib and was active against bortezomib-resistant MM cell lines and samples from patients with clinical bortezomib resistance. Carfilzomib also overcame resistance to other conventional agents and acted synergistically with dexamethasone to enhance cell death. Taken together, these data provide a rationale for the clinical evaluation of carfilzomib in MM. PMID:17591945

Kuhn, Deborah J.; Chen, Qing; Voorhees, Peter M.; Strader, John S.; Shenk, Kevin D.; Sun, Congcong M.; Demo, Susan D.; Bennett, Mark K.; van Leeuwen, Fijs W. B.; Chanan-Khan, Asher A.

2007-01-01

60

The ubiquitin proteasome system in Caenorhabditis elegans and its regulation?  

PubMed Central

Protein degradation constitutes a major cellular function that is responsible for maintenance of the normal cellular physiology either through the degradation of normal proteins or through the elimination of damaged proteins. The Ubiquitin–Proteasome System (UPS)1 is one of the main proteolytic systems that orchestrate protein degradation. Given that up- and down- regulation of the UPS system has been shown to occur in various normal (such as ageing) and pathological (such as neurodegenerative diseases) processes, the exogenous modulation of the UPS function and activity holds promise of (a) developing new therapeutic interventions against various diseases and (b) establishing strategies to maintain cellular homeostasis. Since the proteasome genes are evolutionarily conserved, their role can be dissected in simple model organisms, such as the nematode, Caenorhabditis elegans. In this review, we survey findings on the redox regulation of the UPS in C. elegans showing that the nematode is an instrumental tool in the identification of major players in the UPS pathway. Moreover, we specifically discuss UPS-related genes that have been modulated in the nematode and in human cells and have resulted in similar effects thus further exhibiting the value of this model in the study of the UPS. PMID:24563851

Papaevgeniou, Nikoletta; Chondrogianni, Niki

2014-01-01

61

Sit4 phosphatase is functionally linked to the ubiquitin-proteasome system.  

PubMed Central

Using a synthetic lethality screen we found that the Sit4 phosphatase is functionally linked to the ubiquitin-proteasome system. Yeast cells harboring sit4 mutations and an impaired proteasome (due to pre1-1 pre4-1 mutations) exhibited defective growth on minimal medium. Nearly identical synthetic effects were found when sit4 mutations were combined with defects of the Rad6/Ubc2- and Cdc34/Ubc3-dependent ubiquitination pathways. Under synthetic lethal conditions, sit4 pre or sit4 ubc mutants formed strongly enlarged unbudded cells with a DNA content of 1N, indicating a defect in the maintenance of cell integrity during starvation-induced G(1) arrest. Sit4-related synthetic effects could be cured by high osmotic pressure or by the addition of certain amino acids to the growth medium. These results suggest a concerted function of the Sit4 phosphatase and the ubiquitin-proteasome system in osmoregulation and in the sensing of nutrients. Further analysis showed that Sit4 is not a target of proteasome-dependent protein degradation. We could also show that Sit4 does not contribute to regulation of proteasome activity. These data suggest that both Sit4 phosphatase and the proteasome act on a common target protein. PMID:12930741

Singer, Thorsten; Haefner, Stefan; Hoffmann, Michael; Fischer, Michael; Ilyina, Julia; Hilt, Wolfgang

2003-01-01

62

Local ubiquitin-proteasome-mediated proteolysis and long-term synaptic plasticity  

PubMed Central

The ubiquitin-proteasome pathway (UPP) of protein degradation has many roles in synaptic plasticity that underlies memory. Work on both invertebrate and vertebrate model systems has shown that the UPP regulates numerous substrates critical for synaptic plasticity. Initial research took a global view of ubiquitin-protein degradation in neurons. Subsequently, the idea of local protein degradation was proposed a decade ago. In this review, we focus on the functions of the UPP in long-term synaptic plasticity and discuss the accumulated evidence in support of the idea that the components of the UPP often have disparate local roles in different neuronal compartments rather than a single cell-wide function. PMID:25520617

Hegde, Ashok N.; Haynes, Kathryn A.; Bach, Svitlana V.

2014-01-01

63

Ubiquitin-proteasome system and mitochondria - reciprocity.  

PubMed

Recently, sporadic links have been published between mitochondria - membrane-confined organelles - and the cytosolic ubiquitin-proteasome system (UPS) for removal of cellular proteins. For example, Fzo1, a mitochondrial outer membrane mitofusin was shown to be ubiquitinated by a ubiquitin ligase, Cdc53(MDM30), and degraded by the proteasome. Two additional ubiquitin ligases, MITOL/MARCH-V and MULAN, as well as a deubiquitinating enzyme, Ubp16/USP30, are embedded in mitochondrial outer membranes and participate in mitochondrial dynamics. Defects in mitochondrial morphology or respiration capacity are also reported for mutations in other UPS components such as the Ub ligases Parkin and Rsp5 as well as in proteasome subunits. These examples are likely to reflect a pervasive involvement of UPS in recycling of mitochondria-associated proteins. The flux of imported proteins and the proximity to oxidative phosphorylation results in abundant damaged or misfolded proteins that generate the need for a responsive quality control system. Within the mitochondrial matrix there is a self-contained ATP-dependent system for protein turnover. However at the outer membrane, the UPS may play a corresponding role in recycling either membrane-embedded or imported proteins. In a parallel process, ubiquitination also partakes in selection of damaged mitochondria to the lysozome/vacuole via autophagy. In the reverse direction, components of the UPS are sensitive to cellular REDOX potential, and as such are affected by reactive oxygen species (ROS) generated as a byproduct of mitochondrial respiration. This review will try to address the regulation of mitochondrial morphology and metabolic function by UPS, as well as the reciprocal relationship between aberrant ROS produced by mitochondria and ubiquitination or proteasome activity. This article is part of a Special Issue entitled The 26S Proteasome: When degradation is just not enough! PMID:20674813

Livnat-Levanon, Nurit; Glickman, Michael H

2011-02-01

64

Role of the ubiquitin proteasome system in hematologic malignancies.  

PubMed

Ubiquitination is a post-translational modification process that regulates several critical cellular processes. Ubiquitination is orchestrated by the ubiquitin proteasome system (UPS), which constitutes a cascade of enzymes that transfer ubiquitin onto protein substrates. The UPS catalyzes the destruction of many critical protein substrates involved in cancer pathogenesis. This review article focuses on components of the UPS that have been demonstrated to be deregulated by a variety of mechanisms in hematologic malignancies. These include E3 ubiquitin ligases and deubiquitinating enzymes. The prospects of specific targeting of key enzymes in this pathway that are critical to the pathogenesis of particular hematologic neoplasia are also discussed. PMID:25510280

Sahasrabuddhe, Anagh A; Elenitoba-Johnson, Kojo S J

2015-01-01

65

Emerging therapies targeting the ubiquitin proteasome system in cancer  

PubMed Central

The ubiquitin proteasome system (UPS) is an essential metabolic constituent of cellular physiology that tightly regulates cellular protein concentrations with specificity and precision to optimize cellular function. Inhibition of the proteasome has proven very effective in the treatment of multiple myeloma, and this approach is being tested for utility in other malignancies. New pharmaceuticals targeting the proteasome itself or specific proximal pathways of the UPS are in development as antiproliferatives or immunomodulatory agents. In this article, we discuss the biology of UPS-targeting drugs, their use as therapy for neoplasia, and the state of clinical and preclinical development for emerging therapeutics. PMID:24382383

Weathington, Nathaniel M.; Mallampalli, Rama K.

2014-01-01

66

Roles and potential therapeutic targets of the ubiquitin proteasome system in muscle wasting  

PubMed Central

Muscle wasting, characterized by the loss of protein mass in myofibers, is in most cases largely due to the activation of intracellular protein degradation by the ubiquitin proteasome system (UPS). During the last decade, mechanisms contributing to this activation have been unraveled and key mediators of this process identified. Even though much remains to be understood, the available information already suggests screens for new compounds inhibiting these mechanisms and highlights the potential for pharmaceutical drugs able to treat muscle wasting when it becomes deleterious. This review presents an overview of the main pathways contributing to UPS activation in muscle and describes the present state of efforts made to develop new strategies aimed at blocking or slowing muscle wasting. Publication history: Republished from Current BioData's Targeted Proteins database (TPdb; ). PMID:18047744

Nury, David; Doucet, Christine; Coux, Olivier

2007-01-01

67

Transcription alterations of members of the ubiquitin–proteasome network in prostate carcinoma  

Microsoft Academic Search

The purpose of this work was to investigate the role of the ubiquitin–proteasome network (UPN) in prostate cancer (PCA) and to elicit potential markers for this disease. The UPN represents a key factor in the maintenance of cellular homoeostasis as a result of its fundamental function in the regulation of intracellular protein degradation. Members of this network have a role

O J C Hellwinkel; L E Asong; J-P Rogmann; H Sültmann; C Wagner; T Schlomm; C Eichelberg; OJC Hellwinkel

2011-01-01

68

Regulation of Akt-mTOR, ubiquitin-proteasome and autophagy-lysosome pathways in response to formoterol administration in rat skeletal muscle.  

PubMed

Administration of ?2-agonists triggers skeletal muscle anabolism and hypertrophy. We investigated the time course of the molecular events responsible for rat skeletal muscle hypertrophy in response to 1, 3 and 10 days of formoterol administration (i.p. 2000?g/kg/day). A marked hypertrophy of rat tibialis anterior muscle culminated at day 10. Phosphorylation of Akt, ribosomal protein S6, 4E-BP1 and ERK1/2 was increased at day 3, but returned to control level at day 10. This could lead to a transient increase in protein translation and could explain previous studies that reported increase in protein synthesis following ?2-agonist administration. Formoterol administration was also associated with a significant reduction in MAFbx/atrogin-1 mRNA level (day 3), suggesting that formoterol can also affect protein degradation of MAFbx/atrogin1 targeted substrates, including MyoD and eukaryotic initiation factor-3f (eIF3-f). Surprisingly, mRNA level of autophagy-related genes, light chain 3 beta (LC3b) and gamma-aminobutyric acid receptor-associated protein-like 1 (Gabarapl1), as well as lysosomal hydrolases, cathepsin B and cathepsin L, was significantly and transiently increased after 1 and/or 3 days, suggesting that autophagosome formation would be increased in response to formoterol administration. However, this has to be relativized since the mRNA level of Unc-51-like kinase1 (Ulk1), BCL2/adenovirus E1B interacting protein3 (Bnip3), and transcription factor EB (TFEB), as well as the protein content of Ulk1, Atg13, Atg5-Atg12 complex and p62/Sqstm1 remained unchanged or was even decreased in response to formoterol administration. These results demonstrate that the effects of formoterol are mediated, in part, through the activation of Akt-mTOR pathway and that other signaling pathways become more important in the regulation of skeletal muscle mass with chronic administration of ?2-agonists. PMID:23916784

Joassard, Olivier Roger; Amirouche, Adel; Gallot, Yann Simon; Desgeorges, Marine Maud; Castells, Josiane; Durieux, Anne-Cécile; Berthon, Phanélie; Freyssenet, Damien Gilles

2013-11-01

69

Back to your heart: Ubiquitin proteasome system-regulated signal transduction  

PubMed Central

Awareness of the regulation of cell signaling by post-translational ubiquitination has emerged over the past 2 decades. Like phosphorylation, post-translational modification of proteins with ubiquitin can result in the regulation of numerous cellular functions, for example, the DNA damage response, apoptosis, cell growth, and the innate immune response. In this review, we discuss recently published mechanisms by which the ubiquitin proteasome system regulates key signal transduction pathways in the heart, including MAPK JNK, calcineurin, FOXO, p53, and estrogen receptors ? and ?. We then explore how ubiquitin proteasome system-specific regulation of these signal transduction pathways plays a role in the pathophysiology of common cardiac diseases, such as cardiac hypertrophy, heart failure, ischemia reperfusion injury, and diabetes. PMID:22085703

Portbury, Andrea L.; Ronnebaum, Sarah M.; Zungu, Makhosazane; Patterson, Cam; Willis, Monte S.

2011-01-01

70

The ubiquitin–proteasome machinery is essential for nuclear translocation of incoming minute virus of mice  

Microsoft Academic Search

Minute virus of mice (MVM) infection is disrupted by proteasome inhibitors. Here, we show that inhibition of the ubiquitin–proteasome pathway did not affect viral entry and had influence neither on the natural proteolytic cleavage of VP2 to VP3 nor on the externalization of the N terminal of VP1. In both MG132-treated and untreated cells, MVM particles accumulated progressively in the

Carlos Ros; Christoph Kempf

2004-01-01

71

Intersecting pathways to neurodegeneration in Parkinson's disease: Effects of the pesticide rotenone on DJ1, ?-synuclein, and the ubiquitin–proteasome system  

Microsoft Academic Search

Sporadic Parkinson's disease (PD) is most likely caused by a combination of environmental exposures and genetic susceptibilities, although there are rare monogenic forms of the disease. Mitochondrial impairment at complex I, oxidative stress, ?-synuclein aggregation, and dysfunctional protein degradation, have been implicated in PD pathogenesis, but how they are related to each other is unclear. To further evaluated PD pathogenesis

Ranjita Betarbet; Rosa M. Canet-Aviles; Todd B. Sherer; Pier G. Mastroberardino; Chris McLendon; Jin-Ho Kim; Serena Lund; Hye-Mee Na; Georgia Taylor; Neil F. Bence; Ron Kopito; Byoung Boo Seo; Takao Yagi; Akemi Yagi; Gary Klinefelter; Mark R. Cookson; J. Timothy Greenamyre

2006-01-01

72

The Role of the Ubiquitin Proteasome System in Ischemia and Ischemic Tolerance  

PubMed Central

Ubiquitin modification targets a protein for rapid degradation by the proteasome. However, poly-ubiquitination of proteins can result in multiple functions depending on the topology of the ubiquitin chain. Therefore ubiquitin signaling offers a more complex and versatile biology compared to many other post translational modifications. One area of potential for the application of this knowledge is the field of ischemia-induced brain damage, as occurs following a stroke. The ubiquitin proteasome system may exert a dual role on neuronal outcome following ischemia. Harmful ischemia results in an overload of the ubiquitin proteasome system, and blocking the proteasome reduces brain infarction following ischemia. However, the rapid and selective degradation of proteins following brief ischemia results in endogenous protection against ischemia. Therefore further understanding of the molecular signaling mechanisms which regulate the ubiquitin proteasome system may reveal novel therapeutic targets to reduce brain damage when ischemia is predicted, or to reduce the activation of the cell death mechanisms and the inflammatory response following stroke. The aim of this review is to discuss some of the recent advances in the understanding of protein ubiquitination and its implications for novel stroke therapies. PMID:19181875

Meller, Robert

2010-01-01

73

Autophagy and the ubiquitin-proteasome system: collaborators in neuroprotection.  

PubMed

Protein degradation is an essential cellular function that, when dysregulated or impaired, can lead to a wide variety of disease states. The two major intracellular protein degradation systems are the ubiquitin-proteasome system (UPS) and autophagy, a catabolic process that involves delivery of cellular components to the lysosome for degradation. While the UPS has garnered much attention as it relates to neurodegenerative disease, important links between autophagy and neurodegeneration have also become evident. Furthermore, recent studies have revealed interaction between the UPS and autophagy, suggesting a coordinated and complementary relationship between these degradation systems that becomes critical in times of cellular stress. Here we describe autophagy and review evidence implicating this system as an important player in the pathogenesis of neurodegenerative disease. We discuss the role of autophagy in neurodegeneration and review its neuroprotective functions as revealed by experimental manipulation in disease models. Finally, we explore potential parallels and connections between autophagy and the UPS, highlighting their collaborative roles in protecting against neurodegenerative disease. PMID:18930136

Nedelsky, Natalia B; Todd, Peter K; Taylor, J Paul

2008-12-01

74

Differential roles of the ubiquitin proteasome system and autophagy in the clearance of soluble and aggregated TDP-43 species  

PubMed Central

ABSTRACT TAR DNA-binding protein (TDP-43, also known as TARDBP) is the major pathological protein in amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD). Large TDP-43 aggregates that are decorated with degradation adaptor proteins are seen in the cytoplasm of remaining neurons in ALS and FTD patients post mortem. TDP-43 accumulation and ALS-linked mutations within degradation pathways implicate failed TDP-43 clearance as a primary disease mechanism. Here, we report the differing roles of the ubiquitin proteasome system (UPS) and autophagy in the clearance of TDP-43. We have investigated the effects of inhibitors of the UPS and autophagy on the degradation, localisation and mobility of soluble and insoluble TDP-43. We find that soluble TDP-43 is degraded primarily by the UPS, whereas the clearance of aggregated TDP-43 requires autophagy. Cellular macroaggregates, which recapitulate many of the pathological features of the aggregates in patients, are reversible when both the UPS and autophagy are functional. Their clearance involves the autophagic removal of oligomeric TDP-43. We speculate that, in addition to an age-related decline in pathway activity, a second hit in either the UPS or the autophagy pathway drives the accumulation of TDP-43 in ALS and FTD. Therapies for clearing excess TDP-43 should therefore target a combination of these pathways. PMID:24424030

Scotter, Emma L.; Vance, Caroline; Nishimura, Agnes L.; Lee, Youn-Bok; Chen, Han-Jou; Urwin, Hazel; Sardone, Valentina; Mitchell, Jacqueline C.; Rogelj, Boris; Rubinsztein, David C.; Shaw, Christopher E.

2014-01-01

75

The ubiquitin-proteasome system in spongiform degenerative disorders  

PubMed Central

Summary Spongiform degeneration is characterized by vacuolation in nervous tissue accompanied by neuronal death and gliosis. Although spongiform degeneration is a hallmark of prion diseases, this pathology is also present in the brains of patients suffering from Alzheimer's disease, diffuse Lewy body disease, human immunodeficiency virus (HIV) infection, and Canavan's spongiform leukodystrophy. The shared outcome of spongiform degeneration in these diverse diseases suggests that common cellular mechanisms must underlie the processes of spongiform change and neurodegeneration in the central nervous system. Immunohistochemical analysis of brain tissues reveals increased ubiquitin immunoreactivity in and around areas of spongiform change, suggesting the involvement of ubiquitin-proteasome system dysfunction in the pathogenesis of spongiform neurodegeneration. The link between aberrant ubiquitination and spongiform neurodegeneration has been strengthened by the discovery that a null mutation in the E3 ubiquitin-protein ligase mahogunin ring finger-1 (Mgrn1) causes an autosomal recessively inherited form of spongiform neurodegeneration in animals. Recent studies have begun to suggest that abnormal ubiquitination may alter intracellular signaling and cell functions via proteasome-dependent and proteasome-independent mechanisms, leading to spongiform degeneration and neuronal cell death. Further elucidation of the pathogenic pathways involved in spongiform neurodegeneration should facilitate the development of novel rational therapies for treating prion diseases, HIV infection, and other spongiform degenerative disorders. PMID:18790052

Whatley, Brandi R.; Li, Lian; Chin, Lih-Shen

2008-01-01

76

Role of the ubiquitin proteasome system in Alzheimer's disease  

Microsoft Academic Search

Though Alzheimer's disease (AD) is a syndrome with well-defined clinical and neuropathological manifestations, an array of molecular defects underlies its pathology. A role for the ubiquitin proteasome system (UPS) was suspected in the pathogenesis of AD since the presence of ubiquitin immunoreactivity in AD-related neuronal inclusions, such as neurofibrillary tangles, is seen in all AD cases. Recent studies have indicated

Sudarshan C Upadhya; Ashok N Hegde

2007-01-01

77

The ubiquitin-proteasome system: opportunities for therapeutic intervention in solid tumors.  

PubMed

The destruction of proteins via the ubiquitin-proteasome system is a multi-step, complex process involving polyubiquitination of substrate proteins, followed by proteolytic degradation by the macromolecular 26S proteasome complex. Inhibitors of the proteasome promote the accumulation of proteins that are deleterious to cell survival, and represent promising anti-cancer agents. In multiple myeloma and mantle cell lymphoma, treatment with the first-generation proteasome inhibitor, bortezomib, or the second-generation inhibitor, carfilzomib, has demonstrated significant therapeutic benefit in humans. This has prompted United States Food and Drug Administration (US FDA) approval of these agents and development of additional second-generation compounds with improved properties. There is considerable interest in extending the benefits of proteasome inhibitors to the treatment of solid tumor malignancies. Herein, we review progress that has been made in the preclinical development and clinical evaluation of different proteasome inhibitors in solid tumors. In addition, we describe several novel approaches that are currently being pursued for the treatment of solid tumors, including drug combinatorial strategies incorporating proteasome inhibitors and the targeting of components of the ubiquitin-proteasome system that are distinct from the 26S proteasome complex. PMID:24659480

Johnson, Daniel E

2015-02-01

78

Regulation of mitochondrial genome inheritance by autophagy and ubiquitin-proteasome system: implications for health, fitness, and fertility.  

PubMed

Mitochondria, the energy-generating organelles, play a role in numerous cellular functions including adenosine triphosphate (ATP) production, cellular homeostasis, and apoptosis. Maternal inheritance of mitochondria and mitochondrial DNA (mtDNA) is universally observed in humans and most animals. In general, high levels of mitochondrial heteroplasmy might contribute to a detrimental effect on fitness and disease resistance. Therefore, a disposal of the sperm-derived mitochondria inside fertilized oocytes assures normal preimplantation embryo development. Here we summarize the current research and knowledge concerning the role of autophagic pathway and ubiquitin-proteasome-dependent proteolysis in sperm mitophagy in mammals, including humans. Current data indicate that sperm mitophagy inside the fertilized oocyte could occur along multiple degradation routes converging on autophagic clearance of paternal mitochondria. The influence of assisted reproductive therapies (ART) such as intracytoplasmic sperm injection (ICSI), mitochondrial replacement (MR), and assisted fertilization of oocytes from patients of advanced reproductive age on mitochondrial function, inheritance, and fitness and for the development and health of ART babies will be of particular interest to clinical audiences. Altogether, the study of sperm mitophagy after fertilization has implications in the timing of evolution and developmental and reproductive biology and in human health, fitness, and management of mitochondrial disease. PMID:25028670

Song, Won-Hee; Ballard, John William Oman; Yi, Young-Joo; Sutovsky, Peter

2014-01-01

79

Regulation of Mitochondrial Genome Inheritance by Autophagy and Ubiquitin-Proteasome System: Implications for Health, Fitness, and Fertility  

PubMed Central

Mitochondria, the energy-generating organelles, play a role in numerous cellular functions including adenosine triphosphate (ATP) production, cellular homeostasis, and apoptosis. Maternal inheritance of mitochondria and mitochondrial DNA (mtDNA) is universally observed in humans and most animals. In general, high levels of mitochondrial heteroplasmy might contribute to a detrimental effect on fitness and disease resistance. Therefore, a disposal of the sperm-derived mitochondria inside fertilized oocytes assures normal preimplantation embryo development. Here we summarize the current research and knowledge concerning the role of autophagic pathway and ubiquitin-proteasome-dependent proteolysis in sperm mitophagy in mammals, including humans. Current data indicate that sperm mitophagy inside the fertilized oocyte could occur along multiple degradation routes converging on autophagic clearance of paternal mitochondria. The influence of assisted reproductive therapies (ART) such as intracytoplasmic sperm injection (ICSI), mitochondrial replacement (MR), and assisted fertilization of oocytes from patients of advanced reproductive age on mitochondrial function, inheritance, and fitness and for the development and health of ART babies will be of particular interest to clinical audiences. Altogether, the study of sperm mitophagy after fertilization has implications in the timing of evolution and developmental and reproductive biology and in human health, fitness, and management of mitochondrial disease. PMID:25028670

Ballard, John William Oman; Yi, Young-Joo

2014-01-01

80

Monitoring of ubiquitin-proteasome activity in living cells using a Degron (dgn)-destabilized green fluorescent protein (GFP)-based reporter protein.  

PubMed

Proteasome is the main intracellular organelle involved in the proteolytic degradation of abnormal, misfolded, damaged or oxidized proteins (1, 2). Maintenance of proteasome activity was implicated in many key cellular processes, like cell's stress response (3), cell cycle regulation and cellular differentiation (4) or in immune system response (5). The dysfunction of the ubiquitin-proteasome system has been related to the development of tumors and neurodegenerative diseases (4, 6). Additionally, a decrease in proteasome activity was found as a feature of cellular senescence and organismal aging (7, 8, 9, 10). Here, we present a method to measure ubiquitin-proteasome activity in living cells using a GFP-dgn fusion protein. To be able to monitor ubiquitin-proteasome activity in living primary cells, complementary DNA constructs coding for a green fluorescent protein (GFP)-dgn fusion protein (GFP-dgn, unstable) and a variant carrying a frameshift mutation (GFP-dgnFS, stable (11)) are inserted in lentiviral expression vectors. We prefer this technique over traditional transfection techniques because it guarantees a very high transfection efficiency independent of the cell type or the age of the donor. The difference between fluorescence displayed by the GFP-dgnFS (stable) protein and the destabilized protein (GFP-dgn) in the absence or presence of proteasome inhibitor can be used to estimate ubiquitin-proteasome activity in each particular cell strain. These differences can be monitored by epifluorescence microscopy or can be measured by flow cytometry. PMID:23169445

Greussing, Ruth; Unterluggauer, Hermann; Koziel, Rafal; Maier, Andrea B; Jansen-Dürr, Pidder

2012-01-01

81

Roles of the ubiquitin proteasome system in the effects of drugs of abuse  

PubMed Central

Because of its ability to regulate the abundance of selected proteins the ubiquitin proteasome system (UPS) plays an important role in neuronal and synaptic plasticity. As a result various stages of learning and memory depend on UPS activity. Drug addiction, another phenomenon that relies on neuroplasticity, shares molecular substrates with memory processes. However, the necessity of proteasome-dependent protein degradation for the development of addiction has been poorly studied. Here we first review evidences from the literature that drugs of abuse regulate the expression and activity of the UPS system in the brain. We then provide a list of proteins which have been shown to be targeted to the proteasome following drug treatment and could thus be involved in neuronal adaptations underlying behaviors associated with drug use and abuse. Finally we describe the few studies that addressed the need for UPS-dependent protein degradation in animal models of addiction-related behaviors. PMID:25610367

Massaly, Nicolas; Francès, Bernard; Moulédous, Lionel

2015-01-01

82

Testosterone protects against dexamethasone-induced muscle atrophy, protein degradation and MAFbx upregulation  

Microsoft Academic Search

Administration of glucocorticoids in pharmacological amounts results in muscle atrophy due, in part, to accelerated degradation of muscle proteins by the ubiquitin–proteasome pathway. The ubiquitin ligase MAFbx is upregulated during muscle loss including that caused by glucocorticoids and has been implicated in accelerated muscle protein catabolism during such loss. Testosterone has been found to reverse glucocorticoid-induced muscle loss due to

Weidong Zhao; Jiangping Pan; Zingbo Zhao; Yong Wu; William A. Bauman; Christopher P. Cardozo

2008-01-01

83

Catabolite Degradation of Fructose-1,6-bisphosphatase in the Yeast Saccharomyces cerevisiae: A Genome-wide Screen Identifies Eight Novel GID Genes and Indicates the Existence of Two Degradation Pathways  

PubMed Central

Metabolic adaptation of Saccharomyces cerevisiae cells from a nonfermentable carbon source to glucose induces selective, rapid breakdown of the gluconeogenetic key enzyme fructose-1,6-bisphosphatase (FBPase), a process called catabolite degradation. Herein, we identify eight novel GID genes required for proteasome-dependent catabolite degradation of FBPase. Four yeast proteins contain the CTLH domain of unknown function. All of them are Gid proteins. The site of catabolite degradation has been controversial until now. Two FBPase degradation pathways have been described, one dependent on the cytosolic ubiquitin-proteasome machinery, and the other dependent on vacuolar proteolysis. Interestingly, three of the novel Gid proteins involved in ubiquitin-proteasome–dependent degradation have also been reported by others to affect the vacuolar degradation pathway. As shown herein, additional genes suggested to be essential for vacuolar degradation are unnecessary for proteasome-dependent degradation. These data raise the question as to whether two FBPase degradation pathways exist that share components. Detailed characterization of Gid2p demonstrates that it is part of a soluble, cytosolic protein complex of at least 600 kDa. Gid2p is necessary for FBPase ubiquitination. Our studies have not revealed any involvement of vesicular intermediates in proteasome-dependent FBPase degradation. The influence of Ubp14p, a deubiquitinating enzyme, on proteasome-dependent catabolite degradation was further uncovered. PMID:12686616

Regelmann, Jochen; Schüle, Thomas; Josupeit, Frank S.; Horak, Jaroslav; Rose, Matthias; Entian, Karl-Dieter; Thumm, Michael; Wolf, Dieter H.

2003-01-01

84

Catabolite degradation of fructose-1,6-bisphosphatase in the yeast Saccharomyces cerevisiae: a genome-wide screen identifies eight novel GID genes and indicates the existence of two degradation pathways.  

PubMed

Metabolic adaptation of Saccharomyces cerevisiae cells from a nonfermentable carbon source to glucose induces selective, rapid breakdown of the gluconeogenetic key enzyme fructose-1,6-bisphosphatase (FBPase), a process called catabolite degradation. Herein, we identify eight novel GID genes required for proteasome-dependent catabolite degradation of FBPase. Four yeast proteins contain the CTLH domain of unknown function. All of them are Gid proteins. The site of catabolite degradation has been controversial until now. Two FBPase degradation pathways have been described, one dependent on the cytosolic ubiquitin-proteasome machinery, and the other dependent on vacuolar proteolysis. Interestingly, three of the novel Gid proteins involved in ubiquitin-proteasome-dependent degradation have also been reported by others to affect the vacuolar degradation pathway. As shown herein, additional genes suggested to be essential for vacuolar degradation are unnecessary for proteasome-dependent degradation. These data raise the question as to whether two FBPase degradation pathways exist that share components. Detailed characterization of Gid2p demonstrates that it is part of a soluble, cytosolic protein complex of at least 600 kDa. Gid2p is necessary for FBPase ubiquitination. Our studies have not revealed any involvement of vesicular intermediates in proteasome-dependent FBPase degradation. The influence of Ubp14p, a deubiquitinating enzyme, on proteasome-dependent catabolite degradation was further uncovered. PMID:12686616

Regelmann, Jochen; Schüle, Thomas; Josupeit, Frank S; Horak, Jaroslav; Rose, Matthias; Entian, Karl-Dieter; Thumm, Michael; Wolf, Dieter H

2003-04-01

85

The ubiquitin–proteasome system as a molecular target in solid tumors: an update on bortezomib  

PubMed Central

The ubiquitin–proteasome system has become a promising molecular target in cancer therapy due to its critical role in cellular protein degradation, interaction with cell cycle and apoptosis regulation, and unique mechanism of action. Bortezomib (PS-341) is a potent and specific reversible proteasome inhibitor, which has shown strong in vitro antitumor activity as single agent and in combination with other cytotoxic drugs in a broad spectrum of hematological and solid malignancies. In preclinical studies, bortezomib induced apoptosis of malignant cells through the inhibition of NF-|B and stabilization of pro-apoptotic proteins. Bortezomib also promotes chemo- and radiosensitization of malignant cells in vitro and inhibits tumor growth in murine xenograft models. The proteasome has been established as a relevant target in hematologic malignancies and bortezomib has been approved for the treatment of multiple myeloma. This review summarizes recent data from clinical trials in solid tumors. PMID:20616904

Milano, A; Perri, F; Caponigro, F

2009-01-01

86

The Ubiquitin–Proteasome System and the Autophagic–Lysosomal System in Alzheimer Disease  

PubMed Central

As neurons age, their survival depends on eliminating a growing burden of damaged, potentially toxic proteins and organelles—a capability that declines owing to aging and disease factors. Here, we review the two proteolytic systems principally responsible for protein quality control in neurons and their important contributions to Alzheimer disease pathogenesis. In the first section, the discovery of paired helical filament ubiquitination is described as a backdrop for discussing the importance of the ubiquitin–proteasome system in Alzheimer disease. In the second section, we review the prominent involvement of the lysosomal system beginning with pathological endosomal–lysosomal activation and signaling at the very earliest stages of Alzheimer disease followed by the progressive failure of autophagy. These abnormalities, which result in part from Alzheimer-related genes acting directly on these lysosomal pathways, contribute to the development of each of the Alzheimer neuropathological hallmarks and represent a promising therapeutic target. PMID:22908190

Ihara, Yasuo; Morishima-Kawashima, Maho; Nixon, Ralph

2012-01-01

87

Transcriptome analysis reveals link between proteasomal and mitochondrial pathways in Parkinson’s disease  

Microsoft Academic Search

There is growing evidence that dysfunction of the mitochondrial respiratory chain and failure of the cellular protein degradation machinery, specifically the ubiquitin–proteasome system, play an important role in the pathogenesis of Parkinson’s disease. We now show that the corresponding pathways of these two systems are linked at the transcriptomic level in Parkinsonian substantia nigra. We examined gene expression in medial

D. C. Duke; L. B. Moran; M. E. Kalaitzakis; M. Deprez; D. T. Dexter; R. K. B. Pearce; M. B. Graeber

2006-01-01

88

Plant Development: Regulation by Protein Degradation  

NSDL National Science Digital Library

Many aspects of eukaryotic development depend on regulated protein degradation by the ubiquitin-proteasome pathway. This highly conserved pathway promotes covalent attachment of ubiquitin to protein substrates through the sequential action of three enzymes called a ubiquitin-activating enzyme (E1), a ubiquitin-conjugating enzyme (E2), and a ubiquitin-protein ligase (E3). Most ubiquitinated proteins are then targeted for degradation by the 26S proteasome. Recent studies have also shown that the ubiquitin-related protein RUB/Nedd8 and the proteasome-related COP9 signalosome complex cooperate with the ubiquitin-proteasome pathway to promote protein degradation. Most of these components are conserved in all three eukaryotic kingdoms. However, the known targets of the pathway in plants, and the developmental processes they regulate, are specific to the plant kingdom.

Hanjo Hellmann (University of Texas;Molecular Cell and Developmental Biology); Mark Estelle (University of Texas;Molecular Cell and Developmental Biology)

2002-08-02

89

Autophagic-Lysosomal Inhibition Compromises Ubiquitin-Proteasome System Performance in a p62 Dependent Manner in Cardiomyocytes  

PubMed Central

Intracellular protein degradation is primarily performed by the ubiquitin-proteasome system (UPS) and the autophagic-lysosomal pathway (ALP). The interplay between these two pathways has been rarely examined in intact animals and the mechanism underlying the interplay remains unclear. Hence, we sought to test in vivo and in vitro the impact of inhibition of the ALP on UPS proteolytic performance in cardiomyocytes and to explore the underlying mechanism. Transgenic mice ubiquitously expressing a surrogate UPS substrate (GFPdgn) were treated with bafilomycin-A1 (BFA) to inhibit the ALP. Myocardial and renal GFPdgn protein levels but not mRNA levels were increased at 24 hours but not 3 hours after the first injection of BFA. Myocardial protein abundance of key proteasome subunits and the activities of proteasomal peptidases were not discernibly altered by the treatment. In cultured neonatal rat ventricular myocytes (NRVMs), the surrogate UPS substrate GFPu and a control red fluorescence protein (RFP) were co-expressed to probe UPS performance. At 12 hours or 24 hours after ALP inhibition by 3-methyladenine (3-MA) or BFA, GFPu/RFP protein ratios and the protein half-life of GFPu were significantly increased, which is accompanied by increases in p62 proteins. Similar findings were obtained when ALP was inhibited genetically via silencing Atg7 or Rab7. ALP inhibition-induced increases in GFPu and p62 are co-localized in NRVMs. siRNA-mediated p62 knockdown prevented ALP inhibition from inducing GFPu accumulation in NRVMs. We conclude that in a p62-dependent fashion, ALP inhibition impairs cardiac UPS proteolytic performance in cardiomyocytes in vitro and in vivo. PMID:24959866

Tian, Zongwen; Wang, Changhua; Hu, Chengjun; Tian, Yihao; Liu, Jinbao; Wang, Xuejun

2014-01-01

90

Ageing has no effect on the regulation of the ubiquitin proteasome-related genes and proteins following resistance exercise  

PubMed Central

Skeletal muscle atrophy is a critical component of the ageing process. Age-related muscle wasting is due to disrupted muscle protein turnover, a process mediated in part by the ubiquitin proteasome pathway (UPP). Additionally, older subjects have been observed to have an attenuated anabolic response, at both the molecular and physiological levels, following a single-bout of resistance exercise (RE). We investigated the expression levels of the UPP-related genes and proteins involved in muscle protein degradation in 10 older (60–75 years) vs. 10 younger (18–30 years) healthy male subjects at basal as well as 2 h after a single-bout of RE. MURF1, atrogin-1 and FBXO40, their substrate targets PKM2, myogenin, MYOD, MHC and EIF3F as well as MURF1 and atrogin-1 transcriptional regulators FOXO1 and FOXO3 gene and/or protein expression levels were measured via real time PCR and western blotting, respectively. At basal, no age-related difference was observed in the gene/protein levels of atrogin-1, MURF1, myogenin, MYOD and FOXO1/3. However, a decrease in FBXO40 mRNA and protein levels was observed in older subjects, while PKM2 protein was increased. In response to RE, MURF1, atrogin-1 and FBXO40 mRNA were upregulated in both the younger and older subjects, with changes observed in protein levels. In conclusion, UPP-related gene/protein expression is comparably regulated in healthy young and old male subjects at basal and following RE. These findings suggest that UPP signaling plays a limited role in the process of age-related muscle wasting. Future studies are required to investigate additional proteolytic mechanisms in conjunction with skeletal muscle protein breakdown (MPB) measurements following RE in older vs. younger subjects. PMID:24550841

Stefanetti, Renae J.; Zacharewicz, Evelyn; Della Gatta, Paul; Garnham, Andrew; Russell, Aaron P.; Lamon, Séverine

2014-01-01

91

Ubiquitin pathway and ovarian cancer  

PubMed Central

The ubiquitin–proteasome pathway is a common cellular process in eukaryotic tissue. Ubiquitin binds to proteins and tags them for destruction; this tagging directs proteins to the proteosome in the cell that degrades and recycles unneeded proteins. The ubiquitin–proteasome pathway plays an important role in the regulation of cellular proteins with respect to cell cycle control, transcription, apoptosis, cell adhesion, angiogenesis, and tumour growth. This review article discusses the various ways that the ubiquitin pathway is involved in ovarian cancer, such as modulating the ovarian-cancer-related gene BRCA1 and tumour suppressor p53, and interfering with the erk pathway, the cyclin-dependent cell cycle regulation process, and ERBB2 gene expression. PMID:23300358

Rao, Z.; Ding, Y.

2012-01-01

92

An extract of Artemisia dracunculus L. inhibits ubiquitin-proteasome activity and preserves skeletal muscle mass in a murine model of diabetes.  

PubMed

Impaired insulin signaling is a key feature of type 2 diabetes and is associated with increased ubiquitin-proteasome-dependent protein degradation in skeletal muscle. An extract of Artemisia dracunculus L. (termed PMI5011) improves insulin action by increasing insulin signaling in skeletal muscle. We sought to determine if the effect of PMI5011 on insulin signaling extends to regulation of the ubiquitin-proteasome system. C2C12 myotubes and the KK-A(y) murine model of type 2 diabetes were used to evaluate the effect of PMI5011 on steady-state levels of ubiquitylation, proteasome activity and expression of Atrogin-1 and MuRF-1, muscle-specific ubiquitin ligases that are upregulated with impaired insulin signaling. Our results show that PMI5011 inhibits proteasome activity and steady-state ubiquitylation levels in vitro and in vivo. The effect of PMI5011 is mediated by PI3K/Akt signaling and correlates with decreased expression of Atrogin-1 and MuRF-1. Under in vitro conditions of hormonal or fatty acid-induced insulin resistance, PMI5011 improves insulin signaling and reduces Atrogin-1 and MuRF-1 protein levels. In the KK-A(y) murine model of type 2 diabetes, skeletal muscle ubiquitylation and proteasome activity is inhibited and Atrogin-1 and MuRF-1 expression is decreased by PMI5011. PMI5011-mediated changes in the ubiquitin-proteasome system in vivo correlate with increased phosphorylation of Akt and FoxO3a and increased myofiber size. The changes in Atrogin-1 and MuRF-1 expression, ubiquitin-proteasome activity and myofiber size modulated by PMI5011 in the presence of insulin resistance indicate the botanical extract PMI5011 may have therapeutic potential in the preservation of muscle mass in type 2 diabetes. PMID:23437325

Kirk-Ballard, Heather; Wang, Zhong Q; Acharya, Priyanka; Zhang, Xian H; Yu, Yongmei; Kilroy, Gail; Ribnicky, David; Cefalu, William T; Floyd, Z Elizabeth

2013-01-01

93

An Extract of Artemisia dracunculus L. Inhibits Ubiquitin-Proteasome Activity and Preserves Skeletal Muscle Mass in a Murine Model of Diabetes  

PubMed Central

Impaired insulin signaling is a key feature of type 2 diabetes and is associated with increased ubiquitin-proteasome-dependent protein degradation in skeletal muscle. An extract of Artemisia dracunculus L. (termed PMI5011) improves insulin action by increasing insulin signaling in skeletal muscle. We sought to determine if the effect of PMI5011 on insulin signaling extends to regulation of the ubiquitin-proteasome system. C2C12 myotubes and the KK-Ay murine model of type 2 diabetes were used to evaluate the effect of PMI5011 on steady-state levels of ubiquitylation, proteasome activity and expression of Atrogin-1 and MuRF-1, muscle-specific ubiquitin ligases that are upregulated with impaired insulin signaling. Our results show that PMI5011 inhibits proteasome activity and steady-state ubiquitylation levels in vitro and in vivo. The effect of PMI5011 is mediated by PI3K/Akt signaling and correlates with decreased expression of Atrogin-1 and MuRF-1. Under in vitro conditions of hormonal or fatty acid-induced insulin resistance, PMI5011 improves insulin signaling and reduces Atrogin-1 and MuRF-1 protein levels. In the KK-Ay murine model of type 2 diabetes, skeletal muscle ubiquitylation and proteasome activity is inhibited and Atrogin-1 and MuRF-1 expression is decreased by PMI5011. PMI5011-mediated changes in the ubiquitin-proteasome system in vivo correlate with increased phosphorylation of Akt and FoxO3a and increased myofiber size. The changes in Atrogin-1 and MuRF-1 expression, ubiquitin-proteasome activity and myofiber size modulated by PMI5011 in the presence of insulin resistance indicate the botanical extract PMI5011 may have therapeutic potential in the preservation of muscle mass in type 2 diabetes. PMID:23437325

Kirk-Ballard, Heather; Wang, Zhong Q.; Acharya, Priyanka; Zhang, Xian H.; Yu, Yongmei; Kilroy, Gail; Ribnicky, David; Cefalu, William T.; Floyd, Z. Elizabeth

2013-01-01

94

The Amazing Ubiquitin-Proteasome System: Structural Components and Implication in Aging.  

PubMed

Proteome quality control (PQC) is critical for the maintenance of cellular functionality and it is assured by the curating activity of the proteostasis network (PN). PN is constituted of several complex protein machines that under conditions of proteome instability aim to, firstly identify, and then, either rescue or degrade nonnative polypeptides. Central to the PN functionality is the ubiquitin-proteasome system (UPS) which is composed from the ubiquitin-conjugating enzymes and the proteasome; the latter is a sophisticated multi-subunit molecular machine that functions in a bimodal way as it degrades both short-lived ubiquitinated normal proteins and nonfunctional polypeptides. UPS is also involved in PQC of the nucleus, the endoplasmic reticulum and the mitochondria and it also interacts with the other main cellular degradation axis, namely the autophagy-lysosome system. UPS functionality is optimum in the young organism but it is gradually compromised during aging resulting in increasing proteotoxic stress; these effects correlate not only with aging but also with most age-related diseases. Herein, we present a synopsis of the UPS components and of their functional alterations during cellular senescence or in vivo aging. We propose that mild UPS activation in the young organism will, likely, promote antiaging effects and/or suppress age-related diseases. PMID:25619718

Tsakiri, Eleni N; Trougakos, Ioannis P

2015-01-01

95

Integrated Analysis of Microarray Data of Atherosclerotic Plaques: Modulation of the Ubiquitin-Proteasome System  

PubMed Central

Atherosclerosis is a typical complex multi-factorial disease and many molecules at different levels and pathways were involved in its development. Some studies have investigated the dysregulation in atherosclerosis at mRNA, miRNA or DNA methylation level, respectively. However, to our knowledge, the studies that integrated these data and revealed the abnormal networks of atherosclerosis have not been reported. Using microarray technology, we analyzed the omics data in atherosclerosis at mRNA, miRNA and DNA methylation levels. Our results demonstrated that the global DNA methylation and expression of miRNA/mRNA were significantly decreased in atherosclerotic plaque than in normal vascular tissue. The interaction network constructed using the integrative data revealed many genes, cellular processes and signaling pathways which were widely considered to play crucial roles in atherosclerosis and also revealed some genes, miRNAs or signaling pathways which have not been investigated in atherosclerosis until now (e.g. miR-519d and SNTB2). Moreover, the overall protein ubiquitination in atherosclerotic plaque was significantly increased. The proteasome activity was increased early but decreased in advanced atherosclerosis. Our study revealed many classic and novel genes and miRNAs involved in atherosclerosis and indicated the effects of ubiquitin-proteasome system on atherosclerosis might be closely related to the course of atherosclerosis. However, the efficacy of proteasome inhibitors in the treatment of atherosclerosis still needs more research. PMID:25333956

Wang, Jian; Wang, Rong; Wang, Xiaowei; Zhang, Qunye

2014-01-01

96

Alterations in the ubiquitin proteasome system in persistent but not reversible proteinuric diseases.  

PubMed

Podocytes are the key cells affected in nephrotic glomerular kidney diseases, and they respond uniformly to injury with cytoskeletal rearrangement. In nephrotic diseases, such as membranous nephropathy and FSGS, persistent injury often leads to irreversible structural damage, whereas in minimal change disease, structural alterations are mostly transient. The factors leading to persistent podocyte injury are currently unknown. Proteolysis is an irreversible process and could trigger persistent podocyte injury through degradation of podocyte-specific proteins. We, therefore, analyzed the expression and functional consequence of the two most prominent proteolytic systems, the ubiquitin proteasome system (UPS) and the autophagosomal/lysosomal system, in persistent and transient podocyte injuries. We show that differential upregulation of both proteolytic systems occurs in persistent human and rodent podocyte injury. The expression of specific UPS proteins in podocytes differentiated children with minimal change disease from children with FSGS and correlated with poor clinical outcome. Degradation of the podocyte-specific protein ?-actinin-4 by the UPS depended on oxidative modification in membranous nephropathy. Notably, the UPS was overwhelmed in podocytes during experimental glomerular disease, resulting in abnormal protein accumulation and compensatory upregulation of the autophagosomal/lysosomal system. Accordingly, inhibition of both proteolytic systems enhanced proteinuria in persistent nephrotic disease. This study identifies altered proteolysis as a feature of persistent podocyte injury. In the future, specific UPS proteins may serve as new biomarkers or therapeutic targets in persistent nephrotic syndrome. PMID:24722446

Beeken, Maire; Lindenmeyer, Maja T; Blattner, Simone M; Radón, Victoria; Oh, Jun; Meyer, Tobias N; Hildebrand, Diana; Schlüter, Hartmut; Reinicke, Anna T; Knop, Jan-Hendrik; Vivekanandan-Giri, Anuradha; Münster, Silvia; Sachs, Marlies; Wiech, Thorsten; Pennathur, Subramaniam; Cohen, Clemens D; Kretzler, Matthias; Stahl, Rolf A K; Meyer-Schwesinger, Catherine

2014-11-01

97

The ubiquitin proteasome system in glia and its role in neurodegenerative diseases  

PubMed Central

The ubiquitin proteasome system (UPS) is crucial for intracellular protein homeostasis and for degradation of aberrant and damaged proteins. The accumulation of ubiquitinated proteins is a hallmark of many neurodegenerative diseases, including amyotrophic lateral sclerosis, Alzheimer’s, Parkinson’s, and Huntington’s disease, leading to the hypothesis that proteasomal impairment is contributing to these diseases. So far, most research related to the UPS in neurodegenerative diseases has been focused on neurons, while glial cells have been largely disregarded in this respect. However, glial cells are essential for proper neuronal function and adopt a reactive phenotype in neurodegenerative diseases, thereby contributing to an inflammatory response. This process is called reactive gliosis, which in turn affects UPS function in glial cells. In many neurodegenerative diseases, mostly neurons show accumulation and aggregation of ubiquitinated proteins, suggesting that glial cells may be better equipped to maintain proper protein homeostasis. During an inflammatory reaction, the immunoproteasome is induced in glia, which may contribute to a more efficient degradation of disease-related proteins. Here we review the role of the UPS in glial cells in various neurodegenerative diseases, and we discuss how studying glial cell function might provide essential information in unraveling mechanisms of neurodegenerative diseases. PMID:25152710

Jansen, Anne H. P.; Reits, Eric A. J.; Hol, Elly M.

2014-01-01

98

Down-regulation of the ubiquitin–proteasome proteolysis system by amino acids and insulin involves the adenosine monophosphate-activated protein kinase and mammalian target of rapamycin pathways in rat hepatocytes  

Microsoft Academic Search

The purpose of this work was to examine whether changes in dietary protein levels could elicit differential responses of tissue\\u000a proteolysis and the pathway involved in this response. In rats fed with a high protein diet (55%) for 14 days, the liver was\\u000a the main organ where adaptations occurred, characterized by an increased protein pool and a strong, meal-induced inhibition\\u000a of

Nattida Chotechuang; Dalila Azzout-Marniche; Cécile Bos; Catherine Chaumontet; Claire Gaudichon; Daniel Tomé

2011-01-01

99

Hepatitis B virus HBx protein interactions with the ubiquitin proteasome system.  

PubMed

The hepatitis B virus (HBV) causes acute and chronic hepatitis, and the latter is a major risk factor for the development of hepatocellular carcinoma (HCC). HBV encodes a 17-kDa regulatory protein, HBx, which is required for virus replication. Although the precise contribution(s) of HBx to virus replication is unknown, many viruses target cellular pathways to create an environment favorable for virus replication. The ubiquitin proteasome system (UPS) is a major conserved cellular pathway that controls several critical processes in the cell by regulating the levels of proteins involved in cell cycle, DNA repair, innate immunity, and other processes. We summarize here the interactions of HBx with components of the UPS, including the CUL4 adaptor DDB1, the cullin regulatory complex CSN, and the 26S proteasome. Understanding how these protein interactions benefit virus replication remains a challenge due to limited models in which to study HBV replication. However, studies from other viral systems that similarly target the UPS provide insight into possible strategies used by HBV. PMID:25421893

Minor, Marissa M; Slagle, Betty L

2014-11-01

100

Conditional Transgenic System for Mouse Aurora A Kinase: Degradation by the Ubiquitin Proteasome Pathway Controls the Level of the Transgenic Protein  

Microsoft Academic Search

Aurora A is a mitotic kinase that localizes to centrosomes. Expression of this protein is normally limited to the mitotic stage (G2-M) of the cell cycle, whereas human cancer cells frequently exhibit overexpression of Aurora A protein regardless of the cell cycle stage. In the present study, Aurora A transgenic mouse lines were generated with a new conditional expression system

Tomokazu Fukuda; Yuji Mishina; Michael P. Walker; Richard P. DiAugustine

2005-01-01

101

The ubiquitin-proteasome system in neurodegenerative diseases: precipitating factor, yet part of the solution  

PubMed Central

The ubiquitin-proteasome system (UPS) has been implicated in neurodegenerative diseases based on the presence of deposits consisting of ubiquitylated proteins in affected neurons. It has been postulated that aggregation-prone proteins associated with these disorders, such as ?-synuclein, ?-amyloid peptide, and polyglutamine proteins, compromise UPS function, and delay the degradation of other proteasome substrates. Many of these substrates play important regulatory roles in signaling, cell cycle progression, or apoptosis, and their inadvertent stabilization due to an overloaded and improperly functioning UPS may thus be responsible for cellular demise in neurodegeneration. Over the past decade, numerous studies have addressed the UPS dysfunction hypothesis using various model systems and techniques that differ in their readout and sensitivity. While an inhibitory effect of some disease proteins on the UPS has been demonstrated, increasing evidence attests that the UPS remains operative in many disease models, which opens new possibilities for treatment. In this review, we will discuss the paradigm shift that repositioned the UPS from being a prime suspect in the pathophysiology of neurodegeneration to an attractive therapeutic target that can be harnessed to accelerate the clearance of disease-linked proteins. PMID:25132814

Dantuma, Nico P.; Bott, Laura C.

2014-01-01

102

The ubiquitin proteasome system – Implications for cell cycle control and the targeted treatment of cancer  

PubMed Central

Two families of E3 ubiquitin ligases are prominent in cell cycle regulation and mediate the timely and precise ubiquitin-proteasome-dependent degradation of key cell cycle proteins: the SCF (Skp1/Cul1/F-box protein) complex and the APC/C (Anaphase Promoting Complex or Cyclosome). While certain SCF ligases drive cell cycle progression throughout the cell cycle, APC/C (in complex with either of two substrate recruiting proteins: Cdc20 and Cdh1) orchestrates exit from mitosis (APC/CCdc20) and establishes a stable G1 phase (APC/CCdh1). Upon DNA damage or perturbation of the normal cell cycle, both ligases are involved in checkpoint activation. Mechanistic insight into these processes has significantly improved over the last ten years, largely due to a better understanding of APC/C and the functional characterization of multiple F-box proteins, the variable substrate recruiting components of SCF ligases. Here, we review the role of SCF- and APC/C-mediated ubiquitylation in the normal and perturbed cell cycle and discuss potential clinical implications of SCF and APC/C functions. PMID:23466868

Bassermann, Florian; Eichner, Ruth; Pagano, Michele

2013-01-01

103

Eukaryotic Cells Producing Ribosomes Deficient in Rpl1 Are Hypersensitive to Defects in the Ubiquitin-Proteasome System  

PubMed Central

It has recently become clear that the misassembly of ribosomes in eukaryotic cells can have deleterious effects that go far beyond a simple shortage of ribosomes. In this work we find that cells deficient in ribosomal protein L1 (Rpl1; Rpl10a in mammals) produce ribosomes lacking Rpl1 that are exported to the cytoplasm and that can be incorporated into polyribosomes. The presence of such defective ribosomes leads to slow growth and appears to render the cells hypersensitive to lesions in the ubiquitin-proteasome system. Several genes that were reasonable candidates for degradation of 60S subunits lacking Rpl1 fail to do so, suggesting that key players in the surveillance of ribosomal subunits remain to be found. Interestingly, in spite of rendering the cells hypersensitive to the proteasome inhibitor MG132, shortage of Rpl1 partially suppresses the stress-invoked temporary repression of ribosome synthesis caused by MG132. PMID:21858174

McIntosh, Kerri B.; Bhattacharya, Arpita; Willis, Ian M.; Warner, Jonathan R.

2011-01-01

104

ATF4 Degradation Relies on a Phosphorylation-Dependent Interaction with the SCF TrCP Ubiquitin Ligase  

Microsoft Academic Search

The ubiquitin-proteasome pathway regulates gene expression through protein degradation. Here we show that the F-box protein bTrCP, the receptor component of the SCF E3 ubiquitin ligase responsible for IkBa and b-catenin degradation, is colocalized in the nucleus with ATF4, a member of the ATF-CREB bZIP family of transcription factors, and controls its stability. Association between the two proteins depends on

IRINA LASSOT; EMMANUEL SEGERAL; CLARISSE BERLIOZ-TORRENT; HERVE DURAND; LIONEL GROUSSIN; TSONWIN HAI; RICHARD BENAROUS; FLORENCE MARGOTTIN-GOGUET

2001-01-01

105

Atrophy, hypertrophy, and hypoxemia induce transcriptional regulators of the ubiquitin proteasome system in the rat heart  

Technology Transfer Automated Retrieval System (TEKTRAN)

In skeletal muscle, transcript levels of proteins regulating the ubiquitin proteasome system (UPS) increase with atrophy and decrease with hypertrophy. Whether the same is true for heart muscle is not known. We set out to characterize the transcriptional profile of regulators of the UPS during atrop...

106

Pesticides that inhibit the ubiquitin-proteasome system: effect measure modification by genetic variation in SKP1 in Parkinson?s disease.  

PubMed

Cytoplasmic inclusions known as Lewy bodies, a hallmark of Parkinson's disease (PD) pathology, may protect against cytotoxic proteins. Since the ubiquitin-proteasome system (UPS) degrades cytotoxic proteins, dysfunction in the UPS may contribute to PD etiology. Our goal in this study was to screen pesticides for proteasome inhibition and investigate (i) whether ambient exposures to pesticides that inhibit the UPS increase PD risk and (ii) whether genetic variation in candidate genes of the UPS pathway modify those increased risks. We assessed 26S UPS activity in SK-N-MC(u) cells by fluorescence. We recruited idiopathic PD cases (n=360) and population-based controls (n=816) from three counties in California with considerable commercial agriculture. We determined ambient pesticide exposure by our validated GIS-based model utilizing residential and workplace address histories. We limited effect measure modification assessment to Caucasians (287 cases, 453 controls). Eleven of 28 pesticides we screened inhibited 26S UPS activity at 10 µM. Benomyl, cyanazine, dieldrin, endosulfan, metam, propargite, triflumizole, and ziram were associated with increased PD risk. We estimated an odds ratio of 2.14 (95% CI: 1.42, 3.22) for subjects with ambient exposure to any UPS-inhibiting pesticide at both residential and workplace addresses; this association was modified by genetic variation in the s-phase kinase-associated protein 1 gene (SKP1; interaction p-value=0.005). Our results provide evidence that UPS-inhibiting pesticides play a role in the etiology of PD and suggest that genetic variation in candidate genes involved in the UPS pathway might exacerbate the toxic effects of pesticide exposures. PMID:23988235

Rhodes, Shannon L; Fitzmaurice, Arthur G; Cockburn, Myles; Bronstein, Jeff M; Sinsheimer, Janet S; Ritz, Beate

2013-10-01

107

Pesticides that Inhibit the Ubiquitin-Proteasome System: Effect Measure Modification by Genetic Variation in SKP1 in Parkinson’s Disease  

PubMed Central

Cytoplasmic inclusions known as Lewy bodies, a hallmark of Parkinson’s disease (PD) pathology, may protect against cytotoxic proteins. Since the ubiquitin-proteasome system (UPS) degrades cytotoxic proteins, dysfunction in the UPS may contribute to PD etiology. Our goal in this study was to screen pesticides for proteasome inhibition and investigate (i) whether ambient exposures to pesticides that inhibit the UPS increase PD risk and (ii) whether genetic variation in candidate genes of the UPS pathway modify those increased risks. We assessed 26S UPS activity in SK-N-MCu cells by fluorescence. We recruited idiopathic PD cases (n=360) and population-based controls (n=816) from three counties in California with considerable commercial agriculture. We determined ambient pesticide exposure by our validated GIS-based model utilizing residential and workplace address histories. We limited effect measure modification assessment to Caucasians (287 cases, 453 controls). 11 of 28 pesticides we screened inhibited 26S UPS activity at 10?M. Benomyl, cyanazine, dieldrin, endosulfan, metam, propargite, triflumizole, and ziram were associated with increased PD risk. We estimated an odds ratio of 2.14 (95%CI: 1.42,3.22) for subjects with ambient exposure to any UPS-inhibiting pesticide at both residential and workplace addresses; this association was modified by genetic variation in the s-phase kinase-associated protein 1 gene (SKP1; interaction p-value=0.005). Our results provide evidence that UPS-inhibiting pesticides play a role in the etiology of PD and suggest that genetic variation in candidate genes involved in the UPS pathway might exacerbate the toxic effects of pesticide exposures. PMID:23988235

Rhodes, Shannon L.; Fitzmaurice, Arthur G.; Cockburn, Myles; Bronstein, Jeff M.; Sinsheimer, Janet S.; Ritz, Beate

2013-01-01

108

Regulation of TRAIL-Receptor Expression by the Ubiquitin-Proteasome System  

PubMed Central

The tumor necrosis factor (TNF)-related apoptosis-inducing ligand- receptor (TRAIL-R) family has emerged as a key mediator of cell fate and survival. Ligation of TRAIL ligand to TRAIL-R1 or TRAIL-R2 initiates the extrinsic apoptotic pathway characterized by the recruitment of death domains, assembly of the death-inducing signaling complex (DISC), caspase activation and ultimately apoptosis. Conversely the decoy receptors TRAIL-R3 and TRAIL-R4, which lack the pro-apoptotic death domain, function to dampen the apoptotic response by competing for TRAIL ligand. The tissue restricted expression of the decoy receptors on normal but not cancer cells provides a therapeutic rational for the development of selective TRAIL-mediated anti-tumor therapies. Recent clinical trials using agonistic antibodies against the apoptosis-inducing TRAIL receptors or recombinant TRAIL have been promising; however the number of patients in complete remission remains stubbornly low. The mechanisms of TRAIL resistance are relatively unexplored but may in part be due to TRAIL-R down-regulation or shedding of TRAIL-R by tumor cells. Therefore a better understanding of the mechanisms underlying TRAIL resistance is required. The ubiquitin-proteasome system (UPS) has been shown to regulate TRAIL-R members suggesting that pharmacological inhibition of the UPS may be a novel strategy to augment TRAIL-based therapies and increase efficacies. We recently identified b-AP15 as an inhibitor of proteasome deubiquitinase (DUB) activity. Interestingly, exposure of tumor cell lines to b-AP15 resulted in increased TRAIL-R2 expression and enhanced sensitivity to TRAIL-mediated apoptosis and cell death in vitro and in vivo. In conclusion, targeting the UPS may represent a novel strategy to increase the cell surface expression of pro-apoptotic TRAIL-R on cancer cells and should be considered in clinical trials targeting TRAIL-receptors in cancer patients. PMID:25318057

Sarhan, Dhifaf; D’Arcy, Padraig; Lundqvist, Andreas

2014-01-01

109

Aging and calorie restriction modulate yeast redox state, oxidized protein removal, and the ubiquitin–proteasome system  

Microsoft Academic Search

The ubiquitin–proteasome system governs the half-life of most cellular proteins. Calorie restriction (CR) extends the maximum life span of a variety of species and prevents oxidized protein accumulation. We studied the effects of CR on the ubiquitin–proteasome system and protein turnover in aging Saccharomyces cerevisiae. CR increased chronological life span as well as proteasome activity compared to control cells. The

Fernanda Marques da Cunha; Marilene Demasi; Alicia J. Kowaltowski

2011-01-01

110

Reproductive Cytotoxicity Is Predicted by Magnetic Resonance Microscopy and Confirmed by Ubiquitin Proteasome Immunohistochemistry in a Theophylline-Induced Model of Rat Testicular and Epididymal Toxicity  

NASA Astrophysics Data System (ADS)

This study investigated the testicular changes in the rat induced by the nonspecific phosphodiesterase inhibitor, theophylline using magnetic resonance microscopy (MRM) and ubiquitin immunostaining techniques. In vivo T1- and T2-weighted images were acquired at 2 T under anesthesia. Increased signal observed in the theophylline-treated rats suggests that leakage of MRM contrast was occurring. In vivo MRM results indicate that day 16 testis displayed an increased T1-weighted water signal in the area of the seminiferous tubule that decreased by day 32. These findings were validated by histopathology, suggesting that in vivo MRM has the sensitivity to predict changes in testis and epididymal tissues. The participation of the ubiquitin system was investigated, using probes for various markers of the ubiquitin-proteasome pathway. MRM can be used to detect subtle changes in the vascular perfusion of organ systems, and the up-regulation/mobilization of ubiquitin-proteasome pathway may be one of the mechanisms used in theophylline-treated epididymis to remove damaged cells before storage in the cauda epididymis. The combined use of in vivo MRM and subsequent tissue or seminal analysis for the presence of ubiquitin in longitudinal studies may become an important biomarker for assessing testis toxicities drug studies.

Tengowski, M. W.; Sutovsky, P.; Hedlund, L. W.; Guyot, D. J.; Burkhardt, J. E.; Thompson, W. E.; Sutovsky, M.; Johnson, G. A.

2005-08-01

111

Pyrrolidine dithiocarbamate inhibits enterovirus 71 replication by down-regulating ubiquitin-proteasome system.  

PubMed

Enterovirus 71 (EV71) is the main causative pathogen of hand, foot, and mouth disease (HFMD). The severe neurological complications caused by EV71 infection and the lack of effective therapeutic medicine underline the importance of searching for antiviral substances. Pyrrolidine dithiocarbamate (PDTC), an antioxidant, has been reported to inhibit the replication of coxsackievirus B (CVB) through dysregulating ubiquitin-proteasome system (UPS). In this study, we demonstrated that PDTC exerted potent antiviral effect on EV71. Viral RNA synthesis, viral protein expression, and the production of viral progeny were significantly reduced by the treatment of PDTC in Vero cells infected with EV71. Similar to the previous report about the inhibitory effect of PDTC on UPS, we found that PDTC treatment led to decreased levels of polyubiquitinated proteins in EV71-infected cells. The inhibitory effect of PDTC on UPS was further confirmed by the increased accumulation of cell cycle regulatory proteins p21 and p53, which are normally degraded through UPS, while the expression levels of both proteins remained unchanged. We also showed that PDTC had no impact on the activity of proteasome. Thus, we demonstrated that the down-regulation of PDTC on UPS was the result of its inhibition on ubiquitination. More importantly, this study provides evidence that the inhibition on UPS was required for the antiviral activity of PDTC, since MG132, a potent proteasome inhibitor, significantly inhibited the cytopathic effect and viral protein synthesis in EV71-infected cells. We also found that the antioxidant property of PDTC did not contribute to its antiviral effect, since N-acetyl-l-cysteine, a potent antioxidant, could not inhibit viral replication. In addition, CPE and viral protein synthesis were not inhibited in the cells pretreated with PDTC 2h before viral infection and then cultured in the media with no PDTC supplement, while the antioxidant effect of PDTC was retained. PDTC also showed significant inhibition on apoptosis induced by EV71 infection when it was applied at the early stage of viral infection. Our results collectively suggest that PDTC could be a potential anti-EV71 compound which possesses both antiviral and anti-apoptotic capacity. PMID:25456405

Lin, Lexun; Qin, Ying; Wu, Heng; Chen, Yang; Wu, Shuo; Si, Xiaoning; Wang, Hui; Wang, Tianying; Zhong, Xiaoyan; Zhai, Xia; Tong, Lei; Pan, Bo; Zhang, Fengmin; Zhong, Zhaohua; Wang, Yan; Zhao, Wenran

2015-01-01

112

BRK Targets Dok1 for Ubiquitin-Mediated Proteasomal Degradation to Promote Cell Proliferation and Migration  

PubMed Central

Breast tumor kinase (BRK), also known as protein tyrosine kinase 6 (PTK6), is a non-receptor tyrosine kinase overexpressed in more that 60% of human breast carcinomas. The overexpression of BRK has been shown to sensitize mammary epithelial cells to mitogenic signaling and to promote cell proliferation and tumor formation. The molecular mechanisms of BRK have been unveiled by the identification and characterization of BRK target proteins. Downstream of tyrosine kinases 1 or Dok1 is a scaffolding protein and a substrate of several tyrosine kinases. Herein we show that BRK interacts with and phosphorylates Dok1 specifically on Y362. We demonstrate that this phosphorylation by BRK significantly downregulates Dok1 in a ubiquitin-proteasome-mediated mechanism. Together, these results suggest a novel mechanism of action of BRK in the promotion of tumor formation, which involves the targeting of tumor suppressor Dok1 for degradation through the ubiquitin proteasomal pathway. PMID:24523872

Miah, Sayem; Goel, Raghuveera Kumar; Dai, Chenlu; Kalra, Natasha; Beaton-Brown, Erika; Bagu, Edward T.; Bonham, Keith; Lukong, Kiven E.

2014-01-01

113

Deregulation of the ubiquitin-proteasome system is the predominant molecular pathology in OPMD animal models and patients  

PubMed Central

Oculopharyngeal muscular dystrophy (OPMD) is a late-onset progressive muscle disorder caused by a poly-alanine expansion mutation in the Poly(A) Binding Protein Nuclear 1 (PABPN1). The molecular mechanisms that regulate disease onset and progression are largely unknown. In order to identify molecular pathways that are consistently associated with OPMD, we performed an integrated high-throughput transcriptome study in affected muscles of OPMD animal models and patients. The ubiquitin-proteasome system (UPS) was found to be the most consistently and significantly OPMD-deregulated pathway across species. We could correlate the association of the UPS OPMD-deregulated genes with stages of disease progression. The expression trend of a subset of these genes is age-associated and therefore, marks the late onset of the disease, and a second group with expression trends relating to disease-progression. We demonstrate a correlation between expression trends and entrapment into PABPN1 insoluble aggregates of OPMD-deregulated E3 ligases. We also show that manipulations of proteasome and immunoproteasome activity specifically affect the accumulation and aggregation of mutant PABPN1. We suggest that the natural decrease in proteasome expression and its activity during muscle aging contributes to the onset of the disease. PMID:21798095

2011-01-01

114

Protective Role of Cell Division Cycle 48 (CDC48) Protein against Neurodegeneration via Ubiquitin-Proteasome System Dysfunction during Zebrafish Development*  

PubMed Central

Cell division cycle 48 (CDC48), a ubiquitin-dependent molecular chaperone, is thought to mediate a variety of degradative and regulatory processes and maintain cellular homoeostasis. To investigate the protective function of CDC48 against accumulated ubiquitinated proteins during neurodevelopment, we developed an in vivo bioassay technique that detects expression and accumulation of fluorescent proteins with a polyubiquitination signal at the N terminus. When we introduced CDC48 antisense morpholino oligonucleotides into zebrafish embryos, the morphant embryos were lethal and showed defects in neuronal outgrowth and neurodegeneration, and polyubiquitinated fluorescent proteins accumulated in the inner plexiform and ganglion cell layers, as well as the diencephalon and mesencephalon, indicating that the degradation of polyubiquitinated proteins by the ubiquitin-proteasome system was blocked. These abnormal phenotypes in the morphant were rescued by CDC48 or human valosin-containing protein overexpression. Therefore, the protective function of CDC48 is essential for neurodevelopment. PMID:22549779

Imamura, Shintaro; Yabu, Takeshi; Yamashita, Michiaki

2012-01-01

115

Ubiquitin/proteasome-mediated proteolysis is involved in the response to flooding stress in soybean roots, independent of oxygen limitation.  

PubMed

Ubiquitin/proteasome-mediated proteolysis plays an important role in the response to several environmental stresses. Here, we described the relationship of the proteolysis in the flooding stress in soybean (Glycine max L. cultivar Enrei). Immunoblot analyses were performed using antibodies against two subunits of 26S proteasome, Rpt5 and Rpn10, 20S proteasome and two subunits of COP9 signalosome (CSN), CSN4 and CSN5, to compare between flooded and untreated roots. We also examined their protein amounts in the condition of low oxygen. Moreover, crude extracts from flooded or untreated roots incubated with or without a proteasome inhibitor MG132 were analyzed by proteomics technique. We revealed that the amount of ubiquitinated proteins in soybean roots decreased after flooding treatment and increased to levels similar to controls after de-submergence. Both CSN4 and CSN5 accumulated following flooding treatment, although no significant difference was observed in proteasome. Low oxygen had no effect on the amount of ubiquitinated proteins or CSN4. By 2D-PAGE, the amount of 6 proteins changed significantly following MG132 treatment in flooding stressed plants. We conclude that the accumulation of CSN proteins might enhance the degradation of ubiquitinated proteins independent of hypoxia caused by flooding, thereby lowering their abundance during flooding stress. PMID:22325888

Yanagawa, Yuki; Komatsu, Setsuko

2012-04-01

116

Multiple Interactions between Peroxisome Proliferators-Activated Receptors and the Ubiquitin-Proteasome System and Implications for Cancer Pathogenesis  

PubMed Central

The peroxisome proliferator-activated receptors (PPAR) ?, ?/?, and ? are ligand-activated nuclear receptors involved in a number of physiological processes, including lipid and glucose homeostasis, inflammation, cell growth, differentiation, and death. PPAR agonists are used in the treatment of human diseases, like type 2 diabetes and dyslipidemia, and PPARs appear as promising therapeutic targets in other conditions, including cancer. A better understanding of the functions and regulation of PPARs in normal and pathological processes is of primary importance to devise appropriate therapeutic strategies. The ubiquitin-proteasome system (UPS) plays an important role in controlling level and activity of many nuclear receptors and transcription factors. PPARs are subjected to UPS-dependent regulation. Interestingly, the three PPAR isotypes are differentially regulated by the UPS in response to ligand-dependent activation, a phenomenon that may be intrinsically connected to their distinct cellular functions and behaviors. In addition to their effects ongene expression, PPARs appear to affect protein levels and downstream pathways also by modulating the activity of the UPS in target-specific manners. Here we review the current knowledge of the interactions between the UPS and PPARs in light of the potential implications for their effects on cell fate and tumorigenesis. PMID:18551186

Genini, Davide; Carbone, Giuseppina M.; Catapano, Carlo V.

2008-01-01

117

Narrative Review: Protein Degradation and Human Diseases: The Ubiquitin Connection  

NSDL National Science Digital Library

Physiology in Medicine review article. When researchers discovered the organelle lysosome, they assumed that cellular proteins were degraded within it. However, several independent lines of evidence strongly suggested that intracellular proteolysis was largely nonlysosomal. It is now recognized that degradation of intracellular proteins by the ubiquitin proteasome system (UPS) is involved in the regulation of a broad array of cellular processes,drugs that target the degradation pathways of a single or a few proteins without affecting other proteins. It seems that pharmacologic manipulation of the UPS might alter the outcome of many diseases, especially malignant conditions and possibly neurodegenerative and chronic inflammatory diseases.

MD/PhD Eyal Reinstein (Tel-Aviv University Deparrment of Internal Medicine)

2006-11-07

118

plantsUPS: a database of plants' Ubiquitin Proteasome System  

Microsoft Academic Search

BACKGROUND: The ubiquitin 26S\\/proteasome system (UPS), a serial cascade process of protein ubiquitination and degradation, is the last step for most cellular proteins. There are many genes involved in this system, but are not identified in many species. The accumulating availability of genomic sequence data is generating more demands in data management and analysis. Genomics data of plants such as

Zhou Du; Xin Zhou; Li Li; Zhen Su

2009-01-01

119

Ubiquitin-proteasome-dependent proteolysis in skeletal muscle  

E-print Network

pathway has recently been reported to be of major importance in the breakdown of skeletal muscle proteins be used to manipulate muscle protein mass. Such concepts are essential for the devel- opment of anti-cachectic therapies for many clinical situations. @ Inra/Elsevier, Paris skeletal muscle / protein breakdown

Paris-Sud XI, Université de

120

SIAH-1 interacts with CtIP and promotes its degradation by the proteasome pathway.  

PubMed

SIAH-1 and SIAH-2 are the human members of an evolutionary highly conserved E3 ligase family. SIAH-1 is a p53 and p21(Waf-1/Cip-1) induced gene during apoptosis and tumor suppression. In stable-transfected clones of MCF-7 cells, SIAH-1 overexpression was associated with apoptosis, mitotic alterations and p21(Waf-1/Cip-1) induction of expression. Using a two-hybrid screening, we identified here the transcriptional corepressor CtBP-interacting protein (CtIP) as a SIAH-1-interacting protein. CtIP has been proposed as a regulator of p21(Waf-1/Cip-1) gene transcription through a protein complex involving BRCA1. We demonstrate that SIAH-1 associates with CtIP both in vitro and in vivo. This interaction led to CtIP degradation by the ubiquitin-proteasome pathway. As expected, SIAH-1 induced p21(Waf-1/Cip-1) transcription in Jurkat-T cell. Surprisingly, a SIAH protein deleted of its RING finger, SIAH-1DeltaN, which is able to interact with CtIP but does not promote its degradation, also induced transcription from the p21(Waf-1) promoter in a similar extent as did SIAH-1. Our results suggest that p21(Waf-1/Cip-1) induction by SIAH-1 could not be mediated by CtIP degradation. PMID:14654780

Germani, Antonia; Prabel, Audrey; Mourah, Samia; Podgorniak, Marie-Pierre; Di Carlo, Anna; Ehrlich, Ricardo; Gisselbrecht, Sylvie; Varin-Blank, Nadine; Calvo, Fabien; Bruzzoni-Giovanelli, Heriberto

2003-12-01

121

Importance of the regulation of nuclear receptor degradation.  

PubMed

Nuclear hormone receptors (NHRs) represent a superfamily of structurally related ligand-activated transcription factors, which regulate diverse biological activities like growth, development, and homeostasis. Recently, it has been demonstrated that certain members of the NHR superfamily are degraded through the ubiquitin-proteasome pathway in a ligand-dependent manner. Though the signal for the down-regulation via the ubiquitin-proteasome pathway is not yet known, phosphorylation at specific amino acid residues or coactivator binding to receptors could lead to their degradation by the 26S proteasome. Activation and degradation seems to be an engineered cyclic mechanism, which provides tight control over diverse cellular processes. The degradation process involves extensive loss of proteins and requires expenditure of cellular ATP. That seems to be inevitable for a more important aim, that is efficient and appropriate regulation of transcription. Down-regulation of receptors would lead to an attenuated transcriptional response because the number of receptor molecules available to activate transcription would decrease over time. One of the obvious reasons for down-regulating NHRs thus seems to be to prevent the cell from overstimulation by the hormones or other activating signals. Nuclear receptor turnover may also reset the transcriptional apparatus in preparation for a subsequent response. Since inhibition of the ubiquitin-proteasome degradation pathway disturbs the transcriptional activitity of some of the nuclear receptors such as estrogen (ER) and progesterone (PR) receptors, it is also possible that the degradation of NHRs may enable recycling of components of receptor-cofactor complexes and general transcriptional machinary. Understanding the mechanism of nuclear hormone receptor degradation and its relation to transcription may lead to novel insights of therapuetic intervention. PMID:11487464

Dennis, A P; Haq, R U; Nawaz, Z

2001-08-01

122

The Ubiquitin–Proteasome System of Saccharomyces cerevisiae  

PubMed Central

Protein modifications provide cells with exquisite temporal and spatial control of protein function. Ubiquitin is among the most important modifiers, serving both to target hundreds of proteins for rapid degradation by the proteasome, and as a dynamic signaling agent that regulates the function of covalently bound proteins. The diverse effects of ubiquitylation reflect the assembly of structurally distinct ubiquitin chains on target proteins. The resulting ubiquitin code is interpreted by an extensive family of ubiquitin receptors. Here we review the components of this regulatory network and its effects throughout the cell. PMID:23028185

Finley, Daniel; Ulrich, Helle D.; Sommer, Thomas; Kaiser, Peter

2012-01-01

123

Activation of the ubiquitin-proteasome system against arsenic trioxide cardiotoxicity involves ubiquitin ligase Parkin for mitochondrial homeostasis.  

PubMed

Parkin is an E3 ubiquitin ligase involved in the elimination of damaged mitochondria. Ubiquitination of mitochondrial substrates by Parkin results in proteasomal as well as lysosomal degradation of mitochondria, the latter of which is executed by the autophagy machinery and is called as mitophagy (mitochondrial autophagy). The aim of this study is to examine the possible role of Parkin against cardiotoxicity elicited by arsenic trioxide (ATO) exposure in HL-1 mouse atrial cardiomyocytes. HL-1 cells were administered 1-10?M ATO for up to 24h, and the involvements of apoptosis, and the ubiquitin-proteasome and autophagy-lysosome systems (UPS and ALS) were examined. ATO dose-dependently reduced mitochondrial membrane potentials (??m) in HL-1 cells, indicating that ATO works as a mitochondrial toxin in these cells. Apoptosis was evident in cells exposed to more than 6?M ATO for 24h. Levels of Parkin in mitochondria-rich fractions were increased, suggesting the recruitment of Parkin to mitochondria. Ubiquitination of the voltage-dependent anion channel1 (VDAC1), a substrate of Parkin, was also proved by immunoprecipitation. Accumulation of ubiquitinated proteins including both K48- and K63-lineages was observed in HL-1 cells after ATO exposure, implying an increased demand for proteasomal as well as lysosomal degradation of cellular proteins. Although UPS was activated by ATO as proved by increased proteasomal activity, only slight activation of the ALS marker LC3 was observed, suggesting differential reactions of UPS and ALS to ATO toxicity. The abrogation of UPS by the proteasome inhibitor bortezomib significantly sensitized HL-1 cells to ATO toxicity, showing the contribution of UPS to the maintenance of cellular homeostasis during ATO exposure. Taken together, our results reveal the activation of Parkin as well as UPS during ATO exposure in HL-1 cardiomyocytes, which contributes to the maintenance of mitochondrial as well as cellular homeostasis. PMID:24801902

Watanabe, Mayumi; Funakoshi, Takeshi; Unuma, Kana; Aki, Toshihiko; Uemura, Koichi

2014-08-01

124

Unfolded protein response and activated degradative pathways regulation in GNE myopathy.  

PubMed

Although intracellular beta amyloid (A?) accumulation is known as an early upstream event in the degenerative course of UDP-N-acetylglucosamine 2-epimerase/N-acetylmannosamine kinase (GNE) myopathy, the process by which A?deposits initiate various degradative pathways, and their relationship have not been fully clarified. We studied the possible secondary responses after amyloid beta precursor protein (A?PP) deposition including unfolded protein response (UPR), ubiquitin proteasome system (UPS) activation and its correlation with autophagy system. Eight GNE myopathy patients and five individuals with normal muscle morphology were included in this study. We performed immunofluorescence and immunoblotting to investigate the expression of A?PP, phosphorylated tau (p-tau) and endoplasmic reticulum molecular chaperones. Proteasome activities were measured by cleavage of fluorogenic substrates. The expression of proteasome subunits and linkers between proteasomal and autophagy systems were also evaluated by immunoblotting and relative quantitative real-time RT-PCR. Four molecular chaperones, glucose-regulated protein 94 (GRP94), glucose-regulated protein 78 (GRP78), calreticulin and calnexin and valosin containing protein (VCP) were highly expressed in GNE myopathy. 20S proteasome subunits, three main proteasome proteolytic activities, and the factors linking UPS and autophagy system were also increased. Our study suggests that A?PP deposition results in endoplasmic reticulum stress (ERS) and highly expressed VCP deliver unfolded proteins from endoplasmic reticulum to proteosomal system which is activated in endoplasmic reticulum associated degradation (ERAD) in GNE myopathy. Excessive ubiquitinated unfolded proteins are exported by proteins that connect UPS and autophagy to autophagy system, which is activated as an alternative pathway for degradation. PMID:23472144

Li, Honghao; Chen, Qi; Liu, Fuchen; Zhang, Xuemei; Li, Wei; Liu, Shuping; Zhao, Yuying; Gong, Yaoqin; Yan, Chuanzhu

2013-01-01

125

Contribution of the autophagy-lysosomal and ubiquitin-proteasomal proteolytic systems to total proteolysis in rainbow trout (Oncorhynchus mykiss) myotubes.  

PubMed

The ubiquitin-proteasome system (UPS) is recognized as the major contributor to total proteolysis in mammalian skeletal muscle, responsible for 50% or more of total protein degradation in skeletal muscle, whereas the autophagic-lysosome system (ALS) plays a more minor role. While the relative contribution of these systems to muscle loss is well documented in mammals, little is known in fish species. The current study uses myotubes derived from rainbow trout myogenic precursor cells as an in vitro model of white muscle tissue. Cells were incubated in complete or serum-deprived media or media supplemented with insulin-like growth factor-1 (IGF-1) and exposed to selective proteolytic inhibitors to determine the relative contribution of the ALS and UPS to total protein degradation in myotubes in different culture conditions. Results indicate that the ALS is responsible for 30-34% and 50% of total protein degradation in myotubes in complete and serum-deprived media, respectively. The UPS appears to contribute much less to total protein degradation at almost 4% in cells in complete media to nearly 17% in serum-deprived cells. IGF-1 decreases activity of both systems, as it inhibited the upregulation of both proteolytic systems induced by serum deprivation. The combined inhibition of both the ALS and UPS reduced degradation by a maximum of 55% in serum-deprived cells, suggesting an important contribution of other proteolytic systems to total protein degradation. Collectively, these data identify the ALS as a potential target for strategies aimed at improving muscle protein retention and fillet yield through reductions in protein degradation. PMID:25274907

Seiliez, Iban; Dias, Karine; Cleveland, Beth M

2014-12-01

126

Cytokine-induced oxidative stress in cardiac inflammation and heart failure-how the ubiquitin proteasome system targets this vicious cycle.  

PubMed

The ubiquitin proteasome system (UPS) is critical for the regulation of many intracellular processes necessary for cell function and survival. The absolute requirement of the UPS for the maintenance of protein homeostasis and thereby for the regulation of protein quality control is reflected by the fact that deviation of proteasome function from the norm was reported in cardiovascular pathologies. Inflammation is a major factor contributing to cardiac pathology. Herein, cytokines induce protein translation and the production of free radicals, thereby challenging the cellular protein equilibrium. Here, we discuss current knowledge on the mechanisms of UPS-functional adaptation in response to oxidative stress in cardiac inflammation. The increasing pool of oxidant-damaged degradation-prone proteins in cardiac pathology accounts for the need for enhanced protein turnover by the UPS. This process is accomplished by an up-regulation of the ubiquitylation machinery and the induction of immunoproteasomes. Thereby, the inflamed heart muscle is cleared from accumulating misfolded proteins. Current advances on immunoproteasome-specific inhibitors in this field question the impact of the proteasome as a therapeutic target in heart failure. PMID:23508734

Voigt, Antje; Rahnefeld, Anna; Kloetzel, Peter M; Krüger, Elke

2013-01-01

127

Degradation of Postsynaptic Scaffold GKAP and Regulation of Dendritic Spine Morphology by the TRIM[subscript 3] Ubiquitin Ligase in Rat Hippocampal Neurons  

E-print Network

Changes in neuronal activity modify the structure of dendritic spines and alter the function and protein composition of synapses. Regulated degradation of postsynaptic density (PSD) proteins by the ubiquitin-proteasome ...

Sheng, Morgan Hwa-Tze

128

Selective histone deacetylase (HDAC) inhibition imparts beneficial effects in Huntington's disease mice: implications for the ubiquitin–proteasomal and autophagy systems  

PubMed Central

We previously demonstrated that the histone deacetylase (HDAC) inhibitor, 4b, which preferentially targets HDAC1 and HDAC3, ameliorates Huntington's disease (HD)-related phenotypes in different HD model systems. In the current study, we investigated extensive behavioral and biological effects of 4b in N171-82Q transgenic mice and further explored potential molecular mechanisms of 4b action. We found that 4b significantly prevented body weight loss, improved several parameters of motor function and ameliorated Huntingtin (Htt)-elicited cognitive decline in N171-82Q transgenic mice. Pathways analysis of microarray data from the mouse brain revealed gene networks involving post-translational modification, including protein phosphorylation and ubiquitination pathways, associated with 4b drug treatment. Using real-time qPCR analysis, we validated differential regulation of several genes in these pathways by 4b, including Ube2K, Ubqln, Ube2e3, Usp28 and Sumo2, as well as several other related genes. Additionally, 4b elicited increases in the expression of genes encoding components of the inhibitor of kappaB kinase (IKK) complex. IKK activation has been linked to phosphorylation, acetylation and clearance of the Htt protein by the proteasome and the lysosome, and accordingly, we found elevated levels of phosphorylated endogenous wild-type (wt) Htt protein at serine 16 and threonine 3, and increased AcK9/pS13/pS16 immunoreactivity in cortical samples from 4b-treated mice. We further show that HDAC inhibitors prevent the formation of nuclear Htt aggregates in the brains of N171-82Q mice. Our findings suggest that one mechanism of 4b action is associated with the modulation of the ubiquitin–proteasomal and autophagy pathways, which could affect accumulation, stability and/or clearance of important disease-related proteins, such as Htt. PMID:22965876

Jia, Haiqun; Kast, Ryan J.; Steffan, Joan S.; Thomas, Elizabeth A.

2012-01-01

129

The Ubiquitin-Proteasome Reporter GFPu Does Not Accumulate in Neurons of the R6/2 Transgenic Mouse Model of Huntington's Disease  

PubMed Central

Impairment of the ubiquitin-proteasome system (UPS) has long been considered an attractive hypothesis to explain the selective dysfunction and death of neurons in polyglutamine disorders such as Huntington's disease (HD). The fact that inclusion bodies in HD mouse models and patient brains are rich in ubiquitin and proteasome components suggests that the UPS may be hindered directly or indirectly by inclusion bodies or their misfolded monomeric or oligomeric precursors. However, studies into UPS function in various polyglutamine disease models have yielded conflicting results, suggesting mutant polyglutamine tracts may exert different effects on the UPS depending on protein context, expression level, subcellular localisation and cell-type. To investigate UPS function in a well-characterised mouse model of HD, we have crossed R6/2 HD mice with transgenic UPS reporter mice expressing the GFPu construct. The GFPu construct comprises GFP fused to a constitutive degradation signal (CL-1) that promotes its rapid degradation under conditions of a healthy UPS. Using a combination of immunoblot analysis, fluorescence and immunofluorescence microscopy studies, we found that steady-state GFPu levels were not detectably different between R6/2 and non-R6/2 brain. We observed no correlation between inclusion body formation and GFPu accumulation, suggesting no direct relationship between protein aggregation and global UPS inhibition in R6/2 mice. These findings suggest that while certain branches of the UPS can be impaired by mutant polyglutamine proteins, such proteins do not necessarily cause total blockade of UPS-dependent degradation. It is therefore likely that the relationship between mutant polyglutamine proteins and the UPS is more complex than originally anticipated. PMID:19352500

Bett, John S.; Cook, Casey; Petrucelli, Leonard; Bates, Gillian P.

2009-01-01

130

Fluorene Degradation Pathway Map  

NSDL National Science Digital Library

The University of Minnesota Biocatalysis/Biodegradation Database (UM/BBD) contains "microbial biocatalytic reactions and biodegradation pathways primarily for xenobiotic, chemical compounds." A new microbial enzyme-catalyzed reactions has been posted for Flourene. Flourene is commonly found in vehicle exhaust emissions, motor oils, crude oils, coal and oil combustion products, industrial effluents, and waste incineration, and is a major component of "fossil fuels and their derivatives." The Fluorene pathway map includes organisms which initiate the pathway, as well as organisms that carry out later steps. The pathway map provides detailed information on each hyperlinked step, including graphics, product/substrate reactions, and external links to further information.

Feng, Jingfeng.

131

The contribution of the autophagy-lysosomal and ubiquitin-proteasomal proteolytic systems to total proteolysis in rainbow trout (Oncorhynchus mykiss) myotubes  

Technology Transfer Automated Retrieval System (TEKTRAN)

Two major proteolytic systems are thought to (co-) operate in the skeletal muscle of vertebrates, the ubiquitin-proteasomal system (UPS) and the autophagic/lysosomal system (ALS). While their relative contribution to muscle loss has been already well documented in mammals, little is known in fish sp...

132

cAMP STIMULATES THE UBIQUITIN/PROTEASOME PATHWAY IN RAT SPINAL CORD NEURONS  

PubMed Central

Proteasome impairment and accumulation of ubiquitinated proteins are implicated in neurodegeneration associated with different forms of spinal cord injury. We show herein that elevating cAMP in rat spinal cord neurons increases 26S proteasome activity in a protein kinase A-dependent manner. Treating spinal cord neurons with dibutyryl-cAMP (db-cAMP) also raised the levels of various components of the UPP including proteasome subunits Rpt6 and ?5, polyubiquitin shuttling factor p62/sequestosome1, E3 ligase CHIP, AAA-ATPase p97 and the ubiquitin gene ubB. Finally, db-cAMP reduced the accumulation of ubiquitinated proteins, proteasome inhibition, and neurotoxicity triggered by the endogenous product of inflammation prostaglandin J2. We propose that optimizing the effects of cAMP/PKA-signaling on the UPP could offer an effective therapeutic approach to prevent UPP-related proteotoxicity in spinal cord neurons. PMID:22982149

Myeku, Natura; Wang, Hu; Figueiredo-Pereira, Maria E.

2012-01-01

133

Hepatic Deficiency of COP9 Signalosome Subunit 8 Induces Ubiquitin-Proteasome System Impairment and Bim-Mediated Apoptosis in Murine Livers  

PubMed Central

The COP9 signalosome (CSN), an evolutionally highly conserved protein complex composed of 8 unique subunits (CSN1 through CSN8) in higher eukaryotes, is purported to modulate protein degradation mediated by the ubiquitin-proteasome system (UPS) but this has not been demonstrated in a critical mitotic parenchymal organ of vertebrates. Hepatocyte-specific knockout of the Cops8 gene (HS-Csn8KO) was shown to cause massive hepatocyte apoptosis and liver malfunction but the underlying mechanism remains unclear. Here, we report that Csn8/CSN exerts profound impacts on hepatic UPS function and is critical to the stability of the pro-apoptotic protein Bim. Significant decreases in CIS (cytokine-inducible Src homology 2 domain-containing protein), a Bim receptor of a cullin2-based ubiquitin ligase, were found to co-exist with a marked increase of Bim proteins. Csn8 deficiency also significantly decreased 19S proteasome subunit Rpt5 and markedly increased high molecular weight neddylated and ubiquitinated proteins. The use of a surrogate UPS substrate further reveals severe impairment of UPS-mediated proteolysis in HS-Csn8KO livers. Inclusion body-like materials were accumulated in Csn8 deficient hepatocytes. In addition to Bim, massive hepatocyte apoptosis in HS-Csn8KO livers is also associated with elevated expression of other members of the Bcl2 family, including pro-apoptotic Bax as well as anti-apoptotic Bcl2 and Bcl-XL. Increased interaction between Bcl2 and Bim, but not between Bcl2 and Bax, was detected. Hence, it is concluded that hepatic CSN8 deficiency impairs the UPS in the liver and the resultant Bim upregulation likely plays an important role in triggering hepatocyte apoptosis via sequestering Bcl2 away from Bax. PMID:23840878

Lei, Daoxiong; Li, Faqian; Su, Huabo; Liu, Jinbao; Wei, Ning; Wang, Xuejun

2013-01-01

134

Bee venom effects on ubiquitin proteasome system in hSOD1G85R-expressing NSC34 motor neuron cells  

PubMed Central

Background Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disease that results from a progressive loss of motor neurons. Familial ALS (fALS) is caused by missense mutations in Cu, Zn-superoxide dismutase 1 (SOD1) that frequently result in the accumulation of mutant protein aggregates that are associated with impairments in the ubiquitin-proteasome system (UPS). UPS impairment has been implicated in many neurological disorders. Bee venom (BV) extracted from honey bees has been used as a traditional medicine for treating inflammatory diseases and has been shown to attenuate the neuroinflammatory events that occur in a symptomatic ALS animal model. Methods NSC34 cells were transiently transfected with a WT or G85R hSOD1-GFP construct for 24 hrs and then stimulated with 2.5 ?g/ml BV for 24 hrs. To determine whether a SOD1 mutation affects UPS function in NSC34 cells, we examined proteasome activity and performed western blotting and immunofluorescence using specific antibodies, such as anti-misfolded SOD1, anti-ubiquitin, anti-GRP78, anti-LC3, and anti-ISG15 antibodies. Results We found that GFP-hSOD1G85R overexpression induced SOD1 inclusions and reduced proteasome activity compared with the overexpression of GFP alone in NSC34 motor neuronal cells. In addition, we also observed that BV treatment restored proteasome activity and reduced the accumulation of ubiquitinated and misfolded SOD1 in GFP-hSOD1G85R-overexpressing NSC34 motor neuronal cells. However, BV treatment did not activate the autophagic pathway in these cells. Conclusion Our findings suggest that BV may rescue the impairment of the UPS in ALS models. PMID:23866691

2013-01-01

135

Aerobic exercise training improves oxidative stress and ubiquitin proteasome system activity in heart of spontaneously hypertensive rats.  

PubMed

The activity of the ubiquitin proteasome system (UPS) and the level of oxidative stress contribute to the transition from compensated cardiac hypertrophy to heart failure in hypertension. Moreover, aerobic exercise training (AET) is an important therapy for the treatment of hypertension, but its effects on the UPS are not completely known. The aim of this study was to evaluate the effect of AET on UPS's activity and oxidative stress level in heart of spontaneously hypertensive rats (SHR). A total of 53 Wistar and SHR rats were randomly divided into sedentary and trained groups. The AET protocol was 5×/week in treadmill for 13 weeks. Exercise tolerance test, non-invasive blood pressure measurement, echocardiographic analyses, and left ventricle hemodynamics were performed during experimental period. The expression of ubiquitinated proteins, 4-hydroxynonenal (4-HNE), Akt, phospho-Akt(ser473), GSK3?, and phospho-GSK3?(ser9) were analyzed by western blotting. The evaluation of lipid hydroperoxide concentration was performed using the xylenol orange method, and the proteasomal chymotrypsin-like activity was measured by fluorimetric assay. Sedentary hypertensive group presented cardiac hypertrophy, unaltered expression of total Akt, phospho-Akt, total GSK3? and phospho-GSK3?, UPS hyperactivity, increased lipid hydroperoxidation as well as elevated expression of 4-HNE but normal cardiac function. In contrast, AET significantly increased exercise tolerance, decreased resting systolic blood pressure and heart rate in hypertensive animals. In addition, the AET increased phospho-Akt expression, decreased phospho-GSK3?, and did not alter the expression of total Akt, total GSK3?, and ubiquitinated proteins, however, significantly attenuated 4-HNE levels, lipid hydroperoxidation, and UPS's activity toward normotensive group levels. Our results provide evidence for the main effect of AET on attenuating cardiac ubiquitin proteasome hyperactivity and oxidative stress in SHR rats. PMID:25626892

de Andrade, Luiz Henrique Soares; de Moraes, Wilson Max Almeida Monteiro; Matsuo Junior, Eduardo Hiroshi; de Orleans Carvalho de Moura, Elizabeth; Antunes, Hanna Karen Moreira; Montemor, Jairo; Antonio, Ednei Luiz; Bocalini, Danilo Sales; Serra, Andrey Jorge; Tucci, Paulo José Ferreira; Brum, Patricia Chakur; Medeiros, Alessandra

2015-04-01

136

The Proteasome Inhibitor, MG132, Attenuates Diabetic Nephropathy by Inhibiting SnoN Degradation In Vivo and In Vitro  

PubMed Central

Transforming growth factor-? (TGF-?) has been shown to be involved in diabetic nephropathy (DN). The SnoN protein can regulate TGF-? signaling through interaction with Smad proteins. Recent studies have shown that SnoN is mainly degraded by the ubiquitin-proteasome pathway. However, the role of SnoN in the regulation of TGF-?/Smad signaling in DN is still unclear. In this study, diabetic rats were randomly divided into a diabetic control group (DC group) and a proteasome inhibitor (MG132) diabetes therapy group (DT group). Kidney damage parameters and the expression of SnoN, Smurf2, and TGF-? were observed. Simultaneously, we cultured rat glomerular mesangial cells (GMCs) stimulated with high glucose, and SnoN and Arkadia expression were measured. Results demonstrated that 24-hour urine protein, ACR, BUN, and the expression of Smurf2 and TGF-? were significantly increased (P < 0.05), whereas SnoN was significantly decreased in the DC group (P < 0.05). However, these changes diminished after treatment with MG132. SnoN expression in GMCs decreased significantly (P < 0.05), but Arkadia expression gradually increased due to high glucose stimulation (P < 0.05), which could be almost completely reversed by MG132 (P < 0.05). The present results support the hypothesis that MG132 may alleviate kidney damage by inhibiting SnoN degradation and TGF-? activation, suggesting that the ubiquitin-proteasome pathway may become a new therapeutic target for DN. PMID:25003128

Huang, Wei; Yang, Chen; Nan, Qinling; Gao, Chenlin; Feng, Hong; Gou, Fang; Chen, Guo; Zhang, Zhihong; Yan, Pijun; Peng, Juan

2014-01-01

137

The F-box Protein FBXO25 Promotes the Proteasome-dependent Degradation of ELK-1 Protein*  

PubMed Central

FBXO25 is one of the 69 known human F-box proteins that serve as specificity factors for a family of ubiquitin ligases composed of SKP1, Rbx1, Cullin1, and F-box protein (SCF1) that are involved in targeting proteins for degradation across the ubiquitin proteasome system. However, the substrates of most SCF E3 ligases remain unknown. Here, we applied an in chip ubiquitination screen using a human protein microarray to uncover putative substrates for the FBXO25 protein. Among several novel putative targets identified, the c-fos protooncogene regulator ELK-1 was characterized as the first endogenous substrate for SCF1(FBXO25) E3 ligase. FBXO25 interacted with and mediated the ubiquitination and proteasomal degradation of ELK-1 in HEK293T cells. In addition, FBXO25 overexpression suppressed induction of two ELK-1 target genes, c-fos and egr-1, in response to phorbol 12-myristate 13-acetate. Together, our findings show that FBXO25 mediates ELK-1 degradation through the ubiquitin proteasome system and thereby plays a role in regulating the activation of ELK-1 pathway in response to mitogens. PMID:23940030

Teixeira, Felipe R.; Manfiolli, Adriana O.; Soares, Cláudia S.; Baqui, Munira M. A.; Koide, Tie; Gomes, Marcelo D.

2013-01-01

138

Recent technical developments in the study of ER-associated degradation.  

PubMed

Endoplasmic reticulum-associated degradation (ERAD) is a mechanism during which native and misfolded proteins are recognized and retrotranslocated across the ER membrane to the cytosol for degradation by the ubiquitin-proteasome system. Like other cellular pathways, the factors required for ERAD have been analyzed using both conventional genetic and biochemical approaches. More recently, however, an integrated top-down approach has identified a functional network that underlies the ERAD system. In turn, bottom-up reconstitution has become increasingly sophisticated and elucidated the molecular mechanisms underlying substrate recognition, ubiquitylation, retrotranslocation, and degradation. In addition, a live cell imaging technique and a site-specific in vivo photo-crosslinking approach have further dissected specific steps during ERAD. These technical developments have revealed an unexpected dynamicity of the membrane-associated ERAD complex. In this article, we will discuss how these technical developments have improved our understanding of the ERAD pathway and have led to new questions. PMID:24867671

Nakatsukasa, Kunio; Kamura, Takumi; Brodsky, Jeffrey L

2014-08-01

139

Photovoltaic Lifetime & Degradation Science Statistical Pathway Development: Acrylic Degradation  

E-print Network

Photovoltaic Lifetime & Degradation Science Statistical Pathway Development: Acrylic Degradation, USA ABSTRACT In order to optimize and extend the life of photovoltaics (PV) modules, scientific photovoltaics. The statisti- cally significant relationships were investigated using lifetime and degradation

Rollins, Andrew M.

140

Melatonin-Induced Temporal Up-Regulation of Gene Expression Related to Ubiquitin/Proteasome System (UPS) in the Human Malaria Parasite Plasmodium falciparum  

PubMed Central

There is an increasing understanding that melatonin and the ubiquitin/proteasome system (UPS) interact to regulate multiple cellular functions. Post-translational modifications such as ubiquitination are important modulators of signaling processes, cell cycle and many other cellular functions. Previously, we reported a melatonin-induced upregulation of gene expression related to ubiquitin/proteasome system (UPS) in Plasmodium falciparum, the human malaria parasite, and that P. falciparum protein kinase 7 influences this process. This implies a role of melatonin, an indolamine, in modulating intraerythrocytic development of the parasite. In this report we demonstrate by qPCR analysis, that melatonin induces gene upregulation in nine out of fourteen genes of the UPS, consisting of the same set of genes previously reported, between 4 to 5 h after melatonin treatment. We demonstrate that melatonin causes a temporally controlled gene expression of UPS members. PMID:25479077

Koyama, Fernanda C.; Azevedo, Mauro F.; Budu, Alexandre; Chakrabarti, Debopam; Garcia, Célia R. S.

2014-01-01

141

Degradation signals recognized by the Ubc6p-Ubc7p ubiquitin-conjugating enzyme pair.  

PubMed

Proteolysis by the ubiquitin-proteasome system is highly selective. Specificity is achieved by the cooperation of diverse ubiquitin-conjugating enzymes (Ubcs or E2s) with a variety of ubiquitin ligases (E3s) and other ancillary factors. These recognize degradation signals characteristic of their target proteins. In a previous investigation, we identified signals directing the degradation of beta-galactosidase and Ura3p fusion proteins via a subsidiary pathway of the ubiquitin-proteasome system involving Ubc6p and Ubc7p. This pathway has recently been shown to be essential for the degradation of misfolded and regulated proteins in the endoplasmic reticulum (ER) lumen and membrane, which are transported to the cytoplasm via the Sec61p translocon. Mutant backgrounds which prevent retrograde transport of ER proteins (hrd1/der3Delta and sec61-2) did not inhibit the degradation of the beta-galactosidase and Ura3p fusions carrying Ubc6p/Ubc7p pathway signals. We therefore conclude that the ubiquitination of these fusion proteins takes place on the cytosolic face of the ER without prior transfer to the ER lumen. The contributions of different sequence elements to a 16-amino-acid-residue Ubc6p-Ubc7p-specific signal were analyzed by mutation. A patch of bulky hydrophobic residues was an essential element. In addition, positively charged residues were found to be essential. Unexpectedly, certain substitutions of bulky hydrophobic or positively charged residues with alanine created novel degradation signals, channeling the degradation of fusion proteins to an unidentified proteasomal pathway not involving Ubc6p and Ubc7p. PMID:10982838

Gilon, T; Chomsky, O; Kulka, R G

2000-10-01

142

Degradation Signals Recognized by the Ubc6p-Ubc7p Ubiquitin-Conjugating Enzyme Pair  

PubMed Central

Proteolysis by the ubiquitin-proteasome system is highly selective. Specificity is achieved by the cooperation of diverse ubiquitin-conjugating enzymes (Ubcs or E2s) with a variety of ubiquitin ligases (E3s) and other ancillary factors. These recognize degradation signals characteristic of their target proteins. In a previous investigation, we identified signals directing the degradation of ?-galactosidase and Ura3p fusion proteins via a subsidiary pathway of the ubiquitin-proteasome system involving Ubc6p and Ubc7p. This pathway has recently been shown to be essential for the degradation of misfolded and regulated proteins in the endoplasmic reticulum (ER) lumen and membrane, which are transported to the cytoplasm via the Sec61p translocon. Mutant backgrounds which prevent retrograde transport of ER proteins (hrd1/der3? and sec61-2) did not inhibit the degradation of the ?-galactosidase and Ura3p fusions carrying Ubc6p/Ubc7p pathway signals. We therefore conclude that the ubiquitination of these fusion proteins takes place on the cytosolic face of the ER without prior transfer to the ER lumen. The contributions of different sequence elements to a 16-amino-acid-residue Ubc6p-Ubc7p-specific signal were analyzed by mutation. A patch of bulky hydrophobic residues was an essential element. In addition, positively charged residues were found to be essential. Unexpectedly, certain substitutions of bulky hydrophobic or positively charged residues with alanine created novel degradation signals, channeling the degradation of fusion proteins to an unidentified proteasomal pathway not involving Ubc6p and Ubc7p. PMID:10982838

Gilon, Tamar; Chomsky, Orna; Kulka, Richard G.

2000-01-01

143

Role of the ubiquitin-proteasome system in cardiac dysfunction of adipose triglyceride lipase-deficient mice.  

PubMed

Systemic deletion of the gene encoding for adipose triglyceride lipase (ATGL) in mice leads to severe cardiac dysfunction due to massive accumulation of neutral lipids in cardiomyocytes. Recently, impaired peroxisome proliferator-activated receptor ? (PPAR?) signaling has been described to substantially contribute to the observed cardiac phenotype. Disturbances of the ubiquitin-proteasome system (UPS) have been implicated in numerous cardiac diseases including cardiomyopathy, ischemic heart disease, and heart failure. The objective of the present study was to investigate the potential role of UPS in cardiac ATGL deficiency. Our results demonstrate prominent accumulation of ubiquitinated proteins in hearts of ATGL-deficient mice, an effect that was abolished upon cardiomyocyte-directed overexpression of ATGL. In parallel, cardiac protein expression of the ubiquitin-activating enzyme E1a, which catalyzes the first step of the ubiquitination cascade, was significantly upregulated in ATGL-deficient hearts. Dysfunction of the UPS was accompanied by activation of NF-?B signaling. Moreover, the endoplasmic reticulum (ER)-resident chaperon protein disulfide isomerase was significantly upregulated in ATGL-deficient hearts. Chronic treatment of ATGL-deficient mice with the PPAR? agonist Wy14,643 improved proteasomal function, prevented NF-?B activation and decreased oxidative stress. In summary, our data point to a hitherto unrecognized link between proteasomal function, PPAR? signaling and cardiovascular disease. PMID:25285770

Mussbacher, Marion; Stessel, Heike; Wölkart, Gerald; Haemmerle, Guenter; Zechner, Rudolf; Mayer, Bernd; Schrammel, Astrid

2014-12-01

144

Influence of electrical stimulation on calpain and ubiquitin-proteasome systems in the denervated and unloaded rat tibialis anterior muscles.  

PubMed

The influence of electrical stimulation on calpain and ubiquitin-proteasome systems was examined in the denervated and unloaded tibialis anterior muscles of male Wistar rats. Animals were divided into 5 groups: control, denervation, denervation plus electrical stimulation, unloading, and hindlimb unloading plus electrical stimulation groups. Due to denervation and unloading for 14 days, muscle atrophy markedly occurred in the denervated and unloading animals, and the atrophy in the former was significantly more severe than that in the latter. In the denervated muscle, the atrophy was significantly attenuated by the electrical stimulation, but not in the unloaded muscle. Overexpression of calpain-2 and ubiquitinated proteins was observed only in denervated muscles. In the unloaded animals, though the expression level of calpain-2 appeared to be slightly higher than that in the control, the expression level of ubiquitinated proteins was almost the same as that in the control. The overexpression of calpain-1, calpain-2, and ubiquitinated proteins in the denervated muscle was inhibited by the electrical stimulation. However, there was no difference in these expressions between the unloaded and unloaded plus electrical stimulation groups. The mechanism of the preventive effect of the electrical stimulation on muscle atrophy might differ between the denervated and unloaded muscles. PMID:24745757

Matsumoto, Ayaka; Fujita, Naoto; Arakawa, Takamitsu; Fujino, Hidemi; Miki, Akinori

2014-06-01

145

Inhibiting the ubiquitin-proteasome system leads to preferential accumulation of toxic N-terminal mutant huntingtin fragments.  

PubMed

An expanded polyglutamine (polyQ) domain in the N-terminal region of huntingtin (htt) causes misfolding and accumulation of htt in neuronal cells and the subsequent neurodegeneration of Huntington's disease (HD). Clearing the misfolded htt is critical for preventing neuropathology, and this process is mediated primarily by both the ubiquitin-proteasome system (UPS) and autophagy. Although overexpression of mutant htt can inhibit UPS activity in cultured cells, mutant htt does not inhibit global UPS activity in the brains of HD transgenic mice. These findings underscore the importance of investigating the function of the UPS and autophagy in the brain when mutant proteins are not overexpressed. When cultured PC12 cells were treated with either UPS or autophagy inhibitors, more N-terminal mutant htt fragments accumulated via inhibition of the UPS. Furthermore, in HD CAG repeat knock-in mouse brain, inhibiting the UPS also resulted in a greater accumulation of N-terminal, but not full-length, mutant htt than inhibiting autophagy did. Our findings suggest that impairment of the UPS may be more important for the accumulation of N-terminal mutant htt and might therefore make an attractive therapeutic target. PMID:20354076

Li, Xiang; Wang, Chuan-En; Huang, Shanshan; Xu, Xingshun; Li, Xiao-Jiang; Li, He; Li, Shihua

2010-06-15

146

The Ubiquitin-associated (UBA) 1 Domain of Schizosaccharomyces pombe Rhp23 Is Essential for the Recognition of Ubiquitin-proteasome System Substrates Both in Vitro and in Vivo*  

PubMed Central

The ubiquitin-proteasome system is essential for maintaining a functional cell. Not only does it remove incorrectly folded proteins, it also regulates protein levels to ensure their appropriate spatial and temporal distribution. Proteins marked for degradation by the addition of Lys48-linked ubiquitin (Ub) chains are recognized by shuttle factors and transported to the 26 S proteasome. One of these shuttle factors, Schizosaccharomyces pombe Rhp23, has an unusual domain architecture. It comprises an N-terminal ubiquitin-like domain that can recognize the proteasome followed by two ubiquitin-associated (UBA) domains, termed UBA1 and UBA2, which can bind Ub. This architecture is conserved up to humans, suggesting that both domains are important for Rhp23 function. Such an extent of conservation raises the question as to why, in contrast to all other shuttle proteins, does Rhp23 require two UBA domains? We performed in vitro Ub binding assays using domain swap chimeric proteins and mutated domains in isolation as well as in the context of the full-length protein to reveal that the Ub binding properties of the UBA domains are context-dependent. In vivo, the internal Rhp23 UBA1 domain provides sufficient Ub recognition for the protein to function without UBA2. PMID:23038266

Medina, Bethan; Paraskevopoulos, Konstantinos; Boehringer, Jonas; Sznajder, Anna; Robertson, Morag; Endicott, Jane; Gordon, Colin

2012-01-01

147

The COP9 signalosome is required for light-dependent timeless degradation and Drosophila clock resetting.  

PubMed

The ubiquitin-proteasome system plays a major role in the rhythmic accumulation and turnover of molecular clock components. In turn, these approximately 24 h molecular rhythms drive circadian rhythms of behavior and physiology. In Drosophila, the ubiquitin-proteasome system also plays a critical role in light-dependent degradation of the clock protein Timeless (TIM), a key step in the entrainment of the molecular clocks to light-dark cycles. Here, we investigated the role of the COP9 signalosome (CSN), a general regulator of protein degradation, in fly circadian rhythms. We found that null mutations in the genes encoding the CSN4 and CSN5 subunits prevent normal TIM degradation by light in the pacemaker lateral neurons (LNs) as does LN-specific expression of a dominant-negative CSN5 transgene. These defects are accompanied by strong reductions in behavioral phase shifts of adult flies lacking normal CSN5 activity in LNs. Defects in TIM degradation and resetting of behavioral phases were rescued by overexpression of Jetlag (JET), the F-box protein required for light-mediated TIM degradation. Flies lacking normal CSN activity in all clock neurons are rhythmic in constant light, a phenotype previously associated with jet mutants. Together, these data indicate that JET and the CSN lie in a common pathway leading to light-dependent TIM degradation. Surprisingly, we found that manipulations that strongly inhibit CSN activity had minimal effects on circadian rhythms in constant darkness, indicating a specific role for the CSN in light-mediated TIM degradation. PMID:19176824

Knowles, Alyson; Koh, Kyunghee; Wu, June-Tai; Chien, Cheng-Ting; Chamovitz, Daniel A; Blau, Justin

2009-01-28

148

Aggresome-like structure induced by isothiocyanates is novel proteasome-dependent degradation machinery  

SciTech Connect

Unwanted or misfolded proteins are either refolded by chaperones or degraded by the ubiquitin-proteasome system (UPS). When UPS is impaired, misfolded proteins form aggregates, which are transported along microtubules by motor protein dynein towards the juxta-nuclear microtubule-organizing center to form aggresome, a single cellular garbage disposal complex. Because aggresome formation results from proteasome failure, aggresome components are degraded through the autophagy/lysosome pathway. Here we report that small molecule isothiocyanates (ITCs) can induce formation of aggresome-like structure (ALS) through covalent modification of cytoplasmic {alpha}- and {beta}-tubulin. The formation of ALS is related to neither proteasome inhibition nor oxidative stress. ITC-induced ALS is a proteasome-dependent assembly for emergent removal of misfolded proteins, suggesting that the cell may have a previously unknown strategy to cope with misfolded proteins.

Mi, Lixin, E-mail: lm293@georgetown.edu [Department of Oncology, Lombardi Comprehensive Cancer Center, Georgetown University, Washington, DC 20057 (United States)] [Department of Oncology, Lombardi Comprehensive Cancer Center, Georgetown University, Washington, DC 20057 (United States); Gan, Nanqin; Chung, Fung-Lung [Department of Oncology, Lombardi Comprehensive Cancer Center, Georgetown University, Washington, DC 20057 (United States)] [Department of Oncology, Lombardi Comprehensive Cancer Center, Georgetown University, Washington, DC 20057 (United States)

2009-10-16

149

A Sporadic Parkinson Disease Model via Silencing of the Ubiquitin-Proteasome/E3 Ligase Component SKP1A*  

PubMed Central

The aim of this study was to develop a new model of sporadic Parkinson disease (PD) based on silencing of the SKP1A gene, a component of the ubiquitin-proteasome/E3 ligase complex, Skp1, Cullin 1, F-box protein, which was found to be highly decreased in the substantia nigra of sporadic PD patients. Initially, an embryonic mouse substantia nigra-derived cell line (SN4741 cells) was infected with short hairpin RNA lentiviruses encoding the murine transcript of the SKP1A gene or with scrambled vector. SKP1A silencing resulted in increased susceptibility to neuronal damages induced by the parkinsonism-inducing neurotoxin 1-methyl-4-phenylpyridinium ion and serum starvation, in parallel with a decline in the expression of the dopaminergic markers, dopamine transporter and vesicular monoamine transporter-2. SKP1A-deficient cells presented a delay in completion of the cell cycle and the inability to arrest at the G0/G1 phase when induced to differentiate. Instead, the cells progressed through S phase, developing rounded aggregates with characteristics of aggresomes including immunoreactivity for ?-tubulin, ?-synuclein, ubiquitin, tyrosine hydroxylase, Hsc-70 (70-kDa heat shock cognate protein), and proteasome subunit, and culminating in a lethal phenotype. Conversely, stably enforced expression of wild type SKP1A duplicated the survival index of naïve SN4741 cells under proteasomal inhibition injury, suggesting a new structural role of SKP1 in dopaminergic neuronal function, besides its E3 ligase activity. These results link, for the first time, SKP1 to dopamine neuronal function and survival, suggesting an essential role in sporadic PD. In summary, this new model has reproduced to a significant extent the molecular alterations described in sporadic PD at the cellular level, implicating Skp1 as a potential modifier in sporadic PD neurodegeneration. PMID:19748892

Fishman-Jacob, Tali; Reznichenko, Lydia; Youdim, Moussa B. H.; Mandel, Silvia A.

2009-01-01

150

A sporadic Parkinson disease model via silencing of the ubiquitin-proteasome/E3 ligase component SKP1A.  

PubMed

The aim of this study was to develop a new model of sporadic Parkinson disease (PD) based on silencing of the SKP1A gene, a component of the ubiquitin-proteasome/E3 ligase complex, Skp1, Cullin 1, F-box protein, which was found to be highly decreased in the substantia nigra of sporadic PD patients. Initially, an embryonic mouse substantia nigra-derived cell line (SN4741 cells) was infected with short hairpin RNA lentiviruses encoding the murine transcript of the SKP1A gene or with scrambled vector. SKP1A silencing resulted in increased susceptibility to neuronal damages induced by the parkinsonism-inducing neurotoxin 1-methyl-4-phenylpyridinium ion and serum starvation, in parallel with a decline in the expression of the dopaminergic markers, dopamine transporter and vesicular monoamine transporter-2. SKP1A-deficient cells presented a delay in completion of the cell cycle and the inability to arrest at the G(0)/G(1) phase when induced to differentiate. Instead, the cells progressed through S phase, developing rounded aggregates with characteristics of aggresomes including immunoreactivity for gamma-tubulin, alpha-synuclein, ubiquitin, tyrosine hydroxylase, Hsc-70 (70-kDa heat shock cognate protein), and proteasome subunit, and culminating in a lethal phenotype. Conversely, stably enforced expression of wild type SKP1A duplicated the survival index of naïve SN4741 cells under proteasomal inhibition injury, suggesting a new structural role of SKP1 in dopaminergic neuronal function, besides its E3 ligase activity. These results link, for the first time, SKP1 to dopamine neuronal function and survival, suggesting an essential role in sporadic PD. In summary, this new model has reproduced to a significant extent the molecular alterations described in sporadic PD at the cellular level, implicating Skp1 as a potential modifier in sporadic PD neurodegeneration. PMID:19748892

Fishman-Jacob, Tali; Reznichenko, Lydia; Youdim, Moussa B H; Mandel, Silvia A

2009-11-20

151

Human cytomegalovirus UL76 elicits novel aggresome formation via interaction with S5a of the ubiquitin proteasome system.  

PubMed

HCMV UL76 is a member of a conserved Herpesviridae protein family (Herpes_UL24) that is involved in viral production, latency, and reactivation. UL76 presents as globular aggresomes in the nuclei of transiently transfected cells. Bioinformatic analyses predict that UL76 has a propensity for aggregation and targets cellular proteins implicated in protein folding and ubiquitin-proteasome systems (UPS). Furthermore, fluorescence recovery after photobleaching experiments suggests that UL76 reduces protein mobility in the aggresome, which indicates that UL76 elicits the aggregation of misfolded proteins. Moreover, in the absence of other viral proteins, UL76 interacts with S5a, which is a major receptor of polyubiquitinated proteins for UPS proteolysis via its conserved region and the von Willebrand factor type A (VWA) domain of S5a. We demonstrate that UL76 sequesters polyubiquitinated proteins and S5a to nuclear aggresomes in biological proximity. After knockdown of endogenous S5a by RNA interference techniques, the UL76 level was only minimally affected in transiently expressing cells. However, a significant reduction in the number of cells containing UL76 nuclear aggresomes was observed, which suggests that S5a may play a key role in aggresome formation. Moreover, we show that UL76 interacts with S5a in the late phase of viral infection and that knockdown of S5a hinders the development of both the replication compartment and the aggresome. In this study, we demonstrate that UL76 induces a novel nuclear aggresome, likely by subverting S5a of the UPS. Given that UL76 belongs to a conserved family, this underlying mechanism may be shared by all members of the Herpesviridae. PMID:23966401

Lin, Shin-Rung; Jiang, Meei Jyh; Wang, Hung-Hsueh; Hu, Cheng-Hui; Hsu, Ming-Shan; Hsi, Edward; Duh, Chang-Yih; Wang, Shang-Kwei

2013-11-01

152

The ubiquitin–proteasome system regulates p53-mediated transcription at p21waf1 promoter  

Microsoft Academic Search

The ubiquitin (Ub)–proteasome system (UPS) promotes the proteasomal degradation of target proteins by decorating them with Ub labels. Emerging evidence indicates a role of UPS in regulating gene transcription. In this study, we provided evidence for the involvement of UPS in the transcriptional activation function of tumor suppressor p53. We showed that both ubiquitylation and proteasomal functions are required for

Q Zhu; G Wani; J Yao; S Patnaik; Q-E Wang; M A El-Mahdy; M Prætorius-Ibba; A A Wani

2007-01-01

153

Histone deacetylase inhibitors prevent the degradation and restore the activity of glucocerebrosidase in Gaucher disease  

PubMed Central

Gaucher disease (GD) is caused by a spectrum of genetic mutations within the gene encoding the lysosomal enzyme glucocerebrosidase (GCase). These mutations often lead to misfolded proteins that are recognized by the unfolded protein response system and are degraded through the ubiquitin–proteasome pathway. Modulating this pathway with histone deacetylase inhibitors (HDACis) has been shown to improve protein stability in other disease settings. To identify the mechanisms involved in the regulation of GCase and determine the effects of HDACis on protein stability, we investigated the most prevalent mutations for nonneuronopathic (N370S) and neuronopathic (L444P) GD in cultured fibroblasts derived from GD patients and HeLa cells transfected with these mutations. The half-lives of mutant GCase proteins correspond to decreases in protein levels and enzymatic activity. GCase was found to bind to Hsp70, which directed the protein to TCP1 for proper folding, and to Hsp90, which directed the protein to the ubiquitin–proteasome pathway. Using a known HDACi (SAHA) and a unique small-molecule HDACi (LB-205), GCase levels increased rescuing enzymatic activity in mutant cells. The increase in the quantity of protein can be attributed to increases in protein half-life that correspond primarily with a decrease in degradation rather than an increase in chaperoned folding. HDACis reduce binding to Hsp90 and prevent subsequent ubiquitination and proteasomal degradation without affecting binding to Hsp70 or TCP1. These findings provide insight into the pathogenesis of GD and indicate a potent therapeutic potential of HDAC inhibitors for the treatment of GD and other human protein misfolding disorders. PMID:22160715

Lu, Jie; Yang, Chunzhang; Chen, Masako; Ye, Donald Y.; Lonser, Russell R.; Brady, Roscoe O.; Zhuang, Zhengping

2011-01-01

154

The N-end rule pathway: emerging functions and molecular principles of substrate recognition.  

PubMed

The N-end rule defines the protein-destabilizing activity of a given amino-terminal residue and its post-translational modification. Since its discovery 25 years ago, the pathway involved in the N-end rule has been thought to target only a limited set of specific substrates of the ubiquitin-proteasome system. Recent studies have provided insights into the components, substrates, functions and structural basis of substrate recognition. The N-end rule pathway is now emerging as a major cellular proteolytic system, in which the majority of proteins are born with or acquire specific N-terminal degradation determinants through protein-specific or global post-translational modifications. PMID:22016057

Sriram, Shashikanth M; Kim, Bo Yeon; Kwon, Yong Tae

2011-11-01

155

TNFR1-activated NF-?B signal transduction: regulation by the ubiquitin/proteasome system.  

PubMed

The Tumor Necrosis Factor Receptor-1 (TNFR1) is a central regulator of inflammation, cell death, and cellular proliferation. As such, alterations in TNFR1 signaling are associated with numerous diseases ranging from autoimmune syndromes to cancer. Understanding the regulation of TNFR1 signaling is therefore of considerable importance. The transduction of signaling events in the TNFR1 pathway - from ligand binding through transcriptional regulation - is regulated at nearly every step by post-translational modifications, including ubiquitination. In this review both endogenous and pharmacologic inhibitors of TNFR1 signaling, and how these impact the ubiquitin system, will be discussed. PMID:25461388

Wertz, Ingrid E

2014-11-12

156

Resveratrol promotes degradation of the human bile acid transporter ASBT (SLC10A2).  

PubMed

The sodium/bile acid co-transporter ASBT [apical sodium-dependent bile acid transporter; SLC10A2 (solute carrier family 10 member 2)] plays a key role in the enterohepatic recycling of the bile acids and indirectly contributes to cholesterol homoeostasis. ASBT inhibitors reportedly lower plasma triglyceride levels and increase HDL (high-density lipoprotein) cholesterol levels. RSV (resveratrol), a major constituent of red wine, is known to lower LDL (low-density lipoprotein) cholesterol levels, but its mechanism of action is still unclear. In the present study, we investigated the possible involvement of ASBT in RSV-mediated cholesterol-lowering effects. We demonstrate that RSV inhibits ASBT protein expression and function via a SIRT1 (sirtuin 1)-independent mechanism. The effect was specific to ASBT since other transporters involved in cholesterol homoeostasis, NTCP (SLC10A1), OST? (SLC51A) and ABCG1 (ATP-binding cassette G1), remained unaffected. ASBT inhibition by RSV was reversed by proteasome inhibitors (MG-132 and lactacystin) and the ubiquitin inhibitor LDN57444, suggesting involvement of the ubiquitin-proteasome pathway. Immunoprecipitation revealed high levels of ubiquitinated ASBT after RSV treatment. Phosphorylation at Ser335 and Thr339 was shown previously to play a role in proteosomal degradation of rat ASBT. However, mutation at corresponding residues in rat ASBT revealed that phosphorylation does not contribute to RSV-mediated degradation of ASBT. Combined, our data indicate that RSV promotes ASBT degradation via the ubiquitin-proteasome pathway without requiring phosphorylation. We conclude that regulation of ASBT expression by RSV may have clinical relevance with regard to the observed cholesterol-lowering effects of RSV. PMID:24498857

Chothe, Paresh P; Swaan, Peter W

2014-04-15

157

The ubiquitin-proteasome system regulates p53-mediated transcription at p21waf1 promoter.  

PubMed

The ubiquitin (Ub)-proteasome system (UPS) promotes the proteasomal degradation of target proteins by decorating them with Ub labels. Emerging evidence indicates a role of UPS in regulating gene transcription. In this study, we provided evidence for the involvement of UPS in the transcriptional activation function of tumor suppressor p53. We showed that both ubiquitylation and proteasomal functions are required for efficient transcription mediated by p53. Disruption of transcription by actinomycin D, 5,6-dichloro-1-beta-D-ribofuranosyl-benzimadazole or alpha-amanitin leads to accumulation of cellular p53 protein. Proteasome inhibition by MG132 increases the occupancy of p53 protein at p53-responsive p21(waf1) promoter. In addition, the Sug-1 component of 19S proteasome physically interacts with p53 in vitro and in vivo. Moreover, in response to ultraviolet-induced DNA damage, both the 19S proteasomal components, Sug1 and S1, are recruited to p21(waf1) promoter region in a kinetic pattern similar to that of p53. These results suggested that UPS positively regulates p53-mediated transcription at p21(waf1) promoter. PMID:17224908

Zhu, Q; Wani, G; Yao, J; Patnaik, S; Wang, Q-E; El-Mahdy, M A; Praetorius-Ibba, M; Wani, A A

2007-06-21

158

Pre-cachexia in patients with stages I-III non-small cell lung cancer: systemic inflammation and functional impairment without activation of skeletal muscle ubiquitin proteasome system.  

PubMed

Cachexia is a prevalent phenomenon of non-small cell lung cancer (NSCLC) which is responsible for increased mortality and deterioration of physical performance. Preclinical research indicates that systemic inflammation induces cachexia-related muscle wasting through muscular Nuclear Factor-kappa B (NF-?B) signaling and subsequent ubiquitin proteasome system (UPS)-mediated proteolysis. As these pathways could be a target for early intervention strategies, it needs to be elucidated whether increased activation of these pathways is already present in early stage NSCLC cachexia. The aim of the present study was therefore to assess muscular NF-?B and UPS activation in patients with NSCLC pre-cachexia. Sixteen patients with newly diagnosed stages I-III NSCLC having <10% weight loss and ten healthy controls were studied. Body composition, systemic inflammation and exercise capacity were assessed in all subjects and NF-?B and UPS activity in vastus lateralis muscle biopsies in a subset. Patients showed increased plasma levels of C-reactive protein (CRP) (P<0.001), soluble Tumor Necrosis Factor receptor 1 (sTNF-R1) (P<0.05), fibrinogen (P<0.001) and decreased levels of albumin (P<0.001). No changes in fat free body mass or skeletal muscle NF-?B and UPS activity were observed, while peak oxygen consumption ( [Formula: see text] ) was significantly decreased in patients compared with healthy controls. In conclusion, this exploratory study demonstrates significantly reduced exercise capacity in NSCLC pre-cachexia despite maintenance of muscle mass and unaltered indices of UPS activation. The absence of muscular NF-?B-dependent inflammatory signaling supports the notion that transition of systemic to local inflammation is required to initiate UPS-dependent muscle wasting characteristic for (experimental) cachexia. PMID:22018880

Op den Kamp, C M; Langen, R C; Minnaard, R; Kelders, M C; Snepvangers, F J; Hesselink, M K; Dingemans, A C; Schols, A M

2012-04-01

159

Systemic perturbation of the ERK signaling pathway by the proteasome inhibitor, MG132.  

PubMed

Inhibition of the ubiquitin-proteasome protein degradation pathway has been identified as a viable strategy for anti-tumor therapy based on its broad effects on cell proliferation. By the same token, the variety of elicited effects confounds the interpretation of cell-based experiments using proteasome inhibitors such as MG132. It has been proposed that MG132 treatment reduces growth factor-stimulated phosphorylation of extracellular signal-regulated kinases (ERKs), at least in part through upregulation of dual specificity phosphatases (DUSPs). Here, we show that the effects of MG132 treatment on ERK signaling are more widespread, leading to a reduction in activation of the upstream kinase MEK. This suggests that MG132 systemically perturbs the intracellular phosphoproteome, impacting ERK signaling by reducing phosphorylation status at multiple levels of the kinase cascade. PMID:23226437

Cirit, Murat; Grant, Kyle G; Haugh, Jason M

2012-01-01

160

L166P MUTANT DJ-1, CAUSATIVE FOR RECESSIVE PARKINSON'S DISEASE IS DEGRADED THROUGH THE UBIQUITIN-PROTEASOME SYSTEM  

EPA Science Inventory

Abstract Mutations in a gene on chromosome 1, DJ-1, have been reported recently to be associated with recessive, early-onset Parkinson's disease. Whilst one mutation is a large deletion that is predicted to produce an effective knockout of the gene, the second is a point ...

161

PKC?-dependent activation of the ubiquitin proteasome system is responsible for high glucose-induced human breast cancer MCF-7 cell proliferation, migration and invasion.  

PubMed

Type 2 diabetes mellitus (T2DM) has contributed to advanced breast cancer development over the past decades. However, the mechanism underlying this contribution is poorly understood. In this study, we determined that high glucose enhanced proteasome activity was accompanied by enhanced proliferation, migration and invasion, as well as suppressed apoptosis, in human breast cancer MCF-7 cells. Proteasome inhibitor bortezomib (BZM) pretreatment mitigated high glucose-induced MCF-7 cell growth and invasion. Furthermore, high glucose increased protein kinase C delta (PKC?)-phosphorylation. Administration of the specific PKC? inhibitor rottlerin attenuated high glucose-stimulated cancer cell growth and invasion. In addition, PKC? inhibition by both rottlerin and PKC? shRNA significantly suppressed high glucose-induced proteasome activity. Our results suggest that PKC?-dependent ubiquitin proteasome system activation plays an important role in high glucose- induced breast cancer cell growth and metastasis. PMID:24289563

Zhu, Shan; Yao, Feng; Li, Wen-Huan; Wan, Jin-Nan; Zhang, Yi-Min; Tang, Zhao; Khan, Shahzad; Wang, Chang-Hua; Sun, Sheng-Rong

2013-01-01

162

Oxidized LDLs alter the activity of the ubiquitin- proteasome pathway: potential role in oxidized LDL induced apoptosis  

Microsoft Academic Search

Oxidized low-density lipoproteins (ox- LDL) play a role in the genesis of atherosclerosis. OxLDL are able to induce apoptosis of vascular cells, which is potentially involved in the formation of the necrotic center of atherosclerotic lesions, plaque rupture, and subsequent thrombotic events. Because oxLDL may induce structural modifications of cell protein and altered proteins may impair cell viability, the present

OTILIA VIEIRA; ISABELLE ESCARGUEIL-BLANC; GUNTHER JURGENS; CHRISTOPH BORNER; LEONOR ALMEIDA; ROBERT SALVAYRE; ANNE NEGRE-SALVAYRE

163

The roles of the ubiquitin-proteasome and autophagy–lysosome pathways in Huntington's disease and related conditions  

Microsoft Academic Search

Huntington's disease (HD) is one of nine known conditions caused by a CAG trinucleotide repeat expansion that is translated into a polyglutamine tract in the disease protein. Genetic and transgenic data suggest that the primary effect of these CAG\\/polyglutamine expansions is to confer a toxic gain-of-function on the mutant protein. The cellular pathology of all known polyglutamine diseases is characterised

Brinda Ravikumar; Sovan Sarkar; Zdenek Berger; David C. Rubinsztein

2003-01-01

164

Interactions of Xanthomonas type-III effector proteins with the plant ubiquitin and ubiquitin-like pathways  

PubMed Central

In eukaryotes, regulated protein turnover is required during many cellular processes, including defense against pathogens. Ubiquitination and degradation of ubiquitinated proteins via the ubiquitin–proteasome system (UPS) is the main pathway for the turnover of intracellular proteins in eukaryotes. The extensive utilization of the UPS in host cells makes it an ideal pivot for the manipulation of cellular processes by pathogens. Like many other Gram-negative bacteria, Xanthomonas species secrete a suite of type-III effector proteins (T3Es) into their host cells to promote virulence. Some of these T3Es exploit the plant UPS to interfere with immunity. This review summarizes T3E examples from the genus Xanthomonas with a proven or suggested interaction with the host UPS or UPS-like systems and also discusses the apparent paradox that arises from the presence of T3Es that inhibit the UPS in general while others rely on its activity for their function.

Üstün, Suayib; Börnke, Frederik

2014-01-01

165

Degradation of Activated Protein Kinases by Ubiquitination  

PubMed Central

Protein kinases are important regulators of intracellular signal transduction pathways and play critical roles in diverse cellular functions. Once a protein kinase is activated, its activity is subsequently downregulated through a variety of mechanisms. Accumulating evidence indicates that the activation of protein kinases commonly initiates their downregulation via the ubiquitin/proteasome pathway. Failure to regulate protein kinase activity or expression levels can cause human diseases. PMID:19489726

Lu, Zhimin; Hunter, Tony

2009-01-01

166

Archaeal Tuc1/Ncs6 Homolog Required for Wobble Uridine tRNA Thiolation Is Associated with Ubiquitin-Proteasome, Translation, and RNA Processing System Homologs  

PubMed Central

While cytoplasmic tRNA 2-thiolation protein 1 (Tuc1/Ncs6) and ubiquitin-related modifier-1 (Urm1) are important in the 2-thiolation of 5-methoxycarbonylmethyl-2-thiouridine (mcm5s2U) at wobble uridines of tRNAs in eukaryotes, the biocatalytic roles and properties of Ncs6/Tuc1 and its homologs are poorly understood. Here we present the first report of an Ncs6 homolog of archaea (NcsA of Haloferax volcanii) that is essential for maintaining cellular pools of thiolated tRNALysUUU and for growth at high temperature. When purified from Hfx. volcanii, NcsA was found to be modified at Lys204 by isopeptide linkage to polymeric chains of the ubiquitin-fold protein SAMP2. The ubiquitin-activating E1 enzyme homolog of archaea (UbaA) was required for this covalent modification. Non-covalent protein partners that specifically associated with NcsA were also identified including UbaA, SAMP2, proteasome activating nucleotidase (PAN)-A/1, translation elongation factor aEF-1? and a ?-CASP ribonuclease homolog of the archaeal cleavage and polyadenylation specificity factor 1 family (aCPSF1). Together, our study reveals that NcsA is essential for growth at high temperature, required for formation of thiolated tRNALysUUU and intimately linked to homologs of ubiquitin-proteasome, translation and RNA processing systems. PMID:24906001

Chavarria, Nikita E.; Hwang, Sungmin; Cao, Shiyun; Fu, Xian; Holman, Mary; Elbanna, Dina; Rodriguez, Suzanne; Arrington, Deanna; Englert, Markus; Uthandi, Sivakumar; Söll, Dieter; Maupin-Furlow, Julie A.

2014-01-01

167

Induction of Caspase-3-like activity in Rice following release of cytochrome-f from the chloroplast and subsequent interaction with the Ubiquitin-Proteasome System  

PubMed Central

It has been known that the process of leaf senescence is accompanied by programmed cell death (PCD), and the previous study indicated that dark-induced senescence in detached leaves from rice led to the release of cytochrome f (Cyt f) from chloroplast into the cytoplasm. In this study, the effects of Cyt f on PCD were studied both in vitro and in vivo. In a cell-free system, purified Cyt f activated caspase-3-like protease and endonuclease OsNuc37, and induced DNA fragmentation. Furthermore, Cyt f-induced caspase-3-like activity could be inhibited by MG132, which suggests that the activity was attributed to the 26S proteasome. Conditional expression of Cyt f in the cytoplasm could also activate caspase-3-like activity and DNA fragmentation. Fluorescein diacetate staining and annexin V-FITC/PI double staining demonstrated that Cyt f expression in cytoplasm significantly increased the percentage of PCD protoplasts. Yeast two-hybrid screening showed that Cyt f might interact with E3-ubiquitin ligase and RPN9b, the subunits of the ubiquitin proteasome system (UPS), and other PCD-related proteins. Taken together, these results suggest that the released Cyt f from the chloroplast into the cytoplasm might activate or rescue caspase-3-like activity by interacting with the UPS, ultimately leading to the induction of PCD. PMID:25103621

Wang, Hongjuan; Zhu, Xiaonan; Li, Huan; Cui, Jing; Liu, Cheng; Chen, Xi; Zhang, Wei

2014-01-01

168

Overview of Proteasome Inhibitor-Based Anti-cancer Therapies: Perspective on Bortezomib and Second Generation Proteasome Inhibitors versus Future Generation Inhibitors of Ubiquitin-Proteasome System  

PubMed Central

Over the past ten years, proteasome inhibition has emerged as an effective therapeutic strategy for treating multiple myeloma (MM) and some lymphomas. In 2003, Bortezomib (BTZ) became the first proteasome inhibitor approved by the U.S. Food and Drug Administration (FDA). BTZ-based therapies have become a staple for the treatment of MM at all stages of the disease. The survival rate of MM patients has improved significantly since clinical introduction of BTZ and other immunomodulatory drugs. However, BTZ has several limitations. Not all patients respond to BTZ-based therapies and relapse occurs in many patients who initially responded. Solid tumors, in particular, are often resistant to BTZ. Furthermore, BTZ can induce dose-limiting peripheral neuropathy (PN). The second generation proteasome inhibitor Carfizomib (CFZ; U.S. FDA approved in August 2012) induces responses in a minority of MM patients relapsed from or refractory to BTZ. There is less PN compared to BTZ. Four other second-generation proteasome inhibitors (Ixazomib, Delanzomib, Oprozomib and Marizomib) with different pharmacologic properties and broader anticancer activities, have also shown some clinical activity in bortezomib-resistant cancers. While the mechanism of resistance to bortezomib in human cancers still remains to be fully understood, targeting the immunoproteasome, ubiquitin E3 ligases, the 19S proteasome and deubiquitinases in pre-clinical studies represents possible directions for future generation inhibitors of ubiquitin-proteasome system in the treatment of MM and other cancers. PMID:25092212

Dou, Q. Ping; Zonder, Jeffrey A.

2014-01-01

169

Urban Renewal in the Nucleus: Is Protein Turnover by Proteasomes Absolutely Required for Nuclear Receptor-Regulated Transcription?  

Microsoft Academic Search

The importance of the ubiquitin proteasome path- way in higher eukaryotes has been well established in cell cycle regulation, signal transduction, and cell differentiation, but has only recently been linked to nuclear hormone receptor-regulated gene transcription. Characterization of a number of ubiquitin proteasome pathway enzymes as co- activators and observations that several nuclear receptors are ubiquitinated and degraded in the

ZAFAR NAWAZ; BERT W. O'MALLEY

2003-01-01

170

Sperm survival versus degradation in the Mammalian epididymis: a hypothesis.  

PubMed

A long-standing problem in epididymal physiology is the fate of unejaculated spermatozoa in the cauda epididymidis under conditions such as congenital absence of the vas deferens, long-term vasectomy, or castration. There is no convincing evidence for significant absorption of spermatozoa, defective or otherwise, by spermiophagy or dissolution in the epididymis of normal animals. Spermiophagy by epithelial cells or intraluminal macrophages may take place if the duct ruptures and granulomas form (e.g., after experimental ligation), although there is no quantitative information on the rate of sperm removal by this means. In one animal model (the rabbit), the epididymis is unusually resistant to granuloma formation and has provided unique insights into a phenomenon that is suggested to be present in all species. Spermatozoa retained in the rabbit cauda epididymidis by placing ligatures on the vas deferens and corpus epididymidis degenerate after several weeks but do not decrease significantly in numbers. After castration, however, they die very rapidly and >90% disappear. It is hypothesized that, in the normal androgen-maintained epididymis, degradative pathways are present in the luminal fluid that are constitutively inhibited by survival signals emanating from the epithelium. In the absence of androgen, the intraluminal mileau changes and death signals predominate that activate degradative pathways via the ubiquitin-proteasome system, DNAses, etc., to mediate dissolution of sperm organelles and nucleoprotein. It is suggested that the latter condition is the default situation and is only prevented by the stimulatory action of androgens on the epididymal epithelium. PMID:15215193

Jones, Roy

2004-11-01

171

Regulation of subtilase cytotoxin-induced cell death by an RNA-dependent protein kinase-like endoplasmic reticulum kinase-dependent proteasome pathway in HeLa cells.  

PubMed

Shiga-toxigenic Escherichia coli (STEC) produces subtilase cytotoxin (SubAB), which cleaves the molecular chaperone BiP in the endoplasmic reticulum (ER), leading to an ER stress response and then activation of apoptotic signaling pathways. Here, we show that an early event in SubAB-induced apoptosis in HeLa cells is mediated by RNA-dependent protein kinase (PKR)-like ER kinase (PERK), not activating transcription factor 6 (ATF6) or inositol-requiring enzyme 1(Ire1), two other ER stress sensors. PERK knockdown suppressed SubAB-induced eIF2? phosphorylation, activating transcription factor 4 (ATF4) expression, caspase activation, and cytotoxicity. Knockdown of eIF2? by small interfering RNA (siRNA) or inhibition of eIF2? dephosphorylation by Sal003 enhanced SubAB-induced caspase activation. Treatment with proteasome inhibitors (i.e., MG132 and lactacystin), but not a general caspase inhibitor (Z-VAD) or a lysosome inhibitor (chloroquine), suppressed SubAB-induced caspase activation and poly(ADP-ribose) polymerase (PARP) cleavage, suggesting that the ubiquitin-proteasome system controls events leading to caspase activation, i.e., Bax/Bak conformational changes, followed by cytochrome c release from mitochondria. Levels of ubiquitinated proteins in HeLa cells were significantly decreased by SubAB treatment. Further, in an early event, some antiapoptotic proteins, which normally turn over rapidly, have their synthesis inhibited, and show enhanced degradation via the proteasome, resulting in apoptosis. In PERK knockdown cells, SubAB-induced loss of ubiquitinated proteins was inhibited. Thus, SubAB-induced ER stress is caused by BiP cleavage, leading to PERK activation, not by accumulation of ubiquitinated proteins, which undergo PERK-dependent degradation via the ubiquitin-proteasome system. PMID:22354021

Yahiro, Kinnosuke; Tsutsuki, Hiroyasu; Ogura, Kohei; Nagasawa, Sayaka; Moss, Joel; Noda, Masatoshi

2012-05-01

172

Protection from Endotoxic Uveitis by Intravitreal Resolvin D1: Involvement of Lymphocytes, miRNAs, Ubiquitin-Proteasome, and M1/M2 Macrophages  

PubMed Central

This study investigated the protective effects of intravitreal Resolvin D1 (RvD1) against LPS-induced rat endotoxic uveitis (EIU). RvD1 was administered into the right eye at a single injection of 5??L volume containing 10–100–1000?ng/kg RvD1 1?h post-LPS injection (200??g, Salmonella minnesota) into thefootpad of Sprague-Dawley rats. 24?h later, the eye was enucleated and examined for clinical, biochemical, and immunohistochemical evaluations. RvD1 significantly and dose-dependently decreased the clinical score attributed to EIU, starting from the dose of 10?ng/kg and further decreased by 100 and 1000?ng/kg. These effects were accompanied by changes in four important determinants of the immune-inflammatory response within the eye: (i) the B and T lymphocytes, (ii) the miRNAs pattern, (iii) the ubiquitin-proteasome system (UPS), and (iv) the M1/M2 macrophage phenotype. LPS+RvD1 treated rats showed reduced presence of B and T lymphocytes and upregulation of miR-200c-3p, miR 203a-3p, miR 29b-3p, and miR 21-5p into the eye compared to the LPS alone. This was paralleled by decreases of the ubiquitin, 20S and 26S proteasome subunits, reduced presence of macrophage M1, and increased presence of macrophage M2 in the ocular tissues. Accordingly, the levels of the cytokine TNF-?, the chemokines MIP1-? and NF-?B were reduced.

Rossi, S.; Di Filippo, C.; Gesualdo, C.; Potenza, N.; Russo, A.; Trotta, M. C.; Zippo, M. V.; Maisto, R.; Ferraraccio, F.; Simonelli, F.; D'Amico, M.

2015-01-01

173

Assaying proteasomal degradation in a cell-free system in plants.  

PubMed

The ubiquitin-proteasome pathway for protein degradation has emerged as one of the most important mechanisms for regulation of a wide spectrum of cellular functions in virtually all eukaryotic organisms. Specifically, in plants, the ubiquitin/26S proteasome system (UPS) regulates protein degradation and contributes significantly to development of a wide range of processes, including immune response, development and programmed cell death. Moreover, increasing evidence suggests that numerous plant pathogens, such as Agrobacterium, exploit the host UPS for efficient infection, emphasizing the importance of UPS in plant-pathogen interactions. The substrate specificity of UPS is achieved by the E3 ubiquitin ligase that acts in concert with the E1 and E2 ligases to recognize and mark specific protein molecules destined for degradation by attaching to them chains of ubiquitin molecules. One class of the E3 ligases is the SCF (Skp1/Cullin/F-box protein) complex, which specifically recognizes the UPS substrates and targets them for ubiquitination via its F-box protein component. To investigate a potential role of UPS in a biological process of interest, it is important to devise a simple and reliable assay for UPS-mediated protein degradation. Here, we describe one such assay using a plant cell-free system. This assay can be adapted for studies of the roles of regulated protein degradation in diverse cellular processes, with a special focus on the F-box protein-substrate interactions. PMID:24747194

García-Cano, Elena; Zaltsman, Adi; Citovsky, Vitaly

2014-01-01

174

Ubiquitin-protein ligases in muscle wasting: multiple parallel pathways?  

NASA Technical Reports Server (NTRS)

PURPOSE OF REVIEW: Studies in a wide variety of animal models of muscle wasting have led to the concept that increased protein breakdown via the ubiquitin-proteasome pathway is responsible for the loss of muscle mass seen as muscle atrophy. The complexity of the ubiquitination apparatus has hampered our understanding of how this pathway is activated in atrophying muscles and which ubiquitin-conjugating enzymes in muscle are responsible. RECENT FINDINGS: Recent experiments have shown that two newly identified ubiquitin-protein ligases (E3s), atrogin-1/MAFbx and MURF-1, are critical in the development of muscle atrophy. Other in-vitro studies also implicated E2(14k) and E3alpha, of the N-end rule pathway, as playing an important role in the process. SUMMARY: It seems likely that multiple pathways of ubiquitin conjugation are activated in parallel in atrophying muscle, perhaps to target for degradation specific classes of muscle proteins. The emerging challenge will be to define the protein targets for, as well as inhibitors of, these E3s.

Lecker, Stewart H.; Goldberg, A. L. (Principal Investigator)

2003-01-01

175

Glycosylation-independent ERAD pathway serves as a backup system under ER stress  

PubMed Central

During endoplasmic reticulum (ER)–associated degradation (ERAD), terminally misfolded proteins are retrotranslocated from the ER to the cytosol and degraded by the ubiquitin-proteasome system. Misfolded glycoproteins are recognized by calnexin and transferred to EDEM1, followed by the ER disulfide reductase ERdj5 and the BiP complex. The mechanisms involved in ERAD of nonglycoproteins, however, are poorly understood. Here we show that nonglycoprotein substrates are captured by BiP and then transferred to ERdj5 without going through the calnexin/EDEM1 pathway; after cleavage of disulfide bonds by ERdj5, the nonglycoproteins are transferred to the ERAD scaffold protein SEL1L by the aid of BiP for dislocation into the cytosol. When glucose trimming of the N-glycan groups of the substrates is inhibited, glycoproteins are also targeted to the nonglycoprotein ERAD pathway. These results indicate that two distinct pathways for ERAD of glycoproteins and nonglycoproteins exist in mammalian cells, and these pathways are interchangeable under ER stress conditions. PMID:23966469

Ushioda, Ryo; Hoseki, Jun; Nagata, Kazuhiro

2013-01-01

176

Bacterial Effectors and Their Functions in the Ubiquitin-Proteasome System: Insight from the Modes of Substrate Recognition  

PubMed Central

Protein ubiquitination plays indispensable roles in the regulation of cell homeostasis and pathogenesis of neoplastic, infectious, and neurodegenerative diseases. Given the importance of this modification, it is to be expected that several pathogenic bacteria have developed the ability to utilize the host ubiquitin system for their own benefit. Modulation of the host ubiquitin system by bacterial effector proteins inhibits innate immune responses and hijacks central signaling pathways. Bacterial effectors mimic enzymes of the host ubiquitin system, but may or may not be structurally similar to the mammalian enzymes. Other effectors bind and modify components of the host ubiquitin system, and some are themselves subject to ubiquitination. This review will describe recent findings, based on structural analyses, regarding how pathogens use post-translational modifications of proteins to establish an infection. PMID:25257025

Kim, Minsoo; Otsubo, Ryota; Morikawa, Hanako; Nishide, Akira; Takagi, Kenji; Sasakawa, Chihiro; Mizushima, Tsunehiro

2014-01-01

177

Torbafylline (HWA 448) inhibits enhanced skeletal muscle ubiquitin-proteasome-dependent proteolysis in cancer and septic rats.  

PubMed Central

The development of new pharmacological approaches for preventing muscle wasting in cancer is an important goal because cachectic patients display a reduced response to chemotherapy and radiotherapy. Xanthine derivatives such as pentoxifylline inhibit tumour necrosis factor-alpha (TNF) production, which has been implicated in the signalling of muscle wasting. However, the effect of pentoxifylline has been inconclusive in clinical trials. We report here the first direct evidence that daily injections of torbafylline (also known as HWA 448), another xanthine derivative, had no effect by itself on muscle proteolysis in control healthy rats. In cancer rats, the drug blocked the lipopolysaccharide-induced hyperproduction of TNF and prevented muscle wasting. In these animals HWA 448 suppressed the enhanced proteasome-dependent proteolysis, which is sensitive to the proteasome inhibitor MG132, and the accumulation of high-molecular-mass ubiquitin (Ub) conjugates in the myofibrillar fraction. The drug also normalized the enhanced muscle expression of Ub, which prevails in the atrophying muscles from cancer rats. In contrast, HWA 448 did not reduce the increased expression of either the 14 kDa Ub conjugating enzyme E2 or the ATPase and non-ATPase subunits of the 19 S regulatory complex of the 26 S proteasome, including the non-ATPase subunit S5a, which recognizes polyUb degradation signals. Finally, the drug also prevented muscle wasting in septic rats (which exhibit increased TNF production), and was much more potent than pentoxifylline or other xanthine derivatives. Taken together, the data indicate that HWA 448 is a powerful inhibitor of muscle wasting that blocks enhanced Ub-proteasome-dependent proteolysis in situations where TNF production rises, including cancer and sepsis. PMID:11772390

Combaret, Lydie; Tilignac, Thomas; Claustre, Agnès; Voisin, Laure; Taillandier, Daniel; Obled, Christiane; Tanaka, Keiji; Attaix, Didier

2002-01-01

178

Gene expression: degrade to derepress.  

PubMed

Chromatin immunoprecipitation and sequencing (ChIP-seq) provides a static snap-shot of DNA-associated proteins which fails to reflect the dynamics of the DNA-bound proteome. Now, Catic and co-workers combine ubiquitin ChIP-seq and proteasome inhibitors to map sites of DNA-associated protein degradation on a genome-wide scale. They identify an ubiquitin ligase which targets a transcriptional repressor for destruction by the proteasome, thus activating transcription of specific genes. These findings reveal that the ubiquitin proteasome system actively regulates transcription. PMID:24473147

McShane, Erik; Selbach, Matthias

2014-03-01

179

Inhibition of protein deubiquitination by PR-619 activates the autophagic pathway in OLN-t40 oligodendroglial cells.  

PubMed

Protein aggregate formation may be the result of an impairment of the protein quality control system, e.g., the ubiquitin proteasome system (UPS) and the lysosomal autophagic pathway. For proteasomal degradation, proteins need to be covalently modified by ubiquitin and deubiquitinated before the substrates are proteolytically degraded. Deubiquitination is performed by a large family of proteases, the deubiquitinating enzymes (DUBs). DUBs display a variety of functions and their inhibition may have pathological consequences. Using the broad specificity DUB inhibitor PR-619 we previously have shown that DUB inhibition leads to an overload of ubiquitinated proteins, to protein aggregate formation and subsequent inhibition of the UPS. This study was undertaken to investigate whether PR-619 modulates autophagic functions to possibly compensate the failure of the proteasomal system. Using the oligodendroglial cell line OLN-t40 and a new oligodendroglial cell line stably expressing GFP-LC3, we show that DUB inhibition leads to the activation of autophagy and to the recruitment of LC3 and of the ubiquitin binding protein p62 to the forming aggresomes without impairing the autophagic flux. Furthermore, PR-619 induced the transport of lysosomes to the forming aggregates in a process requiring an intact microtubule network. Further stimulation of autophagy by rapamycin did not prevent PR-619 aggregate formation but rather exerted cytotoxic effects. Hence, inhibition of DUBs by PR-619 activated the autophagic pathway supporting the hypothesis that the UPS and the autophagy-lysosomal pathway are closely linked together. PMID:23686611

Seiberlich, Veronika; Borchert, Janika; Zhukareva, Victoria; Richter-Landsberg, Christiane

2013-09-01

180

Regulating the 20S Proteasome Ubiquitin-Independent Degradation Pathway  

PubMed Central

For many years, the ubiquitin-26S proteasome degradation pathway was considered the primary route for proteasomal degradation. However, it is now becoming clear that proteins can also be targeted for degradation by the core 20S proteasome itself. Degradation by the 20S proteasome does not require ubiquitin tagging or the presence of the 19S regulatory particle; rather, it relies on the inherent structural disorder of the protein being degraded. Thus, proteins that contain unstructured regions due to oxidation, mutation, or aging, as well as naturally, intrinsically unfolded proteins, are susceptible to 20S degradation. Unlike the extensive knowledge acquired over the years concerning degradation by the 26S proteasome, relatively little is known about the means by which 20S-mediated proteolysis is controlled. Here, we describe our current understanding of the regulatory mechanisms that coordinate 20S proteasome-mediated degradation, and highlight the gaps in knowledge that remain to be bridged. PMID:25250704

Ben-Nissan, Gili; Sharon, Michal

2014-01-01

181

Malin Regulates Wnt Signaling Pathway through Degradation of Dishevelled2*  

PubMed Central

Using yeast-two hybrid screening followed by co-immunoprecipitation assay, we have found that the Lafora disease ubiquitin ligase malin interacts with dishevelled2, a key mediator of Wnt signaling pathway. Overexpression of malin enhances the degradation of dishevelled2 and inhibits Wnt signaling, which is evident from the down-regulation of ?-catenin target genes and the decrease in ?-catenin-mediated transcriptional activity. Partial knockdown of malin significantly increases the level of dishevelled2 and up-regulates Wnt signaling. Several malin mutants are found to be ineffective in degrading dishevelled2 and regulating the Wnt pathway. We have also found that malin enhances K48- and K63-linked ubiquitination of dishevelled2 that could lead to its degradation through both proteasome and autophagy. Altogether, our results indicate that malin regulates Wnt signaling pathway through the degradation of dishevelled2 and suggest possible deregulation of Wnt signaling in Lafora disease. PMID:22223637

Sharma, Jaiprakash; Mulherkar, Shalaka; Mukherjee, Diptendu; Jana, Nihar Ranjan

2012-01-01

182

Malin regulates Wnt signaling pathway through degradation of dishevelled2.  

PubMed

Using yeast-two hybrid screening followed by co-immunoprecipitation assay, we have found that the Lafora disease ubiquitin ligase malin interacts with dishevelled2, a key mediator of Wnt signaling pathway. Overexpression of malin enhances the degradation of dishevelled2 and inhibits Wnt signaling, which is evident from the down-regulation of ?-catenin target genes and the decrease in ?-catenin-mediated transcriptional activity. Partial knockdown of malin significantly increases the level of dishevelled2 and up-regulates Wnt signaling. Several malin mutants are found to be ineffective in degrading dishevelled2 and regulating the Wnt pathway. We have also found that malin enhances K48- and K63-linked ubiquitination of dishevelled2 that could lead to its degradation through both proteasome and autophagy. Altogether, our results indicate that malin regulates Wnt signaling pathway through the degradation of dishevelled2 and suggest possible deregulation of Wnt signaling in Lafora disease. PMID:22223637

Sharma, Jaiprakash; Mulherkar, Shalaka; Mukherjee, Diptendu; Jana, Nihar Ranjan

2012-02-24

183

Reporter-based growth assay for systematic analysis of protein degradation.  

PubMed

Protein degradation by the ubiquitin-proteasome system (UPS) is a major regulatory mechanism for protein homeostasis in all eukaryotes. The standard approach to determining intracellular protein degradation relies on biochemical assays for following the kinetics of protein decline. Such methods are often laborious and time consuming and therefore not amenable to experiments aimed at assessing multiple substrates and degradation conditions. As an alternative, cell growth-based assays have been developed, that are, in their conventional format, end-point assays that cannot quantitatively determine relative changes in protein levels. Here we describe a method that faithfully determines changes in protein degradation rates by coupling them to yeast cell-growth kinetics. The method is based on an established selection system where uracil auxotrophy of URA3-deleted yeast cells is rescued by an exogenously expressed reporter protein, comprised of a fusion between the essential URA3 gene and a degradation determinant (degron). The reporter protein is designed so that its synthesis rate is constant whilst its degradation rate is determined by the degron. As cell growth in uracil-deficient medium is proportional to the relative levels of Ura3, growth kinetics are entirely dependent on the reporter protein degradation. This method accurately measures changes in intracellular protein degradation kinetics. It was applied to: (a) Assessing the relative contribution of known ubiquitin-conjugating factors to proteolysis (b) E2 conjugating enzyme structure-function analyses (c) Identification and characterization of novel degrons. Application of the degron-URA3-based system transcends the protein degradation field, as it can also be adapted to monitoring changes of protein levels associated with functions of other cellular pathways. PMID:25406949

Cohen, Itamar; Geffen, Yifat; Ravid, Guy; Ravid, Tommer

2014-01-01

184

ATR checkpoint kinase and CRL1?TRCP collaborate to degrade ASF1a and thus repress genes overlapping with clusters of stalled replication forks  

PubMed Central

Many agents used for chemotherapy, such as doxorubicin, interfere with DNA replication, but the effect of this interference on transcription is largely unknown. Here we show that doxorubicin induces the firing of dense clusters of neoreplication origins that lead to clusters of stalled replication forks in gene-rich parts of the genome, particularly on expressed genes. Genes that overlap with these clusters of stalled forks are actively dechromatinized, unwound, and repressed by an ATR-dependent checkpoint pathway. The ATR checkpoint pathway causes a histone chaperone normally associated with the replication fork, ASF1a, to degrade through a CRL1?TRCP-dependent ubiquitination/proteasome pathway, leading to the localized dechromatinization and gene repression. Therefore, a globally active checkpoint pathway interacts with local clusters of stalled forks to specifically repress genes in the vicinity of the stalled forks, providing a new mechanism of action of chemotherapy drugs like doxorubicin. Finally, ASF1a-depleted cancer cells are more sensitive to doxorubicin, suggesting that the 7%–10% of prostate adenocarcinomas and adenoid cystic carcinomas reported to have homozygous deletion or significant underexpression of ASF1a should be tested for high sensitivity to doxorubicin. PMID:24700029

Im, Jun-Sub; Keaton, Mignon; Lee, Kyung Yong; Kumar, Pankaj; Park, Jonghoon; Dutta, Anindya

2014-01-01

185

Evolution of Efficient Pathways for Degradation of Anthropogenic Chemicals  

PubMed Central

Anthropogenic compounds used as pesticides, solvents, and explosives often persist in the environment and can cause toxicity to humans and wildlife. The persistence of anthropogenic compounds is due to their recent introduction into the environment; microbes in soil and water have had relatively little time to evolve efficient mechanisms for degradation of these novel compounds. Some anthropogenic compounds are easily degraded, while others are degraded very slowly or only partially, leading to accumulation of toxic products. This review examines the factors that affect the ability of microbes to degrade anthropogenic compounds and the mechanisms by which novel pathways emerge in nature. New approaches for engineering microbes with enhanced degradative abilities include assembly of pathways using enzymes from multiple organisms, directed evolution of inefficient enzymes, and genome shuffling to improve microbial fitness under the challenging conditions posed by contaminated environments. PMID:19620997

Copley, Shelley D.

2010-01-01

186

Understanding Degradation Pathways in Organic Photovoltaics (Poster)  

SciTech Connect

Organic Photovoltaics (OPVs) recently attained power conversion efficiencies that are of interest for commercial production. Consequently, one of the most important unsolved issues facing a new industry is understanding what governs lifetime in organic devices and discovering solutions to mitigate degradation mechanisms. Historically, the active organic components are considered vulnerable to photo-oxidation and represent the primary degradation channel. However, we present several (shelf life and light soaking) studies pointing the relative stability of the active layers and instabilities in commonly used electrode materials. We show that engineering of the hole/electron layer at the electrode can lead to environmentally stable devices even without encapsulation.

Lloyd, M. T.; Olson, D. C.; Garcia, A.; Kauvar, I.; Kopidakis, N.; Reese, M. O.; Berry, J. J.; Ginley, D. S.

2011-02-01

187

Degradation of the E7 human papillomavirus oncoprotein by the ubiquitin-proteasome system: targeting via ubiquitination of the N-terminal residue  

Microsoft Academic Search

The E7 oncoprotein of the high risk human papillomavirus type 16 (HPV-16), which is etiologically associated with uterine cervical cancer, is a potent immortalizing and transforming agent. It probably exerts its oncogenic functions by interacting and altering the normal activity of cell cycle control proteins such as p21WAF1, p27KIP1 and pRb, transcriptional activators such as TBP and AP-1, and metabolic

Eyal Reinstein; Martin Scheffner; Moshe Oren; Aaron Ciechanover; Alan Schwartz

2000-01-01

188

Binding-induced folding of prokaryotic ubiquitin-like protein on the mycobacterium proteasomal ATPase targets substrates for degradation  

SciTech Connect

Mycobacterium tuberculosis uses a proteasome system that is analogous to the eukaryotic ubiquitin-proteasome pathway and is required for pathogenesis. However, the bacterial analog of ubiquitin, prokaryotic ubiquitin-like protein (Pup), is an intrinsically disordered protein that bears little sequence or structural resemblance to the highly structured ubiquitin. Thus, it was unknown how pupylated proteins were recruited to the proteasome. Here, we show that the Mycobacterium proteasomal ATPase (Mpa) has three pairs of tentacle-like coiled coils that recognize Pup. Mpa bound unstructured Pup through hydrophobic interactions and a network of hydrogen bonds, leading to the formation of an {alpha}-helix in Pup. Our work describes a binding-induced folding recognition mechanism in the Pup-proteasome system that differs mechanistically from substrate recognition in the ubiquitin-proteasome system. This key difference between the prokaryotic and eukaryotic systems could be exploited for the development of a small molecule-based treatment for tuberculosis.

Wang, T.; Li, H.; Darwin, K. H.

2010-11-01

189

Binding-induced Folding of Prokaryotic Ubiquitin-like Protein on the Mycobacterium Proteasomal ATPase Targets Substrates for Degradation  

SciTech Connect

Mycobacterium tuberculosis uses a proteasome system that is analogous to the eukaryotic ubiquitin-proteasome pathway and is required for pathogenesis. However, the bacterial analog of ubiquitin, prokaryotic ubiquitin-like protein (Pup), is an intrinsically disordered protein that bears little sequence or structural resemblance to the highly structured ubiquitin. Thus, it was unknown how pupylated proteins were recruited to the proteasome. Here, we show that the Mycobacterium proteasomal ATPase (Mpa) has three pairs of tentacle-like coiled coils that recognize Pup. Mpa bound unstructured Pup through hydrophobic interactions and a network of hydrogen bonds, leading to the formation of an {alpha}-helix in Pup. Our work describes a binding-induced folding recognition mechanism in the Pup-proteasome system that differs mechanistically from substrate recognition in the ubiquitin-proteasome system. This key difference between the prokaryotic and eukaryotic systems could be exploited for the development of a small molecule-based treatment for tuberculosis.

T Wang; K Heran Darwin; H Li

2011-12-31

190

Ubiquitin-Dependent Degradation of Ikappa Balpha is Mediated by a Ubiquitin Ligase Skp1\\/Cul 1\\/F-Box Protein FWD1  

Microsoft Academic Search

Activation of the transcription factor nuclear factor kappa B (NF-kappa B) is controlled by proteolysis of its inhibitory subunit (Ikappa B) via the ubiquitin-proteasome pathway. Signal-induced phosphorylation of Ikappa Balpha by a large multisubunit complex containing Ikappa B kinases is a prerequisite for ubiquitination. Here, we show that FWD1 (a mouse homologue of Slimb\\/beta TrCP), a member of the F-box\\/WD40-repeat

Shigetsugu Hatakeyama; Masatoshi Kitagawa; Keiko Nakayama; Michiko Shirane; Masaki Matsumoto; Kimihiko Hattori; Hideaki Higashi; Hiroyasu Nakano; Ko Okumura; Kazunori Onoe; Robert A. Good; Kei-Ichi Nakayama

1999-01-01

191

Genomic and functional profiling of duplicated chromosome 15 cell lines reveal regulatory alterations in UBE3A-associated ubiquitin-proteasome pathway processes  

Microsoft Academic Search

Autism is a complex neurodevelopmental disorder having both genetic and epigenetic etiological elements. Isodicentric chromosome 15 (Idic15), characterized by duplications of the multi-disorder critical region of 15q11-q14, is a relatively common cytogenetic event. When the duplication involves maternally derived con- tent, this abnormality is strongly correlated with autism disorder. However, the mechanistic links between Idic15 and autism are ill-defined. To

Colin A. Baron; Clifford G. Tepper; Stephenie Y. Liu; Ryan R. Davis; Nicholas J. Wang; N. Carolyn Schanen; Jeffrey P. Gregg

2006-01-01

192

Mutations in the tumor suppressors Smad2 and Smad4 inactivate transforming growth factor signaling by targeting Smads to the ubiquitin-proteasome pathway  

Microsoft Academic Search

Biological signals for transforming growth factor (TGF-) are transduced through transmembrane serine\\/threonine kinase receptors that signal to a family of intracellular mediators known as Smads. Smad2 and Smad4 are important for transcriptional and antiproliferative responses to TGF-, and their inactivation in human cancers indicates that they are tumor suppressors. A missense mutation at a conserved arginine residue in the amino-terminal

Jing Xu; Liliana Attisano

2000-01-01

193

Ubiquitin-mediated NF?B degradation pathway.  

PubMed

The nuclear factor ?B (NF?B) transcription factor plays critical roles in inflammation and immunity. The dysregulation of NF?B is associated with inflammatory and autoimmune diseases and cancer. NF?B activation is negatively regulated by the ubiquitin-dependent proteasomal degradation pathway. In the present review, we discuss recent advances in our understanding of how ubiquitin ligases regulate the NF?B degradation pathway.Cellular & Molecular Immunology advance online publication, 27 October 2014; doi:10.1038/cmi.2014.99. PMID:25345807

Xu, Huimin; You, Mengli; Shi, Haifeng; Hou, Yongzhong

2014-10-27

194

Profiling of Ubiquitination Pathway Genes in Peripheral Cells from Patients with Frontotemporal Dementia due to C9ORF72 and GRN Mutations.  

PubMed

We analysed the expression levels of 84 key genes involved in the regulated degradation of cellular protein by the ubiquitin-proteasome system in peripheral cells from patients with frontotemporal dementia (FTD) due to C9ORF72 and GRN mutations, as compared with sporadic FTD and age-matched controls. A SABiosciences PCR array was used to investigate the transcription profile in a discovery population consisting of six patients each in C9ORF72, GRN, sporadic FTD and age-matched control groups. A generalized down-regulation of gene expression compared with controls was observed in C9ORF72 expansion carriers and sporadic FTD patients. In particular, in both groups, four genes, UBE2I, UBE2Q1, UBE2E1 and UBE2N, were down-regulated at a statistically significant (p < 0.05) level. All of them encode for members of the E2 ubiquitin-conjugating enzyme family. In GRN mutation carriers, no statistically significant deregulation of ubiquitination pathway genes was observed, except for the UBE2Z gene, which displays E2 ubiquitin conjugating enzyme activity, and was found to be statistically significant up-regulated (p = 0.006). These preliminary results suggest that the proteasomal degradation pathway plays a role in the pathogenesis of FTD associated with TDP-43 pathology, although different proteins are altered in carriers of GRN mutations as compared with carriers of the C9ORF72 expansion. PMID:25580532

Serpente, Maria; Fenoglio, Chiara; Cioffi, Sara M G; Bonsi, Rossana; Arighi, Andrea; Fumagalli, Giorgio G; Ghezzi, Laura; Scarpini, Elio; Galimberti, Daniela

2015-01-01

195

Profiling of Ubiquitination Pathway Genes in Peripheral Cells from Patients with Frontotemporal Dementia due to C9ORF72 and GRN Mutations  

PubMed Central

We analysed the expression levels of 84 key genes involved in the regulated degradation of cellular protein by the ubiquitin-proteasome system in peripheral cells from patients with frontotemporal dementia (FTD) due to C9ORF72 and GRN mutations, as compared with sporadic FTD and age-matched controls. A SABiosciences PCR array was used to investigate the transcription profile in a discovery population consisting of six patients each in C9ORF72, GRN, sporadic FTD and age-matched control groups. A generalized down-regulation of gene expression compared with controls was observed in C9ORF72 expansion carriers and sporadic FTD patients. In particular, in both groups, four genes, UBE2I, UBE2Q1, UBE2E1 and UBE2N, were down-regulated at a statistically significant (p < 0.05) level. All of them encode for members of the E2 ubiquitin-conjugating enzyme family. In GRN mutation carriers, no statistically significant deregulation of ubiquitination pathway genes was observed, except for the UBE2Z gene, which displays E2 ubiquitin conjugating enzyme activity, and was found to be statistically significant up-regulated (p = 0.006). These preliminary results suggest that the proteasomal degradation pathway plays a role in the pathogenesis of FTD associated with TDP-43 pathology, although different proteins are altered in carriers of GRN mutations as compared with carriers of the C9ORF72 expansion. PMID:25580532

Serpente, Maria; Fenoglio, Chiara; Cioffi, Sara M. G.; Bonsi, Rossana; Arighi, Andrea; Fumagalli, Giorgio G.; Ghezzi, Laura; Scarpini, Elio; Galimberti, Daniela

2015-01-01

196

Degradation pathways of PCB upon gamma irradiation.  

PubMed Central

In order to understand the modifications of the chromatographic profile of Aroclor 1260 upon gamma irradiation, a total of 14 pure polychlorinated biphenyl (PCB) congeners were separately irradiated in solution. Dechlorination was observed, and the generated products were investigated by gas chromatography and mass spectrometry. Degradation proceeds more rapidly in methanol/water mixture than in petroleum ether, but the relative amount of ortho-dechlorinated congeners formed upon irradiation was smaller in the former solvent. Ortho chlorines are preferentially lost in petroleum ether except when they are involved in a 2.5 (or 3.6) substitution pattern, in which case para dechlorination becomes predominant. The precursors of some toxicologically important congeners such as congeners 77, 118, 167, and 189 have been identified. These data are useful to rationalize the modifications of the chromatographic profile of PCB complex mixture upon gamma irradiation. PMID:2088745

Lépine, F; Massé, R

1990-01-01

197

The Type II Hsp40 Sis1 Cooperates with Hsp70 and the E3 Ligase Ubr1 to Promote Degradation of Terminally Misfolded Cytosolic Protein  

PubMed Central

Mechanisms for cooperation between the cytosolic Hsp70 system and the ubiquitin proteasome system during protein triage are not clear. Herein, we identify new mechanisms for selection of misfolded cytosolic proteins for degradation via defining functional interactions between specific cytosolic Hsp70/Hsp40 pairs and quality control ubiquitin ligases. These studies revolved around the use of S. cerevisiae to elucidate the degradation pathway of a terminally misfolded reporter protein, short-lived GFP (slGFP). The Type I Hsp40 Ydj1 acts with Hsp70 to suppress slGFP aggregation. In contrast, the Type II Hsp40 Sis1 is required for proteasomal degradation of slGFP. Sis1 and Hsp70 operate sequentially with the quality control E3 ubiquitin ligase Ubr1 to target slGFP for degradation. Compromise of Sis1 or Ubr1 function leads slGFP to accumulate in a Triton X-100-soluble state with slGFP degradation intermediates being concentrated into perinuclear and peripheral puncta. Interestingly, when Sis1 activity is low the slGFP that is concentrated into puncta can be liberated from puncta and subsequently degraded. Conversely, in the absence of Ubr1, slGFP and the puncta that contain slGFP are relatively stable. Ubr1 mediates proteasomal degradation of slGFP that is released from cytosolic protein handling centers. Pathways for proteasomal degradation of misfolded cytosolic proteins involve functional interplay between Type II Hsp40/Hsp70 chaperone pairs, PQC E3 ligases, and storage depots for misfolded proteins. PMID:23341891

Summers, Daniel W.; Wolfe, Katie J.; Ren, Hong Yu; Cyr, Douglas M.

2013-01-01

198

The deubiquitinating enzyme USP11 controls an IkappaB kinase alpha (IKKalpha)-p53 signaling pathway in response to tumor necrosis factor alpha (TNFalpha).  

PubMed

Post-translational modification and degradation of proteins by the ubiquitin-proteasome system are key regulatory events in cellular responses to various stimuli. The NF-kappaB signaling pathway is controlled by the ubiquitin-mediated proteolysis. Although mechanisms of ubiquitination in the NF-kappaB pathway have been extensively studied, deubiquitination-mediated regulation of the NF-kappaB signaling remains poorly understood. The present studies show that a deubiquitinating enzyme, USP11, specifically regulates IkappaB kinase alpha (IKKalpha) among the NF-kappaB signaling molecules. Knocking down USP11 attenuates expression of IKKalpha in the transcriptional, but not the post-translational, level. However, down-regulation of USP11 dramatically enhances NF-kappaB activity in response to tumor necrosis factor-alpha, indicating that IKKalpha does not require activation of NF-kappaB. Instead, knock down of USP11 or IKKalpha is associated with abrogation of p53 expression upon exposure to tumor necrosis factor-alpha. In concert with these results, silencing of USP11 is associated with transcriptional attenuation of the p53-responsive genes, such as p21 or Bax. Importantly, the ectopic expression of IKKalpha into cells silenced for USP11 restores p53 expression, demonstrating that USP11 functions as an upstream regulator of an IKKalpha-p53 signaling pathway. PMID:17897950

Yamaguchi, Tomoko; Kimura, Junko; Miki, Yoshio; Yoshida, Kiyotsugu

2007-11-23

199

Redox-Regulated Pathway of Tyrosine Phosphorylation Underlies NF-?B Induction by an Atypical Pathway Independent of the 26S Proteasome.  

PubMed

Alternative redox stimuli such as pervanadate or hypoxia/reoxygenation, induce transcription factor NF-?B by phospho-tyrosine-dependent and proteasome-independent mechanisms. While considerable attention has been paid to the absence of proteasomal regulation of tyrosine phosphorylated I?B?, there is a paucity of information regarding proteasomal regulation of signaling events distinct from tyrosine phosphorylation of I?B?. To delineate roles for the ubiquitin-proteasome pathway in the phospho-tyrosine dependent mechanism of NF-?B induction, we employed the proteasome inhibitor, Aclacinomycin, and the phosphotyrosine phosphatase inhibitor, pervanadate (PV). Results from these studies demonstrate that phospho-I?B? (Tyr-42) is not subject to proteasomal degradation in a murine stromal epithelial cell line, confirming results previously reported. Correspondingly, proteasome inhibition had no discernable effect on the key signaling intermediaries, Src and ERK1/2, involved in the phospho-tyrosine mechanisms regulating PV-mediated activation of NF-?B. Consistent with previous reports, a significant redox imbalance leading to the activation of tyrosine kinases, as occurs with pervanadate, is required for the induction of NF-?B. Strikingly, our studies demonstrate that proteasome inhibition can potentiate oxidative stress associated with PV-stimulation without impacting kinase activation, however, other cellular implications for this increase in intracellular oxidation remain to be fully delineated. PMID:25671697

Cullen, Sarah; Ponnappan, Subramaniam; Ponnappan, Usha

2015-01-01

200

Degradation Pathway Models for Photovoltaics Module Lifetime Performance  

E-print Network

.S. Department of Energy workshop on Science for Energy Technologies [1] identified photovoltaics (PV) lifetimeDegradation Pathway Models for Photovoltaics Module Lifetime Performance Nicholas R. Wheeler, Laura data from Underwriter Labs, featuring measurements taken on 18 identical photovoltaic (PV) modules

Rollins, Andrew M.

201

Vacuole import and degradation pathway: Insights into a specialized autophagy pathway  

PubMed Central

Glucose deprivation induces the synthesis of pivotal gluconeogenic enzymes such as fructose-1,6-bisphosphatase, malate dehydrogenase, phosphoenolpyruvate carboxykinase and isocitrate lyase in Saccharomyces cerevisiae. However, following glucose replenishment, these gluconeogenic enzymes are inactivated and degraded. Studies have characterized the mechanisms by which these enzymes are inactivated in response to glucose. The site of degradation of these proteins has also been ascertained to be dependent on the duration of starvation. Glucose replenishment of short-term starved cells results in these proteins being degraded in the proteasome. In contrast, addition of glucose to cells starved for a prolonged period results in these proteins being degraded in the vacuole. In the vacuole dependent pathway, these proteins are sequestered in specialized vesicles termed vacuole import and degradation (Vid). These vesicles converge with the endocytic pathway and deliver their cargo to the vacuole for degradation. Recent studies have identified that internalization, as mediated by actin polymerization, is essential for delivery of cargo proteins to the vacuole for degradation. In addition, components of the target of rapamycin complex 1 interact with cargo proteins during glucose starvation. Furthermore, Tor1p dissociates from cargo proteins following glucose replenishment. Future studies will be needed to elaborate on the importance of internalization at the plasma membrane and the subsequent import of cargo proteins into Vid vesicles in the vacuole dependent degradation pathway. PMID:22125667

Alibhoy, Abbas A; Chiang, Hui-Ling

2011-01-01

202

Evolution of a new bacterial pathway for 4-nitrotoluene degradation.  

PubMed

Bacteria that assimilate synthetic nitroarene compounds represent unique evolutionary models, as their metabolic pathways are in the process of adaptation and optimization for the consumption of these toxic chemicals. We used Acidovorax sp. strain JS42, which is capable of growth on nitrobenzene and 2-nitrotoluene, in experiments to examine how a nitroarene degradation pathway evolves when its host strain is challenged with direct selective pressure to assimilate non-native substrates. Although the same enzyme that initiates the degradation of nitrobenzene and 2-nitrotoluene also oxidizes 4-nitrotoluene to 4-methylcatechol, which is a growth substrate for JS42, the strain is incapable of growth on 4-nitrotoluene. Using long-term laboratory evolution experiments, we obtained JS42 mutants that gained the ability to grow on 4-nitrotoluene via a new degradation pathway. The underlying basis for this new activity resulted from the accumulation of specific mutations in the gene encoding the dioxygenase that catalyses the initial oxidation of nitroarene substrates, but at positions distal to the active site and previously unknown to affect activity in this or related enzymes. We constructed additional mutant dioxygenases to identify the order of mutations that led to the improved enzymes. Biochemical analyses revealed a defined, step-wise pathway for the evolution of the improved dioxygenases. PMID:21895789

Ju, Kou-San; Parales, Rebecca E

2011-10-01

203

Validation of microarray data in human lymphoblasts shows a role of the ubiquitin-proteasome system and NF-kB in the pathogenesis of Down syndrome  

PubMed Central

Background Down syndrome (DS) is a complex disorder caused by the trisomy of either the entire, or a critical region of chromosome 21 (21q22.1-22.3). Despite representing the most common cause of mental retardation, the molecular bases of the syndrome are still largely unknown. Methods To better understand the pathogenesis of DS, we analyzed the genome-wide transcription profiles of lymphoblastoid cell lines (LCLs) from six DS and six euploid individuals and investigated differential gene expression and pathway deregulation associated with trisomy 21. Connectivity map and PASS-assisted exploration were used to identify compounds whose molecular signatures counteracted those of DS lymphoblasts and to predict their therapeutic potential. An experimental validation in DS LCLs and fetal fibroblasts was performed for the most deregulated GO categories, i.e. the ubiquitin mediated proteolysis and the NF-kB cascade. Results We show, for the first time, that the level of protein ubiquitination is reduced in human DS cell lines and that proteasome activity is increased in both basal conditions and oxidative microenvironment. We also provide the first evidence that NF-kB transcription levels, a paradigm of gene expression control by ubiquitin-mediated degradation, is impaired in DS due to reduced IkB-alfa ubiquitination, increased NF-kB inhibitor (IkB-alfa) and reduced p65 nuclear fraction. Finally, the DSCR1/DYRK1A/NFAT genes were analysed. In human DS LCLs, we confirmed the presence of increased protein levels of DSCR1 and DYRK1A, and showed that the levels of the transcription factor NFATc2 were decreased in DS along with a reduction of its nuclear translocation upon induction of calcium fluxes. Conclusions The present work offers new perspectives to better understand the pathogenesis of DS and suggests a rationale for innovative approaches to treat some pathological conditions associated to DS. PMID:23830204

2013-01-01

204

Identification of the major degradation pathways of ticagrelor.  

PubMed

Ticagrelor is a direct-acting and reversible P2Y12-adenosine diphosphate (ADP) receptor blocker used as antiplatelet drug. Forced degradation under various stress conditions was carried out. The degradation products have been detected and identified by high-pressure liquid chromatography multistage mass spectrometry (LC-MS(n)) along with high-resolution mass spectrometry. C18 XTerra MS column combined with a linear gradient mobile phase composed of a mixture of 10mM acetate ammonium/acetonitrile was shown suitable for drug and impurity determinations and validated as a stability indicating method. Structural elucidation of the degradation products relied on MS(n) studies and accurate mass measurements giving access to elemental compositions. Up to nine degradation products resulting from oxidation/auto-oxidation, S-dealkylation and N-dealkylation have been identified, covering a range of possible degradation pathways for derivatives with such functional groups. Kinetics was also studied in order to assess the molecule's shelf-life and to identify the most important degradation factors. PMID:25543285

Hassane, Sadou Yaye; Secrétan, Philippe-Henri; Henriet, Théo; Bernard, Mélisande; Amrani, Fatma; Akrout, Wiem; Tilleul, Patrick; Yagoubi, Najet; Do, Bernard

2015-02-01

205

The trans-Anethole Degradation Pathway in an Arthrobacter sp  

Microsoft Academic Search

A bacterial strain (TA13) capable of utilizing t-anethole as the sole carbon source was isolated from soil. The strain was identified as Arthrobacter aurescens based on its 16 S rRNA gene sequence. Key steps of the degradation pathway of t-anethole were identified by the use of t-anethole-blocked mutants and specific inducible enzy- matic activities. In addition to t-anethole, strain TA13

Eyal Shimoni; Timor Baasov; Uzi Ravid; Yuval Shoham

2002-01-01

206

Characterization of MRFAP1 turnover and interactions downstream of the NEDD8 pathway.  

PubMed

The NEDD8-Cullin E3 ligase pathway plays an important role in protein homeostasis, in particular the degradation of cell cycle regulators and transcriptional control networks. To characterize NEDD8-cullin target proteins, we performed a quantitative proteomic analysis of cells treated with MLN4924, a small molecule inhibitor of the NEDD8 conjugation pathway. MRFAP1 and its interaction partner, MORF4L1, were among the most up-regulated proteins after NEDD8 inhibition in multiple human cell lines. We show that MRFAP1 has a fast turnover rate in the absence of MLN4924 and is degraded via the ubiquitin-proteasome system. The increased abundance of MRFAP1 after MLN4924 treatment results from a decreased rate of degradation. Characterization of the binding partners of both MRFAP1 and MORF4L1 revealed a complex protein-protein interaction network. MRFAP1 bound to a number of E3 ubiquitin ligases, including CUL4B, but not to components of the NuA4 complex, including MRGBP, which bound to MORF4L1. These data indicate that MRFAP1 may regulate the ability of MORF4L1 to interact with chromatin-modifying enzymes by binding to MORF4L1 in a mutually exclusive manner with MRGBP. Analysis of MRFAP1 expression in human tissues by immunostaining with a MRFAP1-specific antibody revealed that it was detectable in only a small number of tissues, in particular testis and brain. Strikingly, analysis of the seminiferous tubules of the testis showed the highest nuclear staining in the spermatogonia and much weaker staining in the spermatocytes and spermatids. MRGBP was inversely correlated with MRFAP1 expression in these cell types, consistent with an exchange of MORF4L1 interaction partners as cells progress through meiosis in the testis. These data highlight an important new arm of the NEDD8-cullin pathway. PMID:22038470

Larance, Mark; Kirkwood, Kathryn J; Xirodimas, Dimitris P; Lundberg, Emma; Uhlen, Mathias; Lamond, Angus I

2012-03-01

207

Order and specificity of the Plasmodium falciparum hemoglobin degradation pathway.  

PubMed Central

The human malaria parasite, Plasmodium falciparum, degrades nearly all its host cell hemoglobin during a short segment of its intraerythrocytic development. This massive catabolic process occurs in an acidic organelle, the digestive vacuole. Aspartic and cysteine proteases have been implicated in this pathway. We have isolated three vacuolar proteases that account for most of the globin-degrading activity of the digestive vacuole. One is the previously described aspartic hemoglobinase that initiates hemoglobin degradation. A second aspartic protease is capable of cleaving hemoglobin with an overlapping specificity, but seems to prefer acid-denatured globin. The third is a cysteine protease that does not recognize native hemoglobin but readily cleaves denatured globin. It is synergistic with the aspartic hemoglobinase, both by in vitro assay of hemoglobin degradation, and by isobologram analysis of protease inhibitor-treated parasites in culture. The cysteine protease is highly sensitive to chloroquine-heme complex, suggesting a possible mechanism of 4-aminoquinoline antimalarial action. The data suggest an ordered pathway of hemoglobin catabolism that presents an excellent target for chemotherapy. Images PMID:8163662

Gluzman, I Y; Francis, S E; Oksman, A; Smith, C E; Duffin, K L; Goldberg, D E

1994-01-01

208

Oestrogen causes ATBF1 protein degradation through the oestrogen-responsive E3 ubiquitin ligase EFP.  

PubMed

We reported previously that the tumour suppressor ATBF1 (AT motif-binding factor 1) formed an autoregulatory feedback loop with oestrogen-ER? (oestrogen receptor ?) signalling to regulate oestrogen-dependent cell proliferation in breast cancer cells. In this loop ATBF1 inhibits the function of oestrogen-ER? signalling, whereas ATBF1 protein levels are fine-tuned by oestrogen-induced transcriptional up-regulation as well as UPP (ubiquitin-proteasome pathway)-mediated protein degradation. In the present study we show that EFP (oestrogen-responsive finger protein) is an E3 ubiquitin ligase mediating oestrogen-induced ATBF1 protein degradation. Knockdown of EFP increases ATBF1 protein levels, whereas overexpression of EFP decreases ATBF1 protein levels. EFP interacts with and ubiquitinates ATBF1 protein. Furthermore, we show that EFP is an important factor in oestrogen-induced ATBF1 protein degradation in which some other factors are also involved. In human primary breast tumours the levels of ATBF1 protein are positively correlated with the levels of EFP protein, as both are directly up-regulated ER? target gene products. However, the ratio of ATBF1 protein to EFP protein is negatively correlated with EFP protein levels. Functionally, ATBF1 antagonizes EFP-mediated cell proliferation. These findings not only establish EFP as the E3 ubiquitin ligase for oestrogen-induced ATBF1 protein degradation, but further support the autoregulatory feedback loop between ATBF1 and oestrogen-ER? signalling and thus implicate ATBF1 in oestrogen-dependent breast development and carcinogenesis. PMID:22452784

Dong, Xue-Yuan; Fu, Xiaoying; Fan, Songqing; Guo, Peng; Su, Dan; Dong, Jin-Tang

2012-06-15

209

Degradation of toluene-2,4-diamine by persulfate: kinetics, intermediates and degradation pathway.  

PubMed

Abstract In this study, the degradation of toluene-2,4-diamine (TDA) by persulfate (PS) in an aqueous solution at near-neutral pH was examined. The result showed that degradation rate of TDA increased with increasing PS concentrations. The optimal dosage of PS in the reaction system was determined by efficiency indicator (I) coupling in the consumption of PS and decay half-life of TDA. Calculation showed that 0.74?mM of PS was the most effective dosage for TDA degradation, at that level the maximum I of 24.51 was obtained. PS can oxidize TDA for an extended reaction time period. Under neutral condition without activation, four degradation intermediates, 2,4-diamino-3-hydroxy-5-sulfonicacidtoluene, 2,4-diaminobenzaldehyde, 2,4-bis (vinylamino) benzaldehyde, and 3,5-diamino-4-hydroxy-2-pentene were identified by high-performance liquid chromatography-mass spectrometry. The tentative degradation pathway of TDA was proposed as well. It was found that hydroxyl radical played an important role in degradation of TDA with activation of Fe(2+), whereas PS anion and sulfate radicals were responsible for the degradation without activation of Fe(2+). PMID:25442404

Jiang, Yong-Hai; Zhang, Jin-Bao; Xi, Bei-Dou; An, Da; Yang, Yu; Li, Ming-Xiao

2014-12-01

210

Degradation of phenazone in aqueous solution with ozone: influencing factors and degradation pathways.  

PubMed

Oxidation kinetics and degradation pathways of phenazone (an analgesic and antipyretic drug) upon reaction with O3 were investigated. Kinetic studies on degradation of phenazone were carried out under different operating conditions such as temperature, pH, anions and H2O2 addition. Results showed that the degradation followed the pseudo-first-order kinetic model. The reaction rate constant (kobs) of phenazone reached the maximum at 20 °C (9.653×10(-3) s(-1)). The presence of NO3(-) could enhance the degradation rate, while the addition of HCO3(-), SO4(2)(-), Cl(-) and the rise of pH showed negative effects on the ozonation of phenazone. H2O2 addition increased the phenazone degradation efficiency by 45.9% with the optimal concentration of 0.135 mM. Reaction by-products were evaluated by UPLC-Q-TOF-MS, which allowed the identification of a total of 10 by-products. The transformation pathways of phenazone ozonation consisted mainly of electrophilic addition and substitution, pyrazole ring opening, hydroxylation, dephenylization and coupling. The toxicity of these intermediate products showed that they are expected not to be more toxic than phenazone, with the exception of P7 (aniline) and P10 (1,5-dimethyl-4-((1-methyl-2-phenylhydrazinyl)methoxy)-2-phenyl-1H-pyrazol-3(2H)-one). PMID:25038548

Miao, Heng-Feng; Cao, Meng; Xu, Dan-Yao; Ren, Hong-Yan; Zhao, Ming-Xing; Huang, Zhen-Xing; Ruan, Wen-Quan

2015-01-01

211

Aerobic Degradation of Dinitrotoluenes and Pathway for Bacterial Degradation of 2,6-Dinitrotoluene  

PubMed Central

An oxidative pathway for the mineralization of 2,4-dinitrotoluene (2,4-DNT) by Burkholderia sp. strain DNT has been reported previously. We report here the isolation of additional strains with the ability to mineralize 2,4-DNT by the same pathway and the isolation and characterization of bacterial strains that mineralize 2,6-dinitrotoluene (2,6-DNT) by a different pathway. Burkholderia cepacia strain JS850 and Hydrogenophaga palleronii strain JS863 grew on 2,6-DNT as the sole source of carbon and nitrogen. The initial steps in the pathway for degradation of 2,6-DNT were determined by simultaneous induction, enzyme assays, and identification of metabolites through mass spectroscopy and nuclear magnetic resonance. 2,6-DNT was converted to 3-methyl-4-nitrocatechol by a dioxygenation reaction accompanied by the release of nitrite. 3-Methyl-4-nitrocatechol was the substrate for extradiol ring cleavage yielding 2-hydroxy-5-nitro-6-oxohepta-2,4-dienoic acid, which was converted to 2-hydroxy-5-nitropenta-2,4-dienoic acid. 2,4-DNT-degrading strains also converted 2,6-DNT to 3-methyl-4-nitrocatechol but did not metabolize the 3-methyl-4-nitrocatechol. Although 2,6-DNT prevented the degradation of 2,4-DNT by 2,4-DNT-degrading strains, the effect was not the result of inhibition of 2,4-DNT dioxygenase by 2,6-DNT or of 4-methyl-5-nitrocatechol monooxygenase by 3-methyl-4-nitrocatechol. PMID:10788393

Nishino, Shirley F.; Paoli, George C.; Spain, Jim C.

2000-01-01

212

Protein degradation and quality control in cells from laforin and malin knockout mice.  

PubMed

Lafora disease is a progressive myoclonus epilepsy caused by mutations in the EPM2A or EPM2B genes that encode a glycogen phosphatase, laforin, and an E3 ubiquitin ligase, malin, respectively. Lafora disease is characterized by accumulation of insoluble, poorly branched, hyperphosphorylated glycogen in brain, muscle, heart, and liver. The laforinmalin complex has been proposed to play a role in the regulation of glycogen metabolism and protein quality control. We evaluated three arms of the protein degradation/ quality control process (the autophago-lysosomal pathway, the ubiquitin-proteasomal pathway, and the endoplasmic reticulum (ER) stress response) in mouse embryonic fibroblasts from Epm2a(-/-), Epm2b(-/-), and Epm2a(-/-) Epm2b(-/-) mice. The levels of LC3-II, a marker of autophagy, were decreased in all knock-out cells as compared with wild type even though they still showed a slight response to starvation and rapamycin. Furthermore, ribosomal protein S6 kinase and S6 phosphorylation were increased. Under basal conditions there was no effect on the levels of ubiquitinated proteins in the knock-out cells, but ubiquitinated protein degradation was decreased during starvation or stress. Lack of malin (Epm2b(-/-) and Epm2a(-/-) Epm2b(-/-) cells) but not laforin (Epm2a(-/-) cells) decreased LAMP1, a lysosomal marker. CHOP expression was similar in wild type and knock-out cells under basal conditions or with ER stress-inducing agents. In conclusion, both laforin and malin knock-out cells display mTOR-dependent autophagy defects and reduced proteasomal activity but no defects in the ER stress response. We speculate that these defects may be secondary to glycogen overaccumulation. This study also suggests a malin function independent of laforin, possibly in lysosomal biogenesis and/or lysosomal glycogen disposal. PMID:24914213

Garyali, Punitee; Segvich, Dyann M; DePaoli-Roach, Anna A; Roach, Peter J

2014-07-25

213

Autophagy as a Regulated Pathway of Cellular Degradation  

NSDL National Science Digital Library

Macroautophagy is a dynamic process involving the rearrangement of subcellular membranes to sequester cytoplasm and organelles for delivery to the lysosome or vacuole where the sequestered cargo is degraded and recycled. This process takes place in all eukaryotic cells. It is highly regulated through the action of various kinases, phosphatases, and guanosine triphosphatases (GTPases). The core protein machinery that is necessary to drive formation and consumption of intermediates in the macroautophagy pathway includes a ubiquitin-like protein conjugation system and a protein complex that directs membrane docking and fusion at the lysosome or vacuole. Macroautophagy plays an important role in developmental processes, human disease, and cellular response to nutrient deprivation.

Daniel Klionsky (University of Michigan; Department of Biology)

2000-12-01

214

Degradation pathway(s) of chlorophyll: what has gene cloning revealed?  

PubMed

The mechanism responsible for the degreening of plants and the degradation of chlorophyll was unclear for many years. However, recent studies have identified the colorless intermediates and helped to construct a basic pathway for degradation. After the successive removal of phytol and Mg21 from the chlorophyll molecule by chlorophyllase and 'Mg dechelatase', pheophorbide a is cleaved and reduced to yield a colorless, open tetrapyrrole intermediate. After further modifications, this is finally transported to the vacuole. Cloning the genes for chlorophyllase isozymes and the reductase should help to elucidate the physiological roles of each enzyme at a molecular level. PMID:11044719

Takamiya, K I; Tsuchiya, T; Ohta, H

2000-10-01

215

A label-free quantitative proteomics strategy to identify E3 ubiquitin ligase substrates targeted to proteasome degradation.  

PubMed

The ubiquitin-proteasome system is a central mechanism for controlled proteolysis that regulates numerous cellular processes in eukaryotes. As such, defects in this system can contribute to disease pathogenesis. In this pathway, E3 ubiquitin ligases provide platforms for binding specific substrates, thereby coordinating their ubiquitylation and subsequent degradation by the proteasome. Despite the identification of many E3 ubiquitin ligases, the identities of their specific substrates are still largely unresolved. The ankyrin repeat-containing protein with a suppressor of cytokine signaling box 2 (ASB2) gene that we initially identified as a retinoic acid-response gene in acute promyelocytic leukemia cells encodes the specificity subunit of an E3 ubiquitin ligase complex that is involved in hematopoietic cell differentiation. We have recently identified filamin A and filamin B as the first ASB2 targets and shown that ASB2 triggers ubiquitylation and proteasome-mediated degradation of these proteins. Here a global quantitative proteomics strategy is provided to identify substrates of E3 ubiquitin ligases targeted to proteasomal degradation. Indeed we used label-free methods for quantifying proteins identified by shotgun proteomics in extracts of cells expressing wild-type ASB2 or an E3 ubiquitin ligase-defective mutant of ASB2 under the control of an inducible promoter. Measurements of spectral count and mass spectrometric signal intensity demonstrated a drastic decrease of filamin A and filamin B in myeloid leukemia cells expressing wild-type ASB2 compared with cells expressing an E3 ubiquitin ligase-defective mutant of ASB2. Altogether we provide an original strategy that enables identification of E3 ubiquitin ligase substrates that have to be degraded. PMID:19376791

Burande, Clara F; Heuzé, Mélina L; Lamsoul, Isabelle; Monsarrat, Bernard; Uttenweiler-Joseph, Sandrine; Lutz, Pierre G

2009-07-01

216

Emerging Mechanistic Insights into AAA Complexes Regulating Proteasomal Degradation  

PubMed Central

The 26S proteasome is an integral element of the ubiquitin-proteasome system (UPS) and, as such, responsible for regulated degradation of proteins in eukaryotic cells. It consists of the core particle, which catalyzes the proteolysis of substrates into small peptides, and the regulatory particle, which ensures specificity for a broad range of substrates. The heart of the regulatory particle is an AAA-ATPase unfoldase, which is surrounded by non-ATPase subunits enabling substrate recognition and processing. Cryo-EM-based studies revealed the molecular architecture of the 26S proteasome and its conformational rearrangements, providing insights into substrate recognition, commitment, deubiquitylation and unfolding. The cytosol proteasomal degradation of polyubiquitylated substrates is tuned by various associating cofactors, including deubiquitylating enzymes, ubiquitin ligases, shuttling ubiquitin receptors and the AAA-ATPase Cdc48/p97. Cdc48/p97 and its cofactors function upstream of the 26S proteasome, and their modular organization exhibits some striking analogies to the regulatory particle. In archaea PAN, the closest regulatory particle homolog and Cdc48 even have overlapping functions, underscoring their intricate relationship. Here, we review recent insights into the structure and dynamics of the 26S proteasome and its associated machinery, as well as our current structural knowledge on the Cdc48/p97 and its cofactors that function in the ubiquitin-proteasome system (UPS). PMID:25102382

Förster, Friedrich; Schuller, Jan M.; Unverdorben, Pia; Aufderheide, Antje

2014-01-01

217

Iodinated contrast media electro-degradation: Process performance and degradation pathways.  

PubMed

The electrochemical degradation of six of the most widely used iodinated contrast media was investigated. Batch experiments were performed under constant current conditions using two DSA® electrodes (titanium coated with a proprietary and patented mixed metal oxide solution of precious metals such as iridium, ruthenium, platinum, rhodium and tantalum). The degradation removal never fell below 85% (at a current density of 64mA/cm(2) with a reaction time of 150min) when perchlorate was used as the supporting electrolyte; however, when sulphate was used, the degradation performance was above 80% (at a current density of 64mA/cm(2) with a reaction time of 150min) for all of the compounds studied. Three main degradation pathways were identified, namely, the reductive de-iodination of the aromatic ring, the reduction of alkyl aromatic amides to simple amides and the de-acylation of N-aromatic amides to produce aromatic amines. However, as amidotrizoate is an aromatic carboxylate, this is added via the decarboxylation reaction. The investigation did not reveal toxicity except for the lower current density used, which has shown a modest toxicity, most likely for some reaction intermediates that are not further degraded. In order to obtain total removal of the contrast media, it was necessary to employ a current intensity between 118 and 182mA/cm(2) with energy consumption higher than 370kWh/m(3). Overall, the electrochemical degradation was revealed to be a reliable process for the treatment of iodinated contrast media that can be found in contaminated waters such as hospital wastewater or pharmaceutical waste-contaminated streams. PMID:25433384

Del Moro, Guido; Pastore, Carlo; Di Iaconi, Claudio; Mascolo, Giuseppe

2015-02-15

218

NP1EC Degradation Pathways Under Oxic and Microxic Conditions  

SciTech Connect

The degradation pathway of nonylphenol ethoxyacetic acid (NP1EC) and the conditions favoring CAP1EC formation were studied in aerobic microcosms constructed with soil from the Mesa soil aquifer treatment (SAT) facility (Arizona, USA) and pristine sediments from Coyote Creek (California, USA). In the Mesa microcosms, para-NP1EC was transformed to para-NP, before being rapidly transformed to nonyl alcohols via ipso-hydroxylation. While the formation of NP from APEMs has been observed by several researchers under anaerobic conditions, this is the first time the transient formation of NP from APEMs has been observed under aerobic conditions. Unlike the Mesa microcosms, large quantities of CAP1ECs were observed in the Coyote Creek microcosms. Initially, CA8P1ECs were the dominant metabolites, but as biodegradation continued, CA6P1ECs became the dominant metabolites. Compared to the CA8P1ECs, the number of CA6P1ECs peaks observed was small (<6) even though their concentrations were high. This suggests that several CA8P1ECs are degraded to only a few CA6P1EC isomers (i.e., the degradation pathway converges) or that some CA6P1EC metabolites are significantly more recalcitrant than others. The different biodegradation pathways observed in the Mesa and Coyote Creek microcosms result from the limited availability of dissolved oxygen in the Coyote Creek microcosms. In both sets of microcosms, the ortho isomers were transformed more slowly than the para isomers and in the Coyote Creek microcosms several ortho-CAP1ECs were observed. In addition, several unknown metabolites were observed in the Coyote Creek microcosms that were not seen in the abiotic or Mesa microcosms; these metabolites appear to be CAP1EC metabolites, have a -CH2-C6H4- fragment, and contain one carboxylic acid. Nitro-nonylphenol was observed in the Mesa microcosms, however, further experimentation illustrated that it was the product of an abiotic reaction between nitrite and nonylphenol under acidic conditions.

Montgomery-Brown, John; Li, Yongmei; Ding, Wang-Hsien; Mong, Gary M.; Campbell, James A.; Reinhard, Martin

2008-03-22

219

Necdin Promotes Ubiquitin-Dependent Degradation of PIAS1 SUMO E3 Ligase  

PubMed Central

Necdin, a pleiotropic protein that promotes differentiation and survival of mammalian neurons, is a member of MAGE (melanoma antigen) family proteins that share a highly conserved MAGE homology domain. Several MAGE proteins interact with ubiquitin E3 ligases and modulate their activities. However, it remains unknown whether MAGE family proteins interact with SUMO (small ubiquitin-like modifier) E3 ligases such as PIAS (protein inhibitor of activated STAT) family, Nsmce2/Mms21 and Cbx4/Pc2. In the present study, we examined whether necdin interacts with these SUMO E3 ligases. Co-immunoprecipitation analysis revealed that necdin, MAGED1, MAGEF1 and MAGEL2 bound to PIAS1 but not to Nsmce2 or Cbx4. These SUMO E3 ligases bound to MAGEA1 but failed to interact with necdin-like 2/MAGEG1. Necdin bound to PIAS1 central domains that are highly conserved among PIAS family proteins and suppressed PIAS1-dependent sumoylation of the substrates STAT1 and PML (promyelocytic leukemia protein). Remarkably, necdin promoted degradation of PIAS1 via the ubiquitin-proteasome pathway. In transfected HEK293A cells, amino- and carboxyl-terminally truncated mutants of PIAS1 bound to necdin but failed to undergo necdin-dependent ubiquitination. Both PIAS1 and necdin were associated with the nuclear matrix, where the PIAS1 terminal deletion mutants failed to localize, implying that the nuclear matrix is indispensable for necdin-dependent ubiquitination of PIAS1. Our data suggest that necdin suppresses PIAS1 both by inhibiting SUMO E3 ligase activity and by promoting ubiquitin-dependent degradation. PMID:24911587

Gur, Ibrahim; Fujiwara, Kazushiro; Hasegawa, Koichi; Yoshikawa, Kazuaki

2014-01-01

220

Evolution of catabolic pathways for synthetic compounds: bacterial pathways for degradation of 2,4-dinitrotoluene and nitrobenzene  

Microsoft Academic Search

The pathways for 2,4-dinitrotoluene (2,4-DNT) and nitrobenzene offer fine illustrations of how the ability to assimilate new carbon sources evolves in bacteria. Studies of the degradation pathways provide insight about two principal strategies for overcoming the metabolic block imposed by nitro- substituents on aromatic compounds. The 2,4-DNT pathway uses novel oxygenases for oxidative denitration and subsequent ring-fission. The nitrobenzene pathway

G. R. Johnson; J. C. Spain

2003-01-01

221

Arsenic trioxide induces programmed cell death through stimulation of ER stress and inhibition of the ubiquitin-proteasome system in human sarcoma cells.  

PubMed

Sarcoma is a rare form of cancer that differs from the much more common carcinomas because it occurs in a distinct type of tissue. Many patients of sarcoma have poor response to chemotherapy and an increased risk for local recurrence. Arsenic trioxide (ATO) is used to treat certain types of leukemia. Recently, data have revealed that ATO induces sarcoma cell death in several types of solid tumor cell lines. In the present study, we investigated whether ATO induces cancer cell death and elucidated the underlying anti-cancer mechanisms. Our results showed that ATO caused concentration- and time-dependent cell death in human osteosarcoma and fibrosarcoma cells. The types of cell death that were induced by ATO were primarily autophagy and apoptosis. Furthermore, ATO activated p38, JNK and AMPK and inhibited the Akt/mTOR signaling pathways. Specifically, we found that ATO induced endoplasmic reticulum (ER) stress and suppressed proteasome activation in two types of sarcoma cell lines. However, the level of proteasome inhibition in osteosarcoma cells was lower than in fibrosarcoma cells. Thus, we used combined treatment with ATO and a proteasome inhibitor to examine the antitumor activity in fibrosarcoma cells. The data indicated showed that the combination treatment of ATO and MG132 (a proteasome inhibitor) resulted in synergistic cytotoxicity. In a fibrosarcoma xenograft mouse model, the combined treatment significantly reduced tumor progression. Immunohistochemical studies revealed that combined treatment induced autophagy and apoptosis. In summary, our results suggest a potential clinical application of ATO in sarcoma therapy and that combined treatment with a proteasome inhibitor can increase the therapeutic efficacy. PMID:25449439

Chiu, Hui-Wen; Tseng, Yin-Chiu; Hsu, Yung-Ho; Lin, Yuh-Feng; Foo, Ning-Ping; Guo, How-Ran; Wang, Ying-Jan

2015-01-28

222

Degradation of diclofenac by ultrasonic irradiation: kinetic studies and degradation pathways.  

PubMed

Diclofenac (DCF) is a widely used anti-inflammatory drug found in various water bodies, posing threats to human health. In this research, the effects of ultrasonic irradiation at 585kHz on the degradation of DCF were studied under the air, oxygen, argon, and nitrogen saturated conditions. First, the dechlorination efficiencies under the air, oxygen, argon, and nitrogen saturated conditions were calculated to be 67%, 60%, 53% and 59%. Second, there was full mineralization of nitrogen during DCF degradation under the air, oxygen, and argon saturated conditions, but no mineralization of nitrogen under the nitrogen-saturated condition. Different from nitrogen, only partial mineralization of carbon occurred under the four gas-saturated conditions. Third, OH scavengers were added to derive the rate constants in the three reaction zones: cavitation bubble, supercritical interface, and bulk solution. Comparison of the constants indicated that DCF degradation was not limited to the bulk solution as conventionally assumed. Oxidation in the supercritical interface played a dominant role under the air and oxygen saturated conditions, while OH reactions in the cavitation bubble and/or bulk solution were dominant under the nitrogen and argon saturated conditions. After the addition of H2O2, reactions in the cavitation bubble and bulk solution kept their dominant roles under the nitrogen and argon saturated conditions, while reaction in the supercritical interface decreased under the air and oxygen saturated conditions. Finally, LC-MS analysis was used to derive the by-products and propose the main pathways of DCF degradation by ultrasonic irradiation. PMID:25065805

Nie, Er; Yang, Mo; Wang, Dong; Yang, Xiaoying; Luo, Xingzhang; Zheng, Zheng

2014-10-01

223

Integrated control of protein degradation in C. elegans muscle  

PubMed Central

Protein degradation is a fundamental cellular process, the genomic control of which is incompletely understood. The advent of transgene-coded reporter proteins has enabled the development of C. elegans into a model for studying this problem. The regulation of muscle protein degradation is surprisingly complex, integrating multiple signals from hypodermis, intestine, neurons and muscle itself. Within the muscle, degradation is executed by separately regulated autophagy-lysosomal, ubiquitin-proteasome and calpain-mediated systems. The signal-transduction mechanisms, in some instances, involve modules previously identified for their roles in developmental processes, repurposed in terminally differentiated muscle to regulate the activities of pre-formed proteins. Here we review the genes, and mechanisms, which appear to coordinately control protein degradation within C. elegans muscle. We also consider these mechanisms in the context of development, physiology, pathophysiology and disease models. PMID:23457662

Lehmann, Susann; Shephard, Freya; Jacobson, Lewis A.; Szewczyk, Nathaniel J.

2012-01-01

224

APC-targeted RAAI degradation mediates the cell cycle and root development in plants  

PubMed Central

Protein degradation by the ubiquitin-proteasome system is necessary for a normal cell cycle. As compared with knowledge of the mechanism in animals and yeast, that in plants is less known. Here we summarize research into the regulatory mechanism of protein degradation in the cell cycle in plants. Anaphase-promoting complex/cyclosome (APC), in the E3 family of enzymes, plays an important role in maintaining normal mitosis. APC activation and substrate specificity is determined by its activators, which can recognize the destruction box (D-box) in APC target proteins. Oryza sativa root architecture-associated I (OsRAA1) with GTP-binding activity was originally cloned from rice. Overexpression of of OsRAA1 inhibits the growth of primary roots in rice. Knockdown lines showed reduced height of seedlings because of abnormal cell division. OsRAA1 transgenic rice and fission yeast show a higher proportion of metaphase cells than that of controls, which suggests a blocked transition from metaphase to anaphase during mitosis. OsRAA1 co-localizes with spindle tubulin. It contains the D-box motif and interacts with OsRPT4 of the regulatory particle of 26S proteasome. OsRAA1 may be a cell cycle inhibitor that can be degraded by the ubiquitin-proteasome system, and its disruption is necessary for the transition from metaphase to anaphase during root growth in rice. PMID:20037474

Cao, Hong; Chong, Kang

2010-01-01

225

Genome-scale analyses of butanol tolerance in Saccharomyces cerevisiae reveal an essential role of protein degradation  

PubMed Central

Background n-Butanol and isobutanol produced from biomass-derived sugars are promising renewable transport fuels and solvents. Saccharomyces cerevisiae has been engineered for butanol production, but its high butanol sensitivity poses an upper limit to product titers that can be reached by further pathway engineering. A better understanding of the molecular basis of butanol stress and tolerance of S. cerevisiae is important for achieving improved tolerance. Results By combining a screening of the haploid S. cerevisiae knock-out library, gene overexpression, and genome analysis of evolutionary engineered n-butanol-tolerant strains, we established that protein degradation plays an essential role in tolerance. Strains deleted in genes involved in the ubiquitin-proteasome system and in vacuolar degradation of damaged proteins showed hypersensitivity to n-butanol. Overexpression of YLR224W, encoding the subunit responsible for the recognition of damaged proteins of an ubiquitin ligase complex, resulted in a strain with a higher n-butanol tolerance. Two independently evolved n-butanol-tolerant strains carried different mutations in both RPN4 and RTG1, which encode transcription factors involved in the expression of proteasome and peroxisomal genes, respectively. Introduction of these mutated alleles in the reference strain increased butanol tolerance, confirming their relevance in the higher tolerance phenotype. The evolved strains, in addition to n-butanol, were also more tolerant to 2-butanol, isobutanol and 1-propanol, indicating a common molecular basis for sensitivity and tolerance to C3 and C4 alcohols. Conclusions This study shows that maintenance of protein integrity plays an essential role in butanol tolerance and demonstrates new promising targets to engineer S. cerevisiae for improved tolerance. PMID:23552365

2013-01-01

226

Mitofusin 1 degradation is induced by a disruptor of mitochondrial calcium homeostasis, CGP37157: a role in apoptosis in prostate cancer cells.  

PubMed

Mitochondria constantly divide (mitochondrial fission) and fuse (mitochondrial fusion) in a normal cell. Disturbances in the balance between these two physiological processes may lead to cell dysfunction or to cell death. Induction of cell death is the prime goal of prostate cancer chemotherapy. Our previous study demonstrated that androgens increase the expression of a mitochondrial protein involved in fission and facilitate an apoptotic response to CGP37157 (CGP), an inhibitor of mitochondrial calcium efflux, in prostate cancer cells. However, the regulation and role of mitochondrial fusion proteins in the death of these cells have not been examined. Therefore, our objective was to investigate the effect of CGP on a key mitochondrial fusion protein, mitofusin 1 (Mfn1), and the role of Mfn1 in prostate cancer cell apoptosis. We used various prostate cancer cell lines and western blot analysis, qRT-PCR, siRNA, M30 apoptosis assay and immunoprecipitation techniques to determine mechanisms regulating Mfn1. Treatment of prostate cancer cells with CGP resulted in selective degradation of Mfn1. Mfn1 ubiquitination was detected following immunoprecipitation of overexpressed Myc-tagged Mfn1 protein from CGP-treated cells, and treatment with the proteasomal inhibitor lactacystin, as well as siRNA-mediated knockdown of the E3 ubiquitin ligase March5, protected Mfn1 from CGP-induced degradation. These data indicate the involvement of the ubiquitin-proteasome pathway in CGP-induced degradation of Mfn1. We also demonstrated that downregulation of Mfn1 by siRNA enhanced the apoptotic response of LNCaP cells to CGP, suggesting a likely pro-survival role for Mfn1 in these cells. Our results suggest that manipulation of mitofusins may provide a novel therapeutic advantage in treating prostate cancer. PMID:24626641

Choudhary, Vivek; Kaddour-Djebbar, Ismail; Alaisami, Rabei; Kumar, M Vijay; Bollag, Wendy B

2014-05-01

227

Supplementation of carnitine leads to an activation of the IGF-1/PI3K/Akt signalling pathway and down regulates the E3 ligase MuRF1 in skeletal muscle of rats  

PubMed Central

Background Recently, it has been shown that carnitine down-regulates genes involved in the ubiquitin-proteasome system (UPS) in muscle of pigs and rats. The mechanisms underlying this observation are yet unknown. Based on the previous finding that carnitine increases plasma IGF-1 concentration, we investigated the hypothesis that carnitine down-regulates genes of the UPS by modulation of the of the IGF-1/PI3K/Akt signalling pathway which is an important regulator of UPS activity in muscle. Methods Male Sprague–Dawley rats, aged four weeks, were fed either a control diet with a low native carnitine concentration or the same diet supplemented with carnitine (1250 mg/kg diet) for four weeks. Components of the UPS and IGF-1/PI3K/Akt signalling pathway in skeletal muscle were examined. Results Rats fed the diet supplemented with carnitine had lower mRNA and protein levels of MuRF1, the most important E3 ubiquitin ligase in muscle, decreased concentrations of ubiquitin-protein conjugates in skeletal muscle and higher IGF-1 concentration in plasma than control rats (P < 0.05). Moreover, in skeletal muscle of rats fed the diet supplemented with carnitine there was an activation of the PI3K/Akt signalling pathway, as indicated by increased protein levels of phosphorylated (activated) Akt1 (P < 0.05). Conclusion The present study shows that supplementation of carnitine markedly decreases the expression of MuRF1 and concentrations of ubiquitinated proteins in skeletal muscle of rats, indicating a diminished degradation of myofibrillar proteins by the UPS. The study moreover shows that supplementation of carnitine leads to an activation of the IGF-1/PI3K/Akt signalling pathway which in turn might contribute to the observed down-regulation of MuRF1 and muscle protein ubiquitination. PMID:23497226

2013-01-01

228

Isolation of an isocarbophos-degrading strain of Arthrobacter sp. scl-2 and identification of the degradation pathway.  

PubMed

Isocarbophos is a widely used organophosphorus insecticide that has caused environmental pollution in many areas. However, degradation of isocarbophos by pure cultures has not been extensively studied, and the degradation pathway has not been determined. In this paper, a highly effective isocarbophos-degrading strain, scl-2, was isolated from isocarbophos-polluted soil. Strain scl-2 was preliminarily identified as Arthrobacter sp. based on its morphological, physiological, and biochemical properties, as well as 16S rDNA analysis. Strain scl-2 could utilize isocarbophos as its sole source of carbon and phosphorus for growth. One hundred mg/l isocarbophos could be degraded to a nondetectable level in 18 h by scl-2 in cell culture, and isofenphos-methyl, profenofos, and phosmet could also be degraded. During the degradation of isocarbophos, the metabolites isopropyl salicylate, salicylate, and gentisate were detected and identified based on MS/MS analysis and their retention times in HPLC. Transformation of gentisate to pyruvate and fumarate via maleylpyruvate and fumarylpyruvate was detected by assaying for the activities of gentisate 1,2- dioxygenase (GDO) and maleylpyruvate isomerase. Therefore, we have identified the degradation pathway of isocarbophos in Arthrobacter sp. scl-2 for the first time. This study highlights an important potential use of the strain scl-2 for the cleanup of environmental contamination by isocarbophos and presents a mechanism of isocarbophos metabolism. PMID:19996699

Rong, Li; Guo, Xinqiang; Chen, Kai; Zhu, Jianchun; Li, Shunpeng; Jiang, Jiandong

2009-11-01

229

Reaction pathways in the electrochemical degradation of thiocarbamate herbicides in NaCl solution  

Microsoft Academic Search

We have identified the intermediates and end products which are formed during the electrolytic degradation of thiocarbamate pesticides in aqueous NaCl solutions and investigated how the intermediate and end product volumes and ratios depend on reaction conditions. Further, we have defined both the reaction pathways leading to intermediate and end product formation and the methods affecting this process. The degradation

FERENC MOGYORÓDY

2006-01-01

230

Proteasome Inhibitors Activate Autophagy Involving Inhibition of PI3K-Akt-mTOR Pathway as an Anti-Oxidation Defense in Human RPE Cells  

PubMed Central

The two major intracellular protein degradation systems, the ubiquitin-proteasome system (UPS) and autophagy, work collaboratively in many biological processes including development, apoptosis, aging, and countering oxidative injuries. We report here that, in human retinal pigment epithelial cells (RPE), ARPE-19 cells, proteasome inhibitors, clasto-lactacystin?-lactone (LA) or epoxomicin (Epo), at non-lethal doses, increased the protein levels of autophagy-specific genes Atg5 and Atg7 and enhanced the conversion of microtubule-associated protein light chain (LC3) from LC3-I to its lipidative form, LC3-II, which was enhanced by co-addition of the saturated concentration of Bafilomycin A1 (Baf). Detection of co-localization for LC3 staining and labeled-lysosome further confirmed autophagic flux induced by LA or Epo. LA or Epo reduced the phosphorylation of the protein kinase B (Akt), a downstream target of phosphatidylinositol-3-kinases (PI3K), and mammalian target of rapamycin (mTOR) in ARPE-19 cells; by contrast, the induced changes of autophagy substrate, p62, showed biphasic pattern. The autophagy inhibitor, Baf, attenuated the reduction in oxidative injury conferred by treatment with low doses of LA and Epo in ARPE-19 cells exposed to menadione (VK3) or 4-hydroxynonenal (4-HNE). Knockdown of Atg7 with siRNA in ARPE-19 cells reduced the protective effects of LA or Epo against VK3. Overall, our results suggest that treatment with low levels of proteasome inhibitors confers resistance to oxidative injury by a pathway involving inhibition of the PI3K-Akt-mTOR pathway and activation of autophagy. PMID:25062253

Sun, Lin; Li, Yiping; Qu, Jia; Snider, Barbara Joy; Wu, Shengzhou

2014-01-01

231

Molecular mechanisms and signaling pathways of angiotensin II-induced muscle wasting: potential therapeutic targets for cardiac cachexia  

PubMed Central

Cachexia is a serious complication of many chronic diseases, such as congestive heart failure (CHF) and chronic kidney disease (CKD). Many factors are involved in the development of cachexia, and there is increasing evidence that angiotensin II (Ang II), the main effector molecule of the renin-angiotensin system (RAS), plays an important role in this process. Patients with advanced CHF or CKD often have increased Ang II levels and cachexia, and angiotensin-converting enzyme (ACE) inhibitor treatment improves weight loss. In rodent models, an increase in systemic Ang II leads to weight loss through increased protein breakdown, reduced protein synthesis in skeletal muscle and decreased appetite. Ang II activates the ubiquitin-proteasome system via generation of reactive oxygen species and via inhibition of the insulin-like growth factor-1 signaling pathway. Furthermore, Ang II inhibits 5? AMP-activated protein kinase (AMPK) activity and disrupts normal energy balance. Ang II also increases cytokines and circulating hormones such as tumor necrosis factor-?, interleukin-6, serum amyloid-A, glucocorticoids and myostatin, which regulate muscle protein synthesis and degradation. Ang II acts on hypothalamic neurons to regulate orexigenic/anorexigenic neuropeptides, such as neuropeptide-Y, orexin and corticotropin-releasing hormone, leading to reduced appetite. Also, Ang II may regulate skeletal muscle regenerative processes. Several clinical studies have indicated that blockade of Ang II signaling via ACE inhibitors or Ang II type 1 receptor blockers prevents weight loss and improves muscle strength. Thus the RAS is a promising target for the treatment of muscle atrophy in patients with CHF and CKD. PMID:23769949

Yoshida, Tadashi; Tabony, A. Michael; Galvez, Sarah; Mitch, William E.; Higashi, Yusuke; Sukhanov, Sergiy; Delafontaine, Patrice

2013-01-01

232

Synergetic effect of alkaline earth metal oxides and iron oxides on the degradation of hexachlorobenzene and its degradation pathway.  

PubMed

The degradation of hexachlorobenzene (HCB) was carried out over physical mixtures of a series of alkaline earth metal oxides (MO: M=Mg, Ca, Sr, Ba) and iron oxides with different crystal types (Fe(x)O(y):Fe(2)O(3) or Fe(3)O(4)) at 300°C. These physical mixtures all showed a synergetic effect toward the degradation of HCB. A range of degradation products were identified by various methods, including tri- to penta-chlorobenzenes by gas chromatography/mass spectrometry (GC-MS), tri- to penta-chlorophenols, tetrachlorocatechol (TCC) and tetrachlorohydroquinone (TCHQ) by GC-MS after derivatization, and formic and acetic acids by ion chromatography. Two degradation pathways, hydrodechlorination and oxidative degradation, appear to occur competitively. However, more sequential chlorinated benzene and phenol congeners were formed over mixed MO/Fe(3)O(4) than over mixed MO/Fe(2)O(3) under the same conditions. The oxidative reaction dominated over mixed MO/Fe(2)O(3) and was promoted as the major reaction by the synergetic effect, while both the oxidative and hydrodechlorination reactions were important over mixed MO/Fe(3)O(4), and both pathways are remarkably promoted by the synergetic effect. The enhanced hydrodechlorination may be attributed to free electrons generated by the transformation of Fe(3)O(4) into Fe(2)O(3), and hydrogen provided by water adsorbed on the MO. PMID:22939896

Su, Guijin; Liu, Yexuan; Huang, Linyan; Shi, Yali; Zhang, Aiqian; Zhang, Lixia; Liu, Wenbin; Gao, Lirong; Zheng, Minghui

2013-01-01

233

Evolution of efficient pathways for degradation of anthropogenic chemicals  

Microsoft Academic Search

Anthropogenic compounds used as pesticides, solvents and explosives often persist in the environment and can cause toxicity to humans and wildlife. The persistence of anthropogenic compounds is due to their recent introduction into the environment; microbes in soil and water have had relatively little time to evolve efficient mechanisms for degradation of these new compounds. Some anthropogenic compounds are easily

Shelley D Copley

2009-01-01

234

PHOSPHOLIPIDS OF FIVE PSEUDOMONAD ARCHETYPES FOR DIFFERENT TOLUENE DEGRADATION PATHWAYS  

EPA Science Inventory

Liquid chromatography/electrospray ionization/mass spectrometry (LC/ESI/MS) was used to determine phospholipid profiles for five reference pseudomonad strains harboring distinct toluene catabolic pathways: Pseudomonas putida mt-2, Pseudomonas putida F1, Burkholderia cepacia G4, B...

235

Accumulation of mutant huntingtin fragments in aggresome-like inclusion bodies as a result of insufficient protein degradation.  

PubMed

The huntingtin exon 1 proteins with a polyglutamine repeat in the pathological range (51 or 83 glutamines), but not with a polyglutamine tract in the normal range (20 glutamines), form aggresome-like perinuclear inclusions in human 293 Tet-Off cells. These structures contain aggregated, ubiquitinated huntingtin exon 1 protein with a characteristic fibrillar morphology. Inclusion bodies with truncated huntingtin protein are formed at centrosomes and are surrounded by vimentin filaments. Inhibition of proteasome activity resulted in a twofold increase in the amount of ubiquitinated, SDS-resistant aggregates, indicating that inclusion bodies accumulate when the capacity of the ubiquitin-proteasome system to degrade aggregation-prone huntingtin protein is exhausted. Immunofluorescence and electron microscopy with immunogold labeling revealed that the 20S, 19S, and 11S subunits of the 26S proteasome, the molecular chaperones BiP/GRP78, Hsp70, and Hsp40, as well as the RNA-binding protein TIA-1, the potential chaperone 14-3-3, and alpha-synuclein colocalize with the perinuclear inclusions. In 293 Tet-Off cells, inclusion body formation also resulted in cell toxicity and dramatic ultrastructural changes such as indentations and disruption of the nuclear envelope. Concentration of mitochondria around the inclusions and cytoplasmic vacuolation were also observed. Together these findings support the hypothesis that the ATP-dependent ubiquitin-proteasome system is a potential target for therapeutic interventions in glutamine repeat disorders. PMID:11359930

Waelter, S; Boeddrich, A; Lurz, R; Scherzinger, E; Lueder, G; Lehrach, H; Wanker, E E

2001-05-01

236

AEROBIC DEGRADATION OF DINITROTOLUENES AND PATHWAY FOR BACTERIAL DEGRADATION OF 2,6-DINITROTOLUENE  

EPA Science Inventory

An oxidative pathway for the mineralization of 2,4-dinitrotoluene (2,4-DNT) by Burkhoderia sp. strain DNT has been reported previously. We report here the isolation of additional strains with the ability to mineralize dinitrotoluene (2,6-DNT) by a different pathway. Burkhoderia ...

237

Timely Activation of Budding Yeast APCCdh1 Involves Degradation of Its Inhibitor, Acm1, by an Unconventional Proteolytic Mechanism  

PubMed Central

Regulated proteolysis mediated by the ubiquitin proteasome system is a fundamental and essential feature of the eukaryotic cell division cycle. Most proteins with cell cycle-regulated stability are targeted for degradation by one of two related ubiquitin ligases, the Skp1-cullin-F box protein (SCF) complex or the anaphase-promoting complex (APC). Here we describe an unconventional cell cycle-regulated proteolytic mechanism that acts on the Acm1 protein, an inhibitor of the APC activator Cdh1 in budding yeast. Although Acm1 can be recognized as a substrate by the Cdc20-activated APC (APCCdc20) in anaphase, APCCdc20 is neither necessary nor sufficient for complete Acm1 degradation at the end of mitosis. An APC-independent, but 26S proteasome-dependent, mechanism is sufficient for complete Acm1 clearance from late mitotic and G1 cells. Surprisingly, this mechanism appears distinct from the canonical ubiquitin targeting pathway, exhibiting several features of ubiquitin-independent proteasomal degradation. For example, Acm1 degradation in G1 requires neither lysine residues in Acm1 nor assembly of polyubiquitin chains. Acm1 was stabilized though by conditional inactivation of the ubiquitin activating enzyme Uba1, implying some requirement for the ubiquitin pathway, either direct or indirect. We identified an amino terminal predicted disordered region in Acm1 that contributes to its proteolysis in G1. Although ubiquitin-independent proteasome substrates have been described, Acm1 appears unique in that its sensitivity to this mechanism is strictly cell cycle-regulated via cyclin-dependent kinase (Cdk) phosphorylation. As a result, Acm1 expression is limited to the cell cycle window in which Cdk is active. We provide evidence that failure to eliminate Acm1 impairs activation of APCCdh1 at mitotic exit, justifying its strict regulation by cell cycle-dependent transcription and proteolytic mechanisms. Importantly, our results reveal that strict cell-cycle expression profiles can be established independent of proteolysis mediated by the APC and SCF enzymes. PMID:25072887

Melesse, Michael; Choi, Eunyoung; Hall, Hana; Walsh, Michael J.; Geer, M. Ariel; Hall, Mark C.

2014-01-01

238

Study of Biochemical Pathways and Enzymes Involved in Pyrene Degradation by Mycobacterium sp. Strain KMS?  

PubMed Central

Pyrene degradation is known in bacteria. In this study, Mycobacterium sp. strain KMS was used to study the metabolites produced during, and enzymes involved in, pyrene degradation. Several key metabolites, including pyrene-4,5-dione, cis-4,5-pyrene-dihydrodiol, phenanthrene-4,5-dicarboxylic acid, and 4-phenanthroic acid, were identified during pyrene degradation. Pyrene-4,5-dione, which accumulates as an end product in some gram-negative bacterial cultures, was further utilized and degraded by Mycobacterium sp. strain KMS. Enzymes involved in pyrene degradation by Mycobacterium sp. strain KMS were studied, using 2-D gel electrophoresis. The first protein in the catabolic pathway, aromatic-ring-hydroxylating dioxygenase, which oxidizes pyrene to cis-4,5-pyrene-dihydrodiol, was induced with the addition of pyrene and pyrene-4,5-dione to the cultures. The subcomponents of dioxygenase, including the alpha and beta subunits, 4Fe-4S ferredoxin, and the Rieske (2Fe-2S) region, were all induced. Other proteins responsible for further pyrene degradation, such as dihydrodiol dehydrogenase, oxidoreductase, and epoxide hydrolase, were also found to be significantly induced by the presence of pyrene and pyrene-4,5-dione. Several nonpathway-related proteins, including sterol-binding protein and cytochrome P450, were induced. A pyrene degradation pathway for Mycobacterium sp. strain KMS was proposed and confirmed by proteomic study by identifying almost all the enzymes required during the initial steps of pyrene degradation. PMID:17041157

Liang, Yanna; Gardner, Dale R.; Miller, Charles D.; Chen, Dong; Anderson, Anne J.; Weimer, Bart C.; Sims, Ronald C.

2006-01-01

239

Eukaryotic starch degradation: integration of plastidial and cytosolic pathways.  

PubMed

Starch is an important plant product widely used as a nutrient, as a source of renewable energy, and for many technological applications. In plants, starch is the almost ubiquitous storage carbohydrate whereas most heterotrophic prokaryotes and eukaryotes rely on glycogen. Despite close similarities in basic chemical features, starch and glycogen differ in both structural and physicochemical properties. Glycogen is a hydrosoluble macromolecule with evenly distributed branching points. Starch exists as a water-insoluble particle having a defined (and evolutionary conserved) internal structure. The biochemistry of starch requires the co-operation of up to 40 distinct (iso)enzymes whilst approximately 10 (iso)enzymes permit glycogen metabolism. The biosynthesis and degradation of native starch include the transition of carbohydrates from the soluble to the solid phase and vice versa. In this review, two novel aspects of the eukaryotic plastidial starch degradation are discussed: Firstly, biochemical reactions that take place at the surface of particulate glucans and mediate the phase transition of carbohydrates. Secondly, processes that occur downstream of the export of starch-derived sugars into the cytosol. Degradation of transitory starch mainly results in the formation of neutral sugars, such as glucose and maltose, that are transported into the cytosol via the respective translocators. The cytosolic metabolism of the neutral sugars includes the action of a hexokinase, a phosphoglucomutase, and a transglucosidase that utilizes high molecular weight glycans as a transient glucosyl acceptor or donor. Data are included on the transglucosidase (disproportionating isozyme 2) in Cyanophora paradoxa that accumulates storage carbohydrates in the cytosol rather than in the plastid. PMID:19325165

Fettke, Joerg; Hejazi, Mahdi; Smirnova, Julia; Höchel, Erik; Stage, Marion; Steup, Martin

2009-01-01

240

Degradation of Phenol via Meta Cleavage Pathway by Pseudomonas fluorescens PU1  

PubMed Central

Degradation of phenolics by members of soil microflora is an important means by which these substances are removed from the environment thus reducing environmental pollution. Biodegradation by microorganisms offers unique opportunities to destroy or render phenolic compounds. A bacterium, PU1, identified as Pseudomonas fluorescens PU1, was investigated for its ability to grow on and degrade phenols as sole carbon sources in aerobic shaking batch culture. The organism degraded up to 1000?ppm of phenol using meta cleavage pathway. The pathways for phenol degradation were proposed by the identification of metabolites and assay of ring cleavage enzymes in cell extracts. Phenol was degraded via catechol with subsequent metaring cleavage. Cell growth increased as the phenol concentrations increased up to 1000 ppm phenol. The biodegradation efficiency, degradation extent, and metabolic pathway of phenol were determined to provide useful clues for further application of this isolate in the engineered bioremediation systems. The paper's results suggest that Pseudomonas fluorescens PU1 strain could be a good candidate for remediation of phenol contaminants from heavily polluted sites. PMID:23724329

Mahiudddin, Md.; Fakhruddin, A. N. M.; Abdullah-Al-Mahin

2012-01-01

241

Photocatalytic degradation of methylene blue with a nanocomposite system: synthesis, photocatalysis and degradation pathways.  

PubMed

Three different composites, including a calcined FeOOH supported ZnAl layered double hydroxide (FeOOH-LDO), a calcined ZnAl layered double hydroxide (ZnAl-LDO) and a calcined ZnFeAl layered double hydroxide (ZnFeAl-LDO), were synthesized via a sol-gel method, and their activity for the visible light photocatalytic degradation of methylene blue (MB) was studied. The composites were characterized by PXRD, SEM, and BET techniques, confirming the formation of highly crystalline structures. The activity performance of MB degradation was in the following order: FeOOH-LDO (?95%) > ZnFeAl-LDO (?60%) > ZnAl-LDO (?23%). In addition, a possible photocatalytic degradation reaction mechanism for MB was also proposed. Moreover, the frontier electron densities on the atoms of MB were calculated, which were in satisfactory agreement with the postulated mechanism. PMID:25611297

Xia, Shengjie; Zhang, Lianyang; Pan, Guoxiang; Qian, Pingping; Ni, Zheming

2015-02-01

242

Photolysis of chlorantraniliprole and cyantraniliprole in water and soil: verification of degradation pathways via kinetics modeling.  

PubMed

Photodegradation of [(14)C]-chlorantraniliprole (CLAP) and [(14)C]-cyantraniliprole (CNAP) was investigated in sterile buffer solutions, in natural water, and on soil surfaces. Both compounds displayed rapid degradation in aqueous buffers when exposed to light at concentrations which could result from direct overspray to a shallow water body. While the main products observed had analogous structures, a substantial difference was noted in the rate of degradation of the two compounds despite minimal differences in their structures. Transformations observed were primarily intramolecular rearrangements and degradations resulting from addition of hydroxyl radicals leading to molecular cleavage. Some of the degradation products were transient, and several degradates had isomeric molecular compositions. The sequence of transformations was established definitively with the help of kinetics modeling. Utility of kinetics analysis in verification of the proposed pathways is illustrated. PMID:24971760

Sharma, Ashok K; Zimmerman, William T; Singles, Suzanne K; Malekani, Kalumbu; Swain, Scott; Ryan, David; Mcquorcodale, Gordon; Wardrope, Laura

2014-07-16

243

Vocal Pathway Degradation in Gonadectomized Xenopus laevis Adults  

PubMed Central

Reproductive behaviors of many vertebrate species are activated in adult males by elevated androgen levels and abolished by castration. Neural and muscular components controlling these behaviors contain numerous hormone-sensitive sites including motor initiation centers (such as the basal ganglia), central pattern generators (CPGs), and muscles; therefore it is difficult to confirm the role of each hormone-activated target using behavioral assays alone. Our goal was to address this issue by determining the site of androgen-induced vocal activation using male Xenopus laevis, a species in which androgen dependence of vocal activation has been previously determined. We compared in vivo calling patterns and functionality of two in vitro preparations—the isolated larynx and an isolated brain from which fictive courtship vocalizations can be evoked—in castrated and control males. The isolated larynx allowed us to test whether castrated males were capable of transducing male-typical nerve signals into vocalizations and the fictively vocalizing brain preparation allowed us to directly examine vocal CPG function separate from the issue of vocal initiation. The results indicate that all three components—vocal initiation, CPG, and larynx—require intact gonads. Vocal production decreased dramatically in castrates and laryngeal contractile properties of castrated males were demasculinized, whereas no changes were observed in control animals. In addition, fictive calls of castrates were degraded compared with those of controls. To our knowledge, this finding represents the first demonstration of gonad-dependent maintenance of a CPG for courtship behavior in adulthood. Because previous studies showed that androgen-replacement can prevent castration-induced vocal impairments, we conclude that degradation of vocal initiation centers, larynx, and CPG function are most likely due to steroid hormone deprivation. PMID:21148092

Yamaguchi, Ayako

2011-01-01

244

Unveiling New Degradation Intermediates/Pathways from the Photocatalytic Degradation of Microcystin-LR  

EPA Science Inventory

This study focuses on the identification of reaction intermediates formed during the photocatalytic degradation of the cyanotoxin microcystin-LR with immobilized TiO2 Tphotocatalysts at neutral pH. To differentiate between impurities already existing in the MC-LR stand...

245

HDAC6 rescues neurodegeneration and provides an essential link between autophagy and the UPS  

Microsoft Academic Search

A prominent feature of late-onset neurodegenerative diseases is accumulation of misfolded protein in vulnerable neurons. When levels of misfolded protein overwhelm degradative pathways, the result is cellular toxicity and neurodegeneration. Cellular mechanisms for degrading misfolded protein include the ubiquitin-proteasome system (UPS), the main non-lysosomal degradative pathway for ubiquitinated proteins, and autophagy, a lysosome-mediated degradative pathway. The UPS and autophagy have

Udai Bhan Pandey; Zhiping Nie; Yakup Batlevi; Brett A. McCray; Gillian P. Ritson; Natalia B. Nedelsky; Stephanie L. Schwartz; Nicholas A. Diprospero; Melanie A. Knight; Oren Schuldiner; Ranjani Padmanabhan; Marc Hild; Deborah L. Berry; Dan Garza; Charlotte C. Hubbert; Tso-Pang Yao; Eric H. Baehrecke; J. Paul Taylor

2007-01-01

246

Degradation pathway of malachite green in a novel dual-tank photoelectrochemical catalytic reactor.  

PubMed

A novel dual-tank photoelectrochemical catalytic reactor was designed to investigate the degradation pathway of malachite green. A thermally formed TiO?/Ti thin film electrode was used as photoanode, graphite was used as cathode, and a saturated calomel electrode was employed as the reference electrode in the reactor. In the reactor, the anode and cathode tanks were connected by a cation exchange membrane. Results showed that the decolorization ratio of malachite green in the anode and cathode was 98.5 and 96.5% after 120 min, respectively. Malachite green in the two anode and cathode tanks was oxidized, achieving the bipolar double effect. Malachite green in both the anode and cathode tanks exhibited similar catalytic degradation pathways. The double bond of the malachite green molecule was attacked by strong oxidative hydroxyl radicals, after which the organic compound was degraded by the two pathways into 4,4-bis(dimethylamino) benzophenone, 4-(dimethylamino) benzophenone, 4-(dimethylamino) phenol, and other intermediate products. Eventually, malachite green was degraded into oxalic acid as a small molecular organic acid, which was degraded by processes such as demethylation, deamination, nitration, substitution, addition, and other reactions. PMID:23811633

Diao, Zenghui; Li, Mingyu; Zeng, Fanyin; Song, Lin; Qiu, Rongliang

2013-09-15

247

Proteasome Inhibition Enhances the Induction and Impairs the Maintenance of Late-Phase Long-Term Potentiation  

ERIC Educational Resources Information Center

Protein degradation by the ubiquitin-proteasome pathway plays important roles in synaptic plasticity, but the molecular mechanisms by which proteolysis regulates synaptic strength are not well understood. We investigated the role of the proteasome in hippocampal late-phase long-term potentiation (L-LTP), a model for enduring synaptic plasticity.…

Dong, Chenghai; Upadhya, Sudarshan C.; Ding, Lan; Smith, Thuy K.; Hegde, Ashok N.

2008-01-01

248

Distinct expression patterns of the E3 ligase SIAH-1 and its partner Kid\\/KIF22 in normal tissues and in the breast tumoral processes  

Microsoft Academic Search

SIAH proteins are the human members of an highly conserved family of E3 ubiquitin ligases. Several data suggest that SIAH proteins may have a role in tumor suppression and apoptosis. Previously, we reported that SIAH-1 induces the degradation of Kid (KIF22), a chromokinesin protein implicated in the normal progression of mitosis and meiosis, by the ubiquitin proteasome pathway. In human

Heriberto Bruzzoni-Giovanelli; Plinio Fernandez; Lucía Veiga; Marie-Pierre Podgorniak; Darren J Powell; Marco M Candeias; Samia Mourah; Fabien Calvo; Mónica Marín

2010-01-01

249

Muscle wasting in diabetic and in tumor-bearing rats: Role of oxidative stress  

Microsoft Academic Search

Cachexia is a debilitating syndrome characterized by body weight loss, muscle wasting, and anemia. Muscle wasting results from an altered balance between protein synthesis and degradation rates. Reactive oxygen species are indicated as crucial players in the onset of muscle protein hypercatabolism by upregulating elements of the ubiquitin–proteasome pathway. The present study has been aimed at evaluating comparatively the involvement

Raffaella Mastrocola; Patrizia Reffo; Fabio Penna; Chiara Elena Tomasinelli; Giuseppe Boccuzzi; Francesco Maria Baccino; Manuela Aragno; Paola Costelli

2008-01-01

250

2011LandesBioscience. Donotdistribute.  

E-print Network

.1093/hmg/ddr048. Key words: Parkinson disease, parkin, ubiquitin-proteasome system, mitophagy, mitochondria). Previous studies have estab- lished its role in mitophagy, a pathway whereby dysfunctional mitochondria are targeted for autophagic degradation. We recently reported that a major function of Parkin in dysfunctional

Chan, David

251

The E3 ubiquitin ligase mind bomb 1 ubiquitinates and promotes the degradation of survival of motor neuron protein  

PubMed Central

Spinal muscular atrophy is an inherited motor neuron disease that results from a deficiency of the survival of motor neuron (SMN) protein. SMN is ubiquitinated and degraded through the ubiquitin proteasome system (UPS). We have previously shown that proteasome inhibition increases SMN protein levels, improves motor function, and reduces spinal cord, muscle, and neuromuscular junction pathology of spinal muscular atrophy (SMA) mice. Specific targets in the UPS may be more efficacious and less toxic. In this study, we show that the E3 ubiquitin ligase, mind bomb 1 (Mib1), interacts with and ubiquitinates SMN and facilitates its degradation. Knocking down Mib1 levels increases SMN protein levels in cultured cells. Also, knocking down the Mib1 orthologue improves neuromuscular function in Caenorhabditis elegans deficient in SMN. These findings demonstrate that Mib1 ubiquitinates and catalyzes the degradation of SMN, and thus represents a novel therapeutic target for SMA. PMID:23615451

Kwon, Deborah Y.; Dimitriadi, Maria; Terzic, Barbara; Cable, Casey; Hart, Anne C.; Chitnis, Ajay; Fischbeck, Kenneth H.; Burnett, Barrington G.

2013-01-01

252

Modelling Pathways to Rubisco Degradation: A Structural Equation Network Modelling Approach  

PubMed Central

‘Omics analysis (transcriptomics, proteomics) quantifies changes in gene/protein expression, providing a snapshot of changes in biochemical pathways over time. Although tools such as modelling that are needed to investigate the relationships between genes/proteins already exist, they are rarely utilised. We consider the potential for using Structural Equation Modelling to investigate protein-protein interactions in a proposed Rubisco protein degradation pathway using previously published data from 2D electrophoresis and mass spectrometry proteome analysis. These informed the development of a prior model that hypothesised a pathway of Rubisco Large Subunit and Small Subunit degradation, producing both primary and secondary degradation products. While some of the putative pathways were confirmed by the modelling approach, the model also demonstrated features that had not been originally hypothesised. We used Bayesian analysis based on Markov Chain Monte Carlo simulation to generate output statistics suggesting that the model had replicated the variation in the observed data due to protein-protein interactions. This study represents an early step in the development of approaches that seek to enable the full utilisation of information regarding the dynamics of biochemical pathways contained within proteomics data. As these approaches gain attention, they will guide the design and conduct of experiments that enable ‘Omics modelling to become a common place practice within molecular biology. PMID:24498339

Tétard-Jones, Catherine; Gatehouse, Angharad M. R.; Cooper, Julia; Leifert, Carlo; Rushton, Steven

2014-01-01

253

New Hydrocarbon Degradation Pathways in the Microbial Metagenome from Brazilian Petroleum Reservoirs  

PubMed Central

Current knowledge of the microbial diversity and metabolic pathways involved in hydrocarbon degradation in petroleum reservoirs is still limited, mostly due to the difficulty in recovering the complex community from such an extreme environment. Metagenomics is a valuable tool to investigate the genetic and functional diversity of previously uncultured microorganisms in natural environments. Using a function-driven metagenomic approach, we investigated the metabolic abilities of microbial communities in oil reservoirs. Here, we describe novel functional metabolic pathways involved in the biodegradation of aromatic compounds in a metagenomic library obtained from an oil reservoir. Although many of the deduced proteins shared homology with known enzymes of different well-described aerobic and anaerobic catabolic pathways, the metagenomic fragments did not contain the complete clusters known to be involved in hydrocarbon degradation. Instead, the metagenomic fragments comprised genes belonging to different pathways, showing novel gene arrangements. These results reinforce the potential of the metagenomic approach for the identification and elucidation of new genes and pathways in poorly studied environments and contribute to a broader perspective on the hydrocarbon degradation processes in petroleum reservoirs. PMID:24587220

Sierra-García, Isabel Natalia; Correa Alvarez, Javier; Pantaroto de Vasconcellos, Suzan; Pereira de Souza, Anete; dos Santos Neto, Eugenio Vaz; de Oliveira, Valéria Maia

2014-01-01

254

New hydrocarbon degradation pathways in the microbial metagenome from Brazilian petroleum reservoirs.  

PubMed

Current knowledge of the microbial diversity and metabolic pathways involved in hydrocarbon degradation in petroleum reservoirs is still limited, mostly due to the difficulty in recovering the complex community from such an extreme environment. Metagenomics is a valuable tool to investigate the genetic and functional diversity of previously uncultured microorganisms in natural environments. Using a function-driven metagenomic approach, we investigated the metabolic abilities of microbial communities in oil reservoirs. Here, we describe novel functional metabolic pathways involved in the biodegradation of aromatic compounds in a metagenomic library obtained from an oil reservoir. Although many of the deduced proteins shared homology with known enzymes of different well-described aerobic and anaerobic catabolic pathways, the metagenomic fragments did not contain the complete clusters known to be involved in hydrocarbon degradation. Instead, the metagenomic fragments comprised genes belonging to different pathways, showing novel gene arrangements. These results reinforce the potential of the metagenomic approach for the identification and elucidation of new genes and pathways in poorly studied environments and contribute to a broader perspective on the hydrocarbon degradation processes in petroleum reservoirs. PMID:24587220

Sierra-García, Isabel Natalia; Correa Alvarez, Javier; de Vasconcellos, Suzan Pantaroto; Pereira de Souza, Anete; dos Santos Neto, Eugenio Vaz; de Oliveira, Valéria Maia

2014-01-01

255

Mechanochemical degradation of tetrabromobisphenol A: performance, products and pathway.  

PubMed

Tetrabromobisphenol A (TBBPA) is the most widely used brominated flame retardant (BFR), which has received more and more concerns due to its high lipophilicity, persistency and endocrine disrupting property in the environment. Considering the possible need for the safe disposal of TBBPA containing wastes in the future, the potential of mechanochemical (MC) destruction as a promising non-combustion technology was investigated in this study. TBBPA was co-ground with calcium oxide (CaO) or the mixture of iron powder and quartz sand (Fe+SiO(2)) in a planetary ball mill at room temperature. The method of Fe+SiO(2) destructed over 98% of initial TBBPA after 3h and acquired 95% debromination rate after 5h, which showed a better performance than the CaO method. Raman spectra and Fourier transform infrared spectroscopy (FTIR) demonstrated the generation of inorganic carbon with the disappearance of benzene ring and CBr bond, indicating the carbonization and debromination process during mechanochemical reaction. LC-MS-MS screening showed that the intermediates of the treatment with Fe+SiO(2) were tri-, bi-, mono-brominated BPA, BPA and other fragments. Finally all the intermediates were also destroyed after 5h grinding. The bromine balance was calculated and a possible reaction pathway was proposed. PMID:23158692

Zhang, Kunlun; Huang, Jun; Zhang, Wang; Yu, Yunfei; Deng, Shubo; Yu, Gang

2012-12-01

256

Cellular mutants define a common mRNA degradation pathway targeting cytokine AU-rich elements.  

PubMed Central

To functionally classify AU-rich elements (AREs) from six different cytokine mRNAs, we made use of two previously described HT1080-derived cellular mutants (slowA, slowC) that lack a function required for the rapid degradation of interleukin-3 (IL-3) mRNA. Here we show that the defect is specific for ARE-containing mRNAs, whereas nonsense-mediated decay is intact. Degradation of beta-globin reporter transcripts mediated by the AREs of IL-3, GM-CSF, and TNFalpha, as well as by the structurally different and less potent AREs of IL-2 and IL-6, is impaired in both mutants. All these reporter transcripts are also sensitive to decay induced by ectopic expression of the RNA-binding protein tristetraprolin in the slowC background. Thus, we concluded that the mutants slowA and slowC define a common mRNA degradation pathway that targets cytokine AREs. In NIH3T3 cells, this decay pathway becomes incapacitated by upstream signaling from p38 MAP- or PI3-kinases, which independently stabilize cytokine ARE-containing transcripts. In contrast, c-fos ARE-directed mRNA degradation proceeds through a different pathway not affected by these kinases. PMID:11720287

Stoecklin, G; Stoeckle, P; Lu, M; Muehlemann, O; Moroni, C

2001-01-01

257

Modulation of apoptosis sensitivity through the interplay with autophagic and proteasomal degradation pathways  

PubMed Central

Autophagic and proteasomal degradation constitute the major cellular proteolysis pathways. Their physiological and pathophysiological adaptation and perturbation modulates the relative abundance of apoptosis-transducing proteins and thereby can positively or negatively adjust cell death susceptibility. In addition to balancing protein expression amounts, components of the autophagic and proteasomal degradation machineries directly interact with and co-regulate apoptosis signal transduction. The influence of autophagic and proteasomal activity on apoptosis susceptibility is now rapidly gaining more attention as a significant modulator of cell death signalling in the context of human health and disease. Here we present a concise and critical overview of the latest knowledge on the molecular interplay between apoptosis signalling, autophagy and proteasomal protein degradation. We highlight that these three pathways constitute an intricate signalling triangle that can govern and modulate cell fate decisions between death and survival. Owing to rapid research progress in recent years, it is now possible to provide detailed insight into the mechanisms of pathway crosstalk, common signalling nodes and the role of multi-functional proteins in co-regulating both protein degradation and cell death. PMID:24457955

Delgado, M E; Dyck, L; Laussmann, M A; Rehm, M

2014-01-01

258

Genomic and metabolic analysis of fluoranthene degradation pathway in Celeribacter indicus P73T  

PubMed Central

Celeribacter indicus P73T, isolated from deep-sea sediment from the Indian Ocean, is capable of degrading a wide range of polycyclic aromatic hydrocarbons (PAHs) and is the first fluoranthene-degrading bacterium within the family Rhodobacteraceae. Here, the complete genome sequence of strain P73T is presented and analyzed. Besides a 4.5-Mb circular chromosome, strain P73T carries five plasmids, and encodes 4827 predicted protein-coding sequences. One hundred and thirty-eight genes, including 14 dioxygenase genes, were predicted to be involved in the degradation of aromatic compounds, and most of these genes are clustered in four regions. P73_0346 is the first fluoranthene 7,8-dioxygenase to be discovered and the first fluoranthene dioxygenase within the toluene/biphenyl family. The degradative genes in regions B and D in P73T are absent in Celeribacter baekdonensis B30, which cannot degrade PAHs. Four intermediate metabolites [acenaphthylene-1(2H)-one, acenaphthenequinone, 1,2-dihydroxyacenaphthylene, and 1,8-naphthalic anhydride] of fluoranthene degradation by strain P73T were detected as the main intermediates, indicating that the degradation of fluoranthene in P73T was initiated by dioxygenation at the C-7,8 positions. Based on the genomic and metabolitic results, we propose a C-7,8 dioxygenation pathway in which fluoranthene is mineralized to TCA cycle intermediates. PMID:25582347

Cao, Junwei; Lai, Qiliang; Yuan, Jun; Shao, Zongze

2015-01-01

259

Genomic and metabolic analysis of fluoranthene degradation pathway in Celeribacter indicus P73(T.).  

PubMed

Celeribacter indicus P73(T), isolated from deep-sea sediment from the Indian Ocean, is capable of degrading a wide range of polycyclic aromatic hydrocarbons (PAHs) and is the first fluoranthene-degrading bacterium within the family Rhodobacteraceae. Here, the complete genome sequence of strain P73(T) is presented and analyzed. Besides a 4.5-Mb circular chromosome, strain P73(T) carries five plasmids, and encodes 4827 predicted protein-coding sequences. One hundred and thirty-eight genes, including 14 dioxygenase genes, were predicted to be involved in the degradation of aromatic compounds, and most of these genes are clustered in four regions. P73_0346 is the first fluoranthene 7,8-dioxygenase to be discovered and the first fluoranthene dioxygenase within the toluene/biphenyl family. The degradative genes in regions B and D in P73(T) are absent in Celeribacter baekdonensis B30, which cannot degrade PAHs. Four intermediate metabolites [acenaphthylene-1(2H)-one, acenaphthenequinone, 1,2-dihydroxyacenaphthylene, and 1,8-naphthalic anhydride] of fluoranthene degradation by strain P73(T) were detected as the main intermediates, indicating that the degradation of fluoranthene in P73(T) was initiated by dioxygenation at the C-7,8 positions. Based on the genomic and metabolitic results, we propose a C-7,8 dioxygenation pathway in which fluoranthene is mineralized to TCA cycle intermediates. PMID:25582347

Cao, Junwei; Lai, Qiliang; Yuan, Jun; Shao, Zongze

2015-01-01

260

Combination of degradation pathways for naphthalene utilization in Rhodococcus sp. strain TFB  

PubMed Central

Rhodococcus sp. strain TFB is a metabolic versatile bacterium able to grow on naphthalene as the only carbon and energy source. Applying proteomic, genetic and biochemical approaches, we propose in this paper that, at least, three coordinated but independently regulated set of genes are combined to degrade naphthalene in TFB. First, proteins involved in tetralin degradation are also induced by naphthalene and may carry out its conversion to salicylaldehyde. This is the only part of the naphthalene degradation pathway showing glucose catabolite repression. Second, a salicylaldehyde dehydrogenase activity that converts salicylaldehyde to salicylate is detected in naphthalene-grown cells but not in tetralin-or salicylate-grown cells. Finally, we describe the chromosomally located nag genes, encoding the gentisate pathway for salicylate conversion into fumarate and pyruvate, which are only induced by salicylate and not by naphthalene. This work shows how biodegradation pathways in Rhodococcus sp. strain TFB could be assembled using elements from different pathways mainly because of the laxity of the regulatory systems and the broad specificity of the catabolic enzymes. PMID:24325207

Tomás-Gallardo, Laura; Gómez-Álvarez, Helena; Santero, Eduardo; Floriano, Belén

2014-01-01

261

Metagenomic Identification of Bacterioplankton Taxa and Pathways Involved in Microcystin Degradation in Lake Erie  

PubMed Central

Cyanobacterial harmful blooms (CyanoHABs) that produce microcystins are appearing in an increasing number of freshwater ecosystems worldwide, damaging quality of water for use by human and aquatic life. Heterotrophic bacteria assemblages are thought to be important in transforming and detoxifying microcystins in natural environments. However, little is known about their taxonomic composition or pathways involved in the process. To address this knowledge gap, we compared the metagenomes of Lake Erie free-living bacterioplankton assemblages in laboratory microcosms amended with microcystins relative to unamended controls. A diverse array of bacterial phyla were responsive to elevated supply of microcystins, including Acidobacteria, Actinobacteria, Bacteroidetes, Planctomycetes, Proteobacteria of the alpha, beta, gamma, delta and epsilon subdivisions and Verrucomicrobia. At more detailed taxonomic levels, Methylophilales (mainly in genus Methylotenera) and Burkholderiales (mainly in genera Bordetella, Burkholderia, Cupriavidus, Polaromonas, Ralstonia, Polynucleobacter and Variovorax) of Betaproteobacteria were suggested to be more important in microcystin degradation than Sphingomonadales of Alphaproteobacteria. The latter taxa were previously thought to be major microcystin degraders. Homologs to known microcystin-degrading genes (mlr) were not overrepresented in microcystin-amended metagenomes, indicating that Lake Erie bacterioplankton might employ alternative genes and/or pathways in microcystin degradation. Genes for xenobiotic metabolism were overrepresented in microcystin-amended microcosms, suggesting they are important in bacterial degradation of microcystin, a phenomenon that has been identified previously only in eukaryotic systems. PMID:23637924

Mou, Xiaozhen; Lu, Xinxin; Jacob, Jisha; Sun, Shulei; Heath, Robert

2013-01-01

262

Effects of chlorobenzoate transformation on the Pseudomonas testosteroni biphenyl and chlorobiphenyl degradation pathway.  

PubMed Central

Bacterial conversion of biphenyl (BP) and chlorobiphenyls (CBPs) to benzoates and chlorobenzoates (CBAs) proceeds by introduction of molecular oxygen at the 2,3 position, followed by a 1,2-meta cleavage of the molecule. Complete mineralization of CBPs requires the presence of two sets of genes, one for the transformation fo CBPs into CBAs and a second for the degradation of CBAs. It has been shown previously that removal of the CBAs produced from the degradation of CBPs is essential for efficient degradation of CBPs. In this study we confirmed that CBAs inhibit BP and CBP transformation in Pseudomonas testosteroni B-356. Among the three monochlorobenzoates tested, 3-chlorobenzoate was the most effective inhibitor. Furthermore, we found that in strain B-356, CBA transformation is controlled by BP-induced oxygenases that are not present in benzoate-grown cells. We found that this BP-linked CBA transformation pathway transformed CBAs produced from CBPs into several metabolites, including chlorocatechols and corresponding muconic semialdehydes. These metabolites inhibited the 2,3-dihydroxybiphenyl 1,2-dioxygenase, while CBAs by themselves had no effect on this enzyme. Therefore, on the basis of this and other observations, it appears that when CBAs produced from CBPs accumulate in the growth medium, they are converted into unproductive metabolites that reduce the flux of the BP and CBP degradation pathway. The practical implications of these interactions on the microbial degradation of polychlorinated BPs are also discussed. PMID:1610172

Sondossi, M; Sylvestre, M; Ahmad, D

1992-01-01

263

Pathways for 3-chloro- and 4-chlorobenzoate degradation in Pseudomonas aeruginosa 3mT.  

PubMed

A bacterial isolate, Pseudomonas aeruginosa 3mT exhibited the ability to degrade high concentrations of 3-chlorobenzoate (3-CBA, 8 g l(-1)) and 4-chlorobenzoate (4-CBA 12 g l(-1)) (Ajithkumar 1998). In this study, by delineating the initial biochemical steps involved in the degradation of these compounds, we investigated how this strain can do so well. Resting cells, permeabilised cells as well as cell-free extracts failed to dechlorinate both 3-CBA and 4-CBA under anaerobic conditions, whereas the former two readily degraded both compounds under aerobic conditions. Accumulation of any intermediary metabolite was not observed during growth as well as reaction with resting cells under highly aerated conditions. However, on modification of reaction conditions, 3-chlorocatechol (3-CC) and 4-chlorocatechol (4-CC) accumulated in 3-CBA and 4-CBA flasks, respectively. Fairly high titres of pyrocatechase II (chlorocatechol 1,2-dioxygenase) activity were obtained in extracts of cells grown on 3-CBA and 4-CBA. Meta-pyrocatechase (catechol 2,3-dioxygenase) activity against 4-CC and catechol, but not against 3-CC, was also detected in low titres. Accumulation of small amounts of 2-chloro-5-hydroxy muconic semialdehyde, the meta-cleavage product of 4-CC, was detected in the medium, when 4-CBA concentration was 4 mM or greater, indicating the presence of a minor meta-pathway in strain 3mT. However, 3-CBA exclusively, and more than 99% of 4-CBA were degraded through the formation of the respective chlorocatechol, via a modified ortho-pathway. This defies the traditional view that the microbes that follow chlorocatechol pathways are not very good degraders of chlorobenzoates. 4-Hydroxybenzoate was readily (and 3-hydroxybenzoate to a lesser extent) degraded by the strain, through the formation of protocatechuate and gentisate, respectively, as intermediary dihydroxy metabolites. PMID:11432583

Ajithkumar, P V; Kunhi, A A

2000-01-01

264

NOVEL ORGANIZATION OF CATECHOL META PATHWAY GENES IN THE NITROBENZENE DEGRADER COMAMONAS SP. JS765 AND ITS EVOLUTIONARY IMPLICATION  

Technology Transfer Automated Retrieval System (TEKTRAN)

The catechol meta cleavage pathway is one of the central metabolic pathways for the degradation of aromatic compounds. A novel organization of the pathway genes, different from that of classical soil microorganisms, has been observed in Sphingomonas sp HV3 and Pseudomonas sp. DJ77. In a Comamonas ...

265

Microbial degradation of chloroaromatics: use of the meta-cleavage pathway for mineralization of chlorobenzene.  

PubMed Central

Pseudomonas putida GJ31 is able to simultaneously grow on toluene and chlorobenzene. When cultures of this strain were inhibited with 3-fluorocatechol while growing on toluene or chlorobenzene, 3-methylcatechol or 3-chlorocatechol, respectively, accumulated in the medium. To establish the catabolic routes for these catechols, activities of enzymes of the (modified) ortho- and meta-cleavage pathways were measured in crude extracts of cells of P. putida GJ31 grown on various aromatic substrates, including chlorobenzene. The enzymes of the modified ortho-cleavage pathway were never present, while the enzymes of the meta-cleavage pathway were detected in all cultures. This indicated that chloroaromatics and methylaromatics are both converted via the meta-cleavage pathway. Meta cleavage of 3-chlorocatechol usually leads to the formation of a reactive acylchloride, which inactivates the catechol 2,3-dioxygenase and blocks further degradation of catechols. However, partially purified catechol 2,3-dioxygenase of P. putida GJ31 converted 3-chlorocatechol to 2-hydroxy-cis,cis-muconic acid. Apparently, P. putida GJ31 has a meta-cleavage enzyme which is resistant to inactivation by the acylchloride, providing this strain with the exceptional ability to degrade both toluene and chlorobenzene via the meta-cleavage pathway. PMID:9226262

Mars, A E; Kasberg, T; Kaschabek, S R; van Agteren, M H; Janssen, D B; Reineke, W

1997-01-01

266

Microbial degradation of chloroaromatics: use of the meta-cleavage pathway for mineralization of chlorobenzene.  

PubMed

Pseudomonas putida GJ31 is able to simultaneously grow on toluene and chlorobenzene. When cultures of this strain were inhibited with 3-fluorocatechol while growing on toluene or chlorobenzene, 3-methylcatechol or 3-chlorocatechol, respectively, accumulated in the medium. To establish the catabolic routes for these catechols, activities of enzymes of the (modified) ortho- and meta-cleavage pathways were measured in crude extracts of cells of P. putida GJ31 grown on various aromatic substrates, including chlorobenzene. The enzymes of the modified ortho-cleavage pathway were never present, while the enzymes of the meta-cleavage pathway were detected in all cultures. This indicated that chloroaromatics and methylaromatics are both converted via the meta-cleavage pathway. Meta cleavage of 3-chlorocatechol usually leads to the formation of a reactive acylchloride, which inactivates the catechol 2,3-dioxygenase and blocks further degradation of catechols. However, partially purified catechol 2,3-dioxygenase of P. putida GJ31 converted 3-chlorocatechol to 2-hydroxy-cis,cis-muconic acid. Apparently, P. putida GJ31 has a meta-cleavage enzyme which is resistant to inactivation by the acylchloride, providing this strain with the exceptional ability to degrade both toluene and chlorobenzene via the meta-cleavage pathway. PMID:9226262

Mars, A E; Kasberg, T; Kaschabek, S R; van Agteren, M H; Janssen, D B; Reineke, W

1997-07-01

267

Degradation of 2,3-Dihydroxybenzoate by a Novel meta-Cleavage Pathway  

PubMed Central

2,3-Dihydroxybenzoate is the precursor in the biosynthesis of several siderophores and an important plant secondary metabolite that, in bacteria, can be degraded via meta-cleavage of the aromatic ring. The dhb cluster of Pseudomonas reinekei MT1 encodes a chimeric meta-cleavage pathway involved in the catabolism of 2,3-dihydroxybenzoate. While the first two enzymes, DhbA and DhbB, are phylogenetically related to those involved in 2,3-dihydroxy-p-cumate degradation, the subsequent steps are catalyzed by enzymes related to those involved in catechol degradation (DhbCDEFGH). Characterization of kinetic properties of DhbA extradiol dioxygenase identified 2,3-dihydroxybenzoate as the preferred substrate. Deletion of the encoding gene impedes growth of P. reinekei MT1 on 2,3-dihydroxybenzoate. DhbA catalyzes 3,4-dioxygenation with 2-hydroxy-3-carboxymuconate as the product, which is then decarboxylated by DhbB to 2-hydroxymuconic semialdehyde. This compound is then subject to dehydrogenation and further degraded to citrate cycle intermediates. Transcriptional analysis revealed genes of the dhB gene cluster to be highly expressed during growth with 2,3-dihydroxybenzoate, whereas a downstream-localized gene encoding 2-hydroxymuconic semialdehyde hydrolase, dispensable for 2,3-dihydroxybenzoate metabolism but crucial for 2,3-dihydroxy-p-cumate degradation, was only marginally expressed. This is the first report describing a gene cluster encoding enzymes for the degradation of 2,3-dihydroxybenzoate. PMID:22609919

Marín, Macarena; Plumeier, Iris

2012-01-01

268

Effects of feeding hexane-extracts of a shochu distillery by-product on skeletal muscle protein degradation in broiler chicken.  

PubMed

We have found that shochu distillery by-product (SDBP) contains a growth promoting factor that can be extracted with ether. In the present study, we administered hexane-extracts of SDBP (HSDBP) to broiler chickens and observed changes in skeletal muscle protein degradation in order to clarify the mechanism of growth promotion due to SDBP feeding. The pectoralis superficial muscle weight was significantly increased by HSDBP feeding. Plasma N(tau)-methylhistidine concentration was significantly decreased by HSDBP, showing that the rate of muscle protein degradation decreased. It was also found that the expression of mRNA of ubiquitin-proteasome system and calpain was decreased by HSDBP. These results indicate that growth promotion due to SDBP is caused by suppression of skeletal muscle protein degradation, which is related to the ubiquitin-ptoteasome system and calpain. PMID:20057147

Kamizono, Tomomi; Nakashima, Kazuki; Ohtsuka, Akira; Hayashi, Kunioki

2010-01-01

269

Organization of metabolic pathways and molecular-genetic mechanisms of xenobiotic degradation in microorganisms: A review  

Microsoft Academic Search

Contemporary data on the mechanism of biodegradation of aromatic hydrocarbons and biodegradation genes (genomic organization\\u000a and pathways of evolution) in diverse groups of microorganisms have been reviewed. Studies of this problem are topical, in\\u000a view of the need in identification and construction of new strains degrading xenobiotics, particularly those halogenated.\\u000a For this reason, emphasis is placed on specific features of

V. G. Khomenkov; A. B. Shevelev; V. G. Zhukov; N. A. Zagustina; A. M. Bezborodov; V. O. Popov

2008-01-01

270

Genetic Evidence for a Defective Xylan Degradation Pathway in Lactococcus lactis  

Microsoft Academic Search

Genetic and biochemical evidence for a defective xylan degradation pathway was found linked to the xylose operon in three lactococcal strains, Lactococcus lactis 210, L. lactis IO-1, and L. lactis NRRL B-4449. Immedi- ately downstream of the xylulose kinase gene (xylB) (K. A. Erlandson, J.-H. Park, W. El Khal, H.-H. Kao, P. Basaran, S. Brydges, and C. A. Batt, Appl.

KARN A. ERLANDSON; SOAZIG C. DELAMARRE; CARL A. BATT

2001-01-01

271

Phenylacetate Catabolism in Rhodococcus sp. Strain RHA1: a Central Pathway for Degradation of Aromatic Compounds  

PubMed Central

In gram-negative bacteria, a pathway for aerobic degradation of phenylacetic acid (PAA) that proceeds via phenylacetyl-coenzyme A (CoA) and hydrolytic ring fission plays a central role in the degradation of a range of aromatic compounds. In contrast, the PAA pathway and its role are not well characterized in gram-positive bacteria. A cluster including 13 paa genes encoding enzymes orthologous to those of gram-negative bacteria was identified on the chromosome of Rhodococcus sp. strain RHA1. These genes were transcribed during growth on PAA, with 11 of the genes apparently in an operon yielding a single transcript. Quantitative proteomic analyses revealed that at least 146 proteins were more than twice as abundant in PAA-grown cells of RHA1 than in pyruvate-grown cells. Of these proteins, 29 were identified, including 8 encoded by the paa genes. Knockout mutagenesis indicated that paaN, encoding a putative ring-opening enzyme, was essential for growth on PAA. However, paaF, encoding phenylacetyl-CoA ligase, and paaR, encoding a putative regulator, were not essential. paaN was also essential for growth of RHA1 on phenylacetaldehyde, phenylpyruvate, 4-phenylbutyrate, 2-phenylethanol, 2-phenylethylamine, and l-phenylalanine. In contrast, growth on 3-hydroxyphenylacetate, ethylbenzene, and styrene was unaffected. These results suggest that the range of substrates degraded via the PAA pathway in RHA1 is somewhat limited relative to the range in previously characterized gram-negative bacteria. PMID:15968060

Navarro-Llorens, Juana María; Patrauchan, Marianna A.; Stewart, Gordon R.; Davies, Julian E.; Eltis, Lindsay D.; Mohn, William W.

2005-01-01

272

Investigation of hexabromocyclododecane thermal degradation pathways by gas chromatography/mass spectrometry.  

PubMed

The decomposition products of hexabromocyclododecane (HBCD), a widely used brominated flame retardant, were investigated by gas chromatography/mass spectrometry (GC/MS). HBCD thermal degradation was conducted under a moderate heating rate (10 degrees C/min) in a batch reactor using both inert and oxidizing atmospheres. GC/MS analysis allowed the identification of substances derived from the primary pyrolysis process at the moderate heating rates used. The presence of oxygen seems to have a negligible influence on the degradation products obtained in HBCD decomposition, at least at moderate heating rates. Based on the identified products, the main pathways of HBCD thermal degradation were assessed and a mechanism for HBCD decomposition was proposed. The results obtained indicate that hexa-, penta- and tetrabrominated polyaromatic structures seem not to be primary products of HBCD decomposition, and may only be obtained by secondary bromination reactions. PMID:11319791

Barontini, F; Cozzani, V; Cuzzola, A; Petarca, L

2001-01-01

273

cis-Inhibition of proteasomal degradation by viral repeats: impact of length and amino acid composition.  

PubMed

The Gly-Ala repeat (GAr) of the Epstein-Barr virus nuclear antigen 1 is a cis acting inhibitor of ubiquitin-proteasome proteolysis. We have investigated the capacity of various repeats to inhibit the turnover of the proteasomal substrate IkappaBalpha. Inhibition of TNFalpha-induced degradation was achieved by insertion of octamers containing three alanines or valines, interspersed by no more then three consecutive glycines. The inhibitory activity was abolished by increasing the length of the spacer, by eliminating the spacers, or by substitution of a single hydrophobic residue with a polar or charged residue. A serine containing octamer was inactive but inhibition was partially restored by insertion of three consecutive repeats. These findings suggest a model where inhibition requires the interaction of at least three alanine residues of the GAr in a beta-strand conformation with adjacent hydrophobic binding pockets of a putative receptor. PMID:11418128

Sharipo, A; Imreh, M; Leonchiks, A; Brändén, C; Masucci, M G

2001-06-15

274

Expression of expanded polyglutamine targets profilin for degradation and alters actin dynamics  

PubMed Central

Huntington’s disease is caused by polyglutamine expansion in the huntingtin protein. Huntingtin directly interacts with profilin, a major actin monomer sequestering protein and a key integrator of signals leading to actin polymerization. We observed a progressive loss of profilin in the cerebral cortex of Huntington’s disease patients, and in cell culture and Drosophila models of polyglutamine disease. This loss of profilin is likely due to increased degradation through the ubiquitin-proteasome system. Profilin loss reduces the F/G actin ratio, indicating a shift in actin polymerization. Overexpression of profilin abolishes mutant huntingtin toxicity in cells and partially ameliorates the morphological and functional eye phenotype and extends lifespan in a transgenic polyglutamine Drosophila model. These results indicate a link between huntingtin and profilin and implicate profilin in Huntington’s disease pathogenesis. PMID:18417352

Burnett, Barrington G.; Andrews, Jaime; Ranganathan, Srikanth; Fischbeck, Kenneth H.; Di Prospero, Nicholas A.

2008-01-01

275

Expression of expanded polyglutamine targets profilin for degradation and alters actin dynamics.  

PubMed

Huntington's disease is caused by polyglutamine expansion in the huntingtin protein. Huntingtin directly interacts with profilin, a major actin monomer sequestering protein and a key integrator of signals leading to actin polymerization. We observed a progressive loss of profilin in the cerebral cortex of Huntington's disease patients, and in cell culture and Drosophila models of polyglutamine disease. This loss of profilin is likely due to increased degradation through the ubiquitin proteasome system. Profilin loss reduces the F/G actin ratio, indicating a shift in actin polymerization. Overexpression of profilin abolishes mutant huntingtin toxicity in cells and partially ameliorates the morphological and functional eye phenotype and extends lifespan in a transgenic polyglutamine Drosophila model. These results indicate a link between huntingtin and profilin and implicate profilin in Huntington's disease pathogenesis. PMID:18417352

Burnett, Barrington G; Andrews, Jaime; Ranganathan, Srikanth; Fischbeck, Kenneth H; Di Prospero, Nicholas A

2008-06-01

276

Production of Pyomelanin, a Second Type of Melanin, via the Tyrosine Degradation Pathway in Aspergillus fumigatus?  

PubMed Central

Aspergillus fumigatus is the most important airborne fungal pathogen of immunosuppressed humans. A. fumigatus is able to produce dihydroxynaphthalene melanin, which is predominantly present in the conidia. Its biosynthesis is an important virulence determinant. Here, we show that A. fumigatus is able to produce an alternative melanin, i.e., pyomelanin, by a different pathway, starting from l-tyrosine. Proteome analysis indicated that the l-tyrosine degradation enzymes are synthesized when the fungus is grown with l-tyrosine in the medium. To investigate the pathway in detail, we deleted the genes encoding essential enzymes for pigment production, homogentisate dioxygenase (hmgA) and 4-hydroxyphenylpyruvate dioxygenase (hppD). Comparative Fourier transform infrared spectroscopy of synthetic pyomelanin and pigment extracted from A. fumigatus cultures confirmed the identity of the observed pigment as pyomelanin. In the hmgA deletion strain, HmgA activity was abolished and the accumulation of homogentisic acid provoked an increased pigment formation. In contrast, homogentisic acid and pyomelanin were not observed with an hppD deletion mutant. Germlings of the hppD deletion mutant showed an increased sensitivity to reactive oxygen intermediates. The transcription of both studied genes was induced by l-tyrosine. These results confirmed the function of the deleted genes and the predicted pathway in A. fumigatus. Homogentisic acid is the major intermediate, and the l-tyrosine degradation pathway leading to pyomelanin is similar to that in humans leading to alkaptomelanin. PMID:19028908

Schmaler-Ripcke, Jeannette; Sugareva, Venelina; Gebhardt, Peter; Winkler, Robert; Kniemeyer, Olaf; Heinekamp, Thorsten; Brakhage, Axel A.

2009-01-01

277

Rhodococcus erythropolis DCL14 Contains a Novel Degradation Pathway for Limonene  

PubMed Central

Strain DCL14, which is able to grow on limonene as a sole source of carbon and energy, was isolated from a freshwater sediment sample. This organism was identified as a strain of Rhodococcus erythropolis by chemotaxonomic and genetic studies. R. erythropolis DCL14 also assimilated the terpenes limonene-1,2-epoxide, limonene-1,2-diol, carveol, carvone, and (?)-menthol, while perillyl alcohol was not utilized as a carbon and energy source. Induction tests with cells grown on limonene revealed that the oxygen consumption rates with limonene-1,2-epoxide, limonene-1,2-diol, 1-hydroxy-2-oxolimonene, and carveol were high. Limonene-induced cells of R. erythropolis DCL14 contained the following four novel enzymatic activities involved in the limonene degradation pathway of this microorganism: a flavin adenine dinucleotide- and NADH-dependent limonene 1,2-monooxygenase activity, a cofactor-independent limonene-1,2-epoxide hydrolase activity, a dichlorophenolindophenol-dependent limonene-1,2-diol dehydrogenase activity, and an NADPH-dependent 1-hydroxy-2-oxolimonene 1,2-monooxygenase activity. Product accumulation studies showed that (1S,2S,4R)-limonene-1,2-diol, (1S,4R)-1-hydroxy-2-oxolimonene, and (3R)-3-isopropenyl-6-oxoheptanoate were intermediates in the (4R)-limonene degradation pathway. The opposite enantiomers [(1R,2R,4S)-limonene-1,2-diol, (1R,4S)-1-hydroxy-2-oxolimonene, and (3S)-3-isopropenyl-6-oxoheptanoate] were found in the (4S)-limonene degradation pathway, while accumulation of (1R,2S,4S)-limonene-1,2-diol from (4S)-limonene was also observed. These results show that R. erythropolis DCL14 metabolizes both enantiomers of limonene via a novel degradation pathway that starts with epoxidation at the 1,2 double bond forming limonene-1,2-epoxide. This epoxide is subsequently converted to limonene-1,2-diol, 1-hydroxy-2-oxolimonene, and 7-hydroxy-4-isopropenyl-7-methyl-2-oxo-oxepanone. This lactone spontaneously rearranges to form 3-isopropenyl-6-oxoheptanoate. In the presence of coenzyme A and ATP this acid is converted further, and this finding, together with the high levels of isocitrate lyase activity in extracts of limonene-grown cells, suggests that further degradation takes place via the ?-oxidation pathway. PMID:10224006

van der Werf, Mariët J.; Swarts, Henk J.; de Bont, Jan A. M.

1999-01-01

278

Enhanced degradation in soil of the herbicide EPTC and determination of its degradative pathway by an isolated soil microorganism  

SciTech Connect

A series of experiments was conducted to examine the ability of Ohio soils to develop enhanced degradation of the herbicide EPTC (s-ethyl N,N-dipropyl carbamothiaote) and to determine its metabolism by an isolated soil microorganism. Three soils selected to obtain an range in pH, texture, and organic carbon were treated with EPTC for 4 consecutive applications (6 weeks between applications). EPTC concentrations as measured by gas chromatography, decreased 80% or more one week after the second application in all three soils. Metabolism of unlabelled and labelled EPTC by an isolated soil microbe was followed by GC/MS and TLC/LSC analysis, respectively. Rapid decrease in 14-C activity in the organic fraction corresponded with rapid {sup 14}CO{sub 2} evolution and transient increase in 14-C activity in the aqueous fraction. Four metabolites were observed in the TLC analysis. Two were identified as EPTC-sulfoxide and N-depropyl EPTC with N-depropyl EPTC being confirmed by GC/MS analysis. The availability of different pathways for EPTC metabolism by soil microbes after repeated applications to the soil results in its very rapid degradation and loss of efficacy.

Ankumah, R.O.

1988-01-01

279

Amyloid-Beta Protein Clearance and Degradation (ABCD) Pathways and their Role in Alzheimer's Disease.  

PubMed

Amyloid-? proteins (A?) of 42 (A?42) and 40 aa (A?40) accumulate as senile plaques (SP) and cerebrovascular amyloid protein deposits that are defining diagnostic features of Alzheimer's disease (AD). A number of rare mutations linked to familial AD (FAD) on the A? precursor protein (APP), Presenilin-1 (PS1), Presenilin- 2 (PS2), Adamalysin10, and other genetic risk factors for sporadic AD such as the ?4 allele of Apolipoprotein E (ApoE-?4) foster the accumulation of A? and also induce the entire spectrum of pathology associated with the disease. A? accumulation is therefore a key pathological event and a prime target for the prevention and treatment of AD. APP is sequentially processed by ?-site APP cleaving enzyme (BACE1) and ?-secretase, a multisubunit PS1/PS2-containing integral membrane protease, to generate A?. Although A? accumulates in all forms of AD, the only pathways known to be affected in FAD increase A? production by APP gene duplication or via base substitutions on APP and ?-secretase subunits PS1 and PS2 that either specifically increase the yield of the longer A?42 or both A?40 and A?42. However, the vast majority of AD patients accumulate A? without these known mutations. This led to proposals that impairment of A? degradation or clearance may play a key role in AD pathogenesis. Several candidate enzymes, including Insulin-degrading enzyme (IDE), Neprilysin (NEP), Endothelin-converting enzyme (ECE), Angiotensin converting enzyme (ACE), Plasmin, and Matrix metalloproteinases (MMPs) have been identified and some have even been successfully evaluated in animal models. Several studies also have demonstrated the capacity of ?-secretase inhibitors to paradoxically increase the yield of A? and we have recently established that the mechanism is by skirting A? degradation. This review outlines major cellular pathways of A? degradation to provide a basis for future efforts to fully characterize the panel of pathways responsible for A? turnover. PMID:25523424

Baranello, Robert J; Bharani, Krishna L; Padmaraju, Vasudevaraju; Chopra, Nipun; Lahiri, Debomoy K; Greig, Nigel H; Pappolla, Miguel A; Sambamurti, Kumar

2015-01-01

280

Anoxic Androgen Degradation by the Denitrifying Bacterium Sterolibacterium denitrificans via the 2,3-seco Pathway  

PubMed Central

The biodegradation of steroids is a crucial biochemical process mediated exclusively by bacteria. So far, information concerning the anoxic catabolic pathways of androgens is largely unknown, which has prevented many environmental investigations. In this work, we show that Sterolibacterium denitrificans DSMZ 13999 can anaerobically mineralize testosterone and some C19 androgens. By using a 13C-metabolomics approach and monitoring the sequential appearance of the intermediates, we demonstrated that S. denitrificans uses the 2,3-seco pathway to degrade testosterone under anoxic conditions. Furthermore, based on the identification of a C17 intermediate, we propose that the A-ring cleavage may be followed by the removal of a C2 side chain at C-5 of 17-hydroxy-1-oxo-2,3-seco-androstan-3-oic acid (the A-ring cleavage product) via retro-aldol reaction. The androgenic activities of the bacterial culture and the identified intermediates were assessed using the lacZ-based yeast androgen assay. The androgenic activity in the testosterone-grown S. denitrificans culture decreased significantly over time, indicating its ability to eliminate androgens. The A-ring cleavage intermediate (?500 ?M) did not exhibit androgenic activity, whereas the sterane-containing intermediates did. So far, only two androgen-degrading anaerobes (Sterolibacterium denitrificans DSMZ 13999 [a betaproteobacterium] and Steroidobacter denitrificans DSMZ 18526 [a gammaproteobacterium]) have been isolated and characterized, and both of them use the 2,3-seco pathway to anaerobically degrade androgens. The key intermediate 2,3-seco-androstan-3-oic acid can be used as a signature intermediate for culture-independent environmental investigations of anaerobic degradation of C19 androgens. PMID:24657867

Wang, Po-Hsiang; Yu, Chang-Ping; Lee, Tzong-Huei; Lin, Ching-Wen; Ismail, Wael; Wey, Shiaw-Pyng; Kuo, An-Ti

2014-01-01

281

Dissection of the Endogenous Cellular Pathways of PCSK9-induced Low Density Lipoprotein Receptor Degradation  

PubMed Central

Elevated levels of plasma low density lipoprotein (LDL)-cholesterol, leading to familial hypercholesterolemia, are enhanced by mutations in at least three major genes, the LDL receptor (LDLR), its ligand apolipoprotein B, and the proprotein convertase PCSK9. Single point mutations in PCSK9 are associated with either hyper- or hypocholesterolemia. Accordingly, PCSK9 is an attractive target for treatment of dyslipidemia. PCSK9 binds the epidermal growth factor domain A (EGF-A) of the LDLR and directs it to endosomes/lysosomes for destruction. Although the mechanism by which PCSK9 regulates LDLR degradation is not fully resolved, it seems to involve both intracellular and extracellular pathways. Here, we show that clathrin light chain small interfering RNAs that block intracellular trafficking from the trans-Golgi network to lysosomes rapidly increased LDLR levels within HepG2 cells in a PCSK9-dependent fashion without affecting the ability of exogenous PCSK9 to enhance LDLR degradation. In contrast, blocking the extracellular LDLR endocytosis/degradation pathway by a 4-, 6-, or 24-h incubation of cells with Dynasore or an EGF-AB peptide or by knockdown of endogenous autosomal recessive hypercholesterolemia did not significantly affect LDLR levels. The present data from HepG2 cells and mouse primary hepatocytes favor a model whereby depending on the dose and/or incubation period, endogenous PCSK9 enhances the degradation of the LDLR both extra- and intracellularly. Therefore, targeting either pathway, or both, would be an effective method to reduce PCSK9 activity in the treatment of hypercholesterolemia and coronary heart disease. PMID:19635789

Poirier, Steve; Mayer, Gaetan; Poupon, Viviane; McPherson, Peter S.; Desjardins, Roxane; Ly, Kevin; Asselin, Marie-Claude; Day, Robert; Duclos, Franck J.; Witmer, Mark; Parker, Rex; Prat, Annik; Seidah, Nabil G.

2009-01-01

282

PINK1-Parkin Pathway Activity Is Regulated by Degradation of PINK1 in the Mitochondrial Matrix  

PubMed Central

Loss-of-function mutations in PINK1, which encodes a mitochondrially targeted serine/threonine kinase, result in an early-onset heritable form of Parkinson's disease. Previous work has shown that PINK1 is constitutively degraded in healthy cells, but selectively accumulates on the surface of depolarized mitochondria, thereby initiating their autophagic degradation. Although PINK1 is known to be a cleavage target of several mitochondrial proteases, whether these proteases account for the constitutive degradation of PINK1 in healthy mitochondria remains unclear. To explore the mechanism by which PINK1 is degraded, we performed a screen for mitochondrial proteases that influence PINK1 abundance in the fruit fly Drosophila melanogaster. We found that genetic perturbations targeting the matrix-localized protease Lon caused dramatic accumulation of processed PINK1 species in several mitochondrial compartments, including the matrix. Knockdown of Lon did not decrease mitochondrial membrane potential or trigger activation of the mitochondrial unfolded protein stress response (UPRmt), indicating that PINK1 accumulation in Lon-deficient animals is not a secondary consequence of mitochondrial depolarization or the UPRmt. Moreover, the influence of Lon on PINK1 abundance was highly specific, as Lon inactivation had little or no effect on the abundance of other mitochondrial proteins. Further studies indicated that the processed forms of PINK1 that accumulate upon Lon inactivation are capable of activating the PINK1-Parkin pathway in vivo. Our findings thus suggest that Lon plays an essential role in regulating the PINK1-Parkin pathway by promoting the degradation of PINK1 in the matrix of healthy mitochondria. PMID:24874806

Thomas, Ruth E.; Andrews, Laurie A.; Burman, Jonathon L.; Lin, Wen-Yang; Pallanck, Leo J.

2014-01-01

283

Involvement of Two Latex-Clearing Proteins during Rubber Degradation and Insights into the Subsequent Degradation Pathway Revealed by the Genome Sequence of Gordonia polyisoprenivorans Strain VH2  

PubMed Central

The increasing production of synthetic and natural poly(cis-1,4-isoprene) rubber leads to huge challenges in waste management. Only a few bacteria are known to degrade rubber, and little is known about the mechanism of microbial rubber degradation. The genome of Gordonia polyisoprenivorans strain VH2, which is one of the most effective rubber-degrading bacteria, was sequenced and annotated to elucidate the degradation pathway and other features of this actinomycete. The genome consists of a circular chromosome of 5,669,805 bp and a circular plasmid of 174,494 bp with average GC contents of 67.0% and 65.7%, respectively. It contains 5,110 putative protein-coding sequences, including many candidate genes responsible for rubber degradation and other biotechnically relevant pathways. Furthermore, we detected two homologues of a latex-clearing protein, which is supposed to be a key enzyme in rubber degradation. The deletion of these two genes for the first time revealed clear evidence that latex-clearing protein is essential for the microbial utilization of rubber. Based on the genome sequence, we predict a pathway for the microbial degradation of rubber which is supported by previous and current data on transposon mutagenesis, deletion mutants, applied comparative genomics, and literature search. PMID:22327575

Hiessl, Sebastian; Schuldes, Jörg; Thürmer, Andrea; Halbsguth, Tobias; Bröker, Daniel; Angelov, Angel; Liebl, Wolfgang; Daniel, Rolf

2012-01-01

284

Competition in chemostat culture between Pseudomonas strains that use different pathways for the degradation of toluene.  

PubMed Central

Pseudomonas putida mt-2, P. cepacia G4, P. mendocina KR1, and P. putida F1 degrade toluene through different pathways. In this study, we compared the competition behaviors of these strains in chemostat culture at a low growth rate (D = 0.05 h-1), with toluene as the sole source of carbon and energy. Either toluene or oxygen was growth limiting. Under toluene-limiting conditions, P. mendocina KR1, in which initial attack is by monooxygenation of the aromatic nucleus at the para position, outcompeted the other three strains. Under oxygen limitation, P. cepacia G4, which hydroxylates toluene in the ortho position, was the most competitive strain. P. putida mt-2, which metabolizes toluene via oxidation of the methyl group, was the least competitive strain under both growth conditions. The apparent superiority of strains carrying toluene degradation pathways that start degradation by hydroxylation of the aromatic nucleus was also found during competition experiments with pairs of strains of P. cepacia, P. fluorescence, and P. putida that were freshly isolated from contaminated soil. PMID:8085826

Duetz, W A; de Jong, C; Williams, P A; van Andel, J G

1994-01-01

285

Uridine-Ribohydrolase Is a Key Regulator in the Uridine Degradation Pathway of Arabidopsis[W  

PubMed Central

Nucleoside degradation and salvage are important metabolic pathways but hardly understood in plants. Recent work on human pathogenic protozoans like Leishmania and Trypanosoma substantiates an essential function of nucleosidase activity. Plant nucleosidases are related to those from protozoans and connect the pathways of nucleoside degradation and salvage. Here, we describe the cloning of such an enzyme from Arabidopsis thaliana, Uridine-Ribohydrolase 1 (URH1) and the characterization by complementation of a yeast mutant. Furthermore, URH1 was synthesized as a recombinant protein in Escherichia coli. The pure recombinant protein exhibited highest hydrolase activity for uridine, followed by inosine and adenosine, the corresponding Km values were 0.8, 1.4, and 0.7 mM, respectively. In addition, URH1 was able to cleave the cytokinin derivative isopentenyladenine-riboside. Promoter ?-glucuronidase fusion studies revealed that URH1 is mainly transcribed in the vascular cells of roots and in root tips, guard cells, and pollen. Mutants expressing the Arabidopsis enzyme or the homolog from rice (Oryza sativa) exhibit resistance toward toxic fluorouridine, fluorouracil, and fluoroorotic acid, providing clear evidence for a pivotal function of URH1 as regulative in pyrimidine degradation. Moreover, mutants with increased and decreased nucleosidase activity are delayed in germination, indicating that this enzyme activity must be well balanced in the early phase of plant development. PMID:19293370

Jung, Benjamin; Flörchinger, Martin; Kunz, Hans-Henning; Traub, Michaela; Wartenberg, Ruth; Jeblick, Wolfgang; Neuhaus, H. Ekkehard; Möhlmann, Torsten

2009-01-01

286

Systematic Unraveling of the Unsolved Pathway of Nicotine Degradation in Pseudomonas  

PubMed Central

Microorganisms such as Pseudomonas putida play important roles in the mineralization of organic wastes and toxic compounds. To comprehensively and accurately elucidate key processes of nicotine degradation in Pseudomonas putida, we measured differential protein abundance levels with MS-based spectral counting in P. putida S16 grown on nicotine or glycerol, a non-repressive carbon source. In silico analyses highlighted significant clustering of proteins involved in a functional pathway in nicotine degradation. The transcriptional regulation of differentially expressed genes was analyzed by using quantitative reverse transcription-PCR. We observed the following key results: (i) The proteomes, containing 1,292 observed proteins, provide a detailed view of enzymes involved in nicotine metabolism. These proteins could be assigned to the functional groups of transport, detoxification, and amino acid metabolism. There were significant differences in the cytosolic protein patterns of cells growing in a nicotine medium and those in a glycerol medium. (ii) The key step in the conversion of 3-succinoylpyridine to 6-hydroxy-3-succinoylpyridine was catalyzed by a multi-enzyme reaction consisting of a molybdopeterin binding oxidase (spmA), molybdopterin dehydrogenase (spmB), and a (2Fe-2S)-binding ferredoxin (spmC) with molybdenum molybdopterin cytosine dinucleotide as a cofactor. (iii) The gene of a novel nicotine oxidoreductase (nicA2) was cloned, and the recombinant protein was characterized. The proteins and functional pathway identified in the current study represent attractive targets for degradation of environmental toxic compounds. PMID:24204321

Tang, Hongzhi; Zhang, Kunzhi; Yao, Yuxiang; Xu, Ping

2013-01-01

287

Degradation pathways of low-ethoxylated nonylphenols by isolated bacteria using an improved method.  

PubMed

Nonylphenol ethoxylates (NPEOs) with low ethoxylation degree (NPav?EO; containing two ethoxy units on average) and estrogenic properties are the intermediate products of nonionic surfactant NPEOs. To better understand the environmental fate of low-ethoxylated NPEOs, phylogenetically diverse low-ethoxylated NPEO-degrading bacteria were isolated from activated sludge using gellan gum as the gelling reagent. Four isolates belonging to four genera, i.e., Pseudomonas sp. NP522b in ?-Proteobacteria, Variovorax sp. NP427b and Ralstonia sp. NP47a in ?-Proteobacteria, and Sphingomonas sp. NP42a in ?-Proteobacteria were acquired. Ralstonia sp. NP47a or Sphingomonas sp. NP42a, have not been reported for the degradation of low-ethoxylated NPEOs previously. The biotransformation pathways of these isolates were investigated. The first three strains (NP522b, NP427b, and NP47a) exhibited high NPav?EO oxidation ability by oxidizing the polyethoxy (EO) chain to form low-ethoxylated nonylphenoxy carboxylates, and then further oxidizing the alkyl chain to form carboxyalkylphenol polyethoxycarboxylates. Furthermore, Sphingomonas sp. NP42a degraded NPav2EO through a nonoxidative pathway with nonylphenol monoethoxylate as the dominant product. PMID:23943000

Zhang, Yu; Gu, Xin; Zhang, Jing; Yang, Min

2014-08-01

288

Genetic immunization based on the ubiquitin-fusion degradation pathway against Trypanosoma cruzi  

SciTech Connect

Cytotoxic CD8{sup +} T cells are particularly important to the development of protective immunity against the intracellular protozoan parasite, Trypanosoma cruzi, the etiological agent of Chagas disease. We have developed a new effective strategy of genetic immunization by activating CD8{sup +} T cells through the ubiquitin-fusion degradation (UFD) pathway. We constructed expression plasmids encoding the amastigote surface protein-2 (ASP-2) of T. cruzi. To induce the UFD pathway, a chimeric gene encoding ubiquitin fused to ASP-2 (pUB-ASP-2) was constructed. Mice immunized with pUB-ASP-2 presented lower parasitemia and longer survival period, compared with mice immunized with pASP-2 alone. Depletion of CD8{sup +} T cells abolished protection against T. cruzi in mice immunized with pUB-ASP-2 while depletion of CD4{sup +} T cells did not influence the effective immunity. Mice deficient in LMP2 or LMP7, subunits of immunoproteasomes, were not able to develop protective immunity induced. These results suggest that ubiquitin-fused antigens expressed in antigen-presenting cells were effectively degraded via the UFD pathway, and subsequently activated CD8{sup +} T cells. Consequently, immunization with pUB-ASP-2 was able to induce potent protective immunity against infection of T. cruzi.

Chou, Bin [Department of Microbiology and Immunology, Faculty of Medicine, Fukuoka University, 7-45-1 Nanakuma, Jonan-ku, Fukuoka 814-0180 (Japan) [Department of Microbiology and Immunology, Faculty of Medicine, Fukuoka University, 7-45-1 Nanakuma, Jonan-ku, Fukuoka 814-0180 (Japan); Department of Parasitology, Graduate School of Medical Science, Kyushu University, Fukuoka 812-8582 (Japan); Hiromatsu, Kenji, E-mail: khiromatsu@fukuoka-u.ac.jp [Department of Microbiology and Immunology, Faculty of Medicine, Fukuoka University, 7-45-1 Nanakuma, Jonan-ku, Fukuoka 814-0180 (Japan)] [Department of Microbiology and Immunology, Faculty of Medicine, Fukuoka University, 7-45-1 Nanakuma, Jonan-ku, Fukuoka 814-0180 (Japan); Hisaeda, Hajime; Duan, Xuefeng; Imai, Takashi [Department of Parasitology, Graduate School of Medical Science, Kyushu University, Fukuoka 812-8582 (Japan)] [Department of Parasitology, Graduate School of Medical Science, Kyushu University, Fukuoka 812-8582 (Japan); Murata, Shigeo; Tanaka, Keiji [Department of Molecular Oncology, The Tokyo Metropolitan Institute of Medical Science, Tokyo 113-8613 (Japan)] [Department of Molecular Oncology, The Tokyo Metropolitan Institute of Medical Science, Tokyo 113-8613 (Japan); Himeno, Kunisuke [Department of Parasitology, Graduate School of Medical Science, Kyushu University, Fukuoka 812-8582 (Japan)] [Department of Parasitology, Graduate School of Medical Science, Kyushu University, Fukuoka 812-8582 (Japan)

2010-02-12

289

Chemical modification and degradation of atrazine in Medicago sativa through multiple pathways.  

PubMed

Atrazine is a member of the triazine herbicide family intensively used to control weeds for crop production. In this study, atrazine residues and its degraded products in alfalfa (Medicago sativa) were characterized using UPLC-TOF-MS/MS. Most of atrazine absorbed in plants was found as chemically modified derivatives like deisopropylated atrazine (DIA), dehydrogenated atrazine (DHA), or methylated atrazine (MEA), and some atrazine derivatives were conjugated through different functional groups such as sugar, glutathione, and amino acids. Interestingly, the specific conjugates DHA+hGSH (homoglutathione) and MEA-HCl+hGSH in alfalfa were detected. These results suggest that atrazine in alfalfa can be degraded through different pathways. The increased activities of glycosyltransferase and glutathione S-transferase were determined to support the atrazine degradation models. The outcome of the work uncovered the detailed mechanism for the residual atrazine accumulation and degradation in alfalfa and will help to evaluate whether the crop is suitable to be cultivated in the atrazine-polluted soil. PMID:25226578

Zhang, Jing Jing; Lu, Yi Chen; Yang, Hong

2014-10-01

290

Sulfamethoxazole degradation by ultrasound/ozone oxidation process in water: kinetics, mechanisms, and pathways.  

PubMed

In this research, sulfamethoxazole (SMX) degradation was investigated using ultrasound (US), ozone (O3) and ultrasound/ozone oxidation process (UOOP). It was proved that ultrasound significantly enhanced SMX ozonation by assisting ozone in producing more hydroxyl radicals in UOOP. Ultrasound also made the rate constants improve by kinetics analysis. When ultrasound was added to the ozonation process, the reaction rate increased by 6-26% under different pH conditions. Moreover, main intermediates oxidized by US, O3 and UOOP system were identified. Although the main intermediates in ozonation and UOOP were similar, the introduction of ultrasound in UOOP had well improved the cleavage of S-N bond. In this condition SMX become much easier to be attacked, which led to enhanced SMX removal rate in UOOP compared to the other two examined processes. Finally, the SMX degradation pathways were proposed. PMID:25107668

Guo, Wan-Qian; Yin, Ren-Li; Zhou, Xian-Jiao; Du, Juan-Shan; Cao, Hai-Ou; Yang, Shan-Shan; Ren, Nan-Qi

2015-01-01

291

Reaction pathways and mechanisms of the electrochemical degradation of phenol on different electrodes.  

PubMed

Laboratory experiments were carried out on the kinetics and pathways of the electrochemical (EC) degradation of phenol at three different types of anodes, Ti/SnO2-Sb, Ti/RuO2, and Pt. Although phenol was oxidised by all of the anodes at a current density of 20 mA/cm2 or a cell voltage of 4.6 V, there was a considerable difference between the three anode types in the effectiveness and performance of EC organic degradation. Phenol was readily mineralized at the Ti/SnO2-Sb anode, but its degradation was much slower at the Ti/RuO2 and Pt anodes. The analytical results of high-performance liquid chromatography (HPLC) and gas chromatography coupled with mass spectrometry (GC/MS) indicated that the intermediate products of EC phenol degradation, including benzoquinone and organic acids, were subsequently oxidised rapidly by the Ti/SnO2-Sb anode, but accumulated in the cells of Ti/RuO2 and Pt. There was also a formation of dark-coloured polymeric compounds and precipitates in the solutions electrolyzed by the Ti/RuO2 and Pt anodes, which was not observed for the Ti/SnO2-Sb cells. It is argued that anodic property not only affects the reaction kinetics of various steps of EC organic oxidation, but also alters the pathway of phenol electrolysis. Favourable surface treatment, such as the SnO2-Sb coating, provides the anode with an apparent catalytic function for rapid organic oxidation that is probably brought about by hydroxyl radicals generated from anodic water electrolysis. PMID:15882890

Li, Xiao-Yan; Cui, Yu-Hong; Feng, Yu-Jie; Xie, Zhao-Ming; Gu, Ji-Dong

2005-05-01

292

The proteasome and the degradation of oxidized proteins: Part III—Redox regulation of the proteasomal system  

PubMed Central

Here, we review shortly the current knowledge on the regulation of the proteasomal system during and after oxidative stress. After addressing the components of the proteasomal system and the degradation of oxidatively damaged proteins in part I and II of this series, we address here which changes in activity undergo the proteasome and the ubiquitin-proteasomal system itself under oxidative conditions. While several components of the proteasomal system undergo direct oxidative modification, a number of redox-regulated events are modulating the proteasomal activity in a way it can address the major tasks in an oxidative stress situation: the removal of oxidized proteins and the adaptation of the cellular metabolism to the stress situation. PMID:24563857

Höhn, Tobias Jung Annika; Grune, Tilman

2014-01-01

293

The cAMP signaling system inhibits the repair of {gamma}-ray-induced DNA damage by promoting Epac1-mediated proteasomal degradation of XRCC1 protein in human lung cancer cells  

SciTech Connect

Highlights: Black-Right-Pointing-Pointer cAMP signaling system inhibits repair of {gamma}-ray-induced DNA damage. Black-Right-Pointing-Pointer cAMP signaling system inhibits DNA damage repair by decreasing XRCC1 expression. Black-Right-Pointing-Pointer cAMP signaling system decreases XRCC1 expression by promoting its proteasomal degradation. Black-Right-Pointing-Pointer The promotion of XRCC1 degradation by cAMP signaling system is mediated by Epac1. -- Abstract: Cyclic AMP is involved in the regulation of metabolism, gene expression, cellular growth and proliferation. Recently, the cAMP signaling system was found to modulate DNA-damaging agent-induced apoptosis by regulating the expression of Bcl-2 family proteins and inhibitors of apoptosis. Thus, we hypothesized that the cAMP signaling may modulate DNA repair activity, and we investigated the effects of the cAMP signaling system on {gamma}-ray-induced DNA damage repair in lung cancer cells. Transient expression of a constitutively active mutant of stimulatory G protein (G{alpha}sQL) or treatment with forskolin, an adenylyl cyclase activator, augmented radiation-induced DNA damage and inhibited repair of the damage in H1299 lung cancer cells. Expression of G{alpha}sQL or treatment with forskolin or isoproterenol inhibited the radiation-induced expression of the XRCC1 protein, and exogenous expression of XRCC1 abolished the DNA repair-inhibiting effect of forskolin. Forskolin treatment promoted the ubiquitin and proteasome-dependent degradation of the XRCC1 protein, resulting in a significant decrease in the half-life of the protein after {gamma}-ray irradiation. The effect of forskolin on XRCC1 expression was not inhibited by PKA inhibitor, but 8-pCPT-2 Prime -O-Me-cAMP, an Epac-selective cAMP analog, increased ubiquitination of XRCC1 protein and decreased XRCC1 expression. Knockdown of Epac1 abolished the effect of 8-pCPT-2 Prime -O-Me-cAMP and restored XRCC1 protein level following {gamma}-ray irradiation. From these results, we conclude that the cAMP signaling system inhibits the repair of {gamma}-ray-induced DNA damage by promoting the ubiquitin-proteasome dependent degradation of XRCC1 in an Epac-dependent pathway in lung cancer cells.

Cho, Eun-Ah [Department of Biochemistry and Molecular Biology, Cancer Research Center, Seoul National University College of Medicine, Seoul 110-799 (Korea, Republic of)] [Department of Biochemistry and Molecular Biology, Cancer Research Center, Seoul National University College of Medicine, Seoul 110-799 (Korea, Republic of); Juhnn, Yong-Sung, E-mail: juhnn@snu.ac.kr [Department of Biochemistry and Molecular Biology, Cancer Research Center, Seoul National University College of Medicine, Seoul 110-799 (Korea, Republic of)] [Department of Biochemistry and Molecular Biology, Cancer Research Center, Seoul National University College of Medicine, Seoul 110-799 (Korea, Republic of)

2012-06-01

294

Perturbing the Ubiquitin Pathway Reveals How Mitosis Is Hijacked to Denucleate and Regulate Cell Proliferation and Differentiation In Vivo  

PubMed Central

Background The eye lens presents a unique opportunity to explore roles for specific molecules in cell proliferation, differentiation and development because cells remain in place throughout life and, like red blood cells and keratinocytes, they go through the most extreme differentiation, including removal of nuclei and cessation of protein synthesis. Ubiquitination controls many critical cellular processes, most of which require specific lysines on ubiquitin (Ub). Of the 7 lysines (K) least is known about effects of modification of K6. Methodology and Principal Findings We replaced K6 with tryptophan (W) because K6 is the most readily modified K and W is the most structurally similar residue to biotin. The backbone of K6W-Ub is indistinguishable from that of Wt-Ub. K6W-Ub is effectively conjugated and deconjugated but the conjugates are not degraded via the ubiquitin proteasome pathways (UPP). Expression of K6W-ubiquitin in the lens and lens cells results in accumulation of intracellular aggregates and also slows cell proliferation and the differentiation program, including expression of lens specific proteins, differentiation of epithelial cells into fibers, achieving proper fiber cell morphology, and removal of nuclei. The latter is critical for transparency, but the mechanism by which cell nuclei are removed has remained an age old enigma. This was also solved by expressing K6W-Ub. p27kip, a UPP substrate accumulates in lenses which express K6W-Ub. This precludes phosphorylation of nuclear lamin by the mitotic kinase, a prerequisite for disassembly of the nuclear membrane. Thus the nucleus remains intact and DNAseII? neither gains entry to the nucleus nor degrades the DNA. These results could not be obtained using chemical proteasome inhibitors that cannot be directed to specific tissues. Conclusions and Significance K6W-Ub provides a novel, genetic means to study functions of the UPP because it can be targeted to specific cells and tissues. A fully functional UPP is required to execute most stages of lens differentiation, specifically removal of cell nuclei. In the absence of a functional UPP, small aggregate prone, cataractous lenses are formed. PMID:20975996

Wawrousek, Eric; Liu, Qing; Avidan, Orna; Cvekl, Ales; Yang, Ying; Haririnia, Aydin; Storaska, Andrew; Fushman, David; Kuszak, Jer; Dudek, Edward; Smith, Donald; Taylor, Allen

2010-01-01

295

A polyomic approach to elucidate the fluoranthene-degradative pathway in Mycobacterium vanbaalenii PYR-1.  

PubMed

Mycobacterium vanbaalenii PYR-1 is capable of degrading a wide range of high-molecular-weight polycyclic aromatic hydrocarbons (PAHs), including fluoranthene. We used a combination of metabolomic, genomic, and proteomic technologies to investigate fluoranthene degradation in this strain. Thirty-seven fluoranthene metabolites including potential isomers were isolated from the culture medium and analyzed by high-performance liquid chromatography, gas chromatography-mass spectrometry, and UV-visible absorption. Total proteins were separated by one-dimensional gel and analyzed by liquid chromatography-tandem mass spectrometry in conjunction with the M. vanbaalenii PYR-1 genome sequence (http://jgi.doe.gov), which resulted in the identification of 1,122 proteins. Among them, 53 enzymes were determined to be likely involved in fluoranthene degradation. We integrated the metabolic information with the genomic and proteomic results and proposed pathways for the degradation of fluoranthene. According to our hypothesis, the oxidation of fluoranthene is initiated by dioxygenation at the C-1,2, C-2,3, and C-7,8 positions. The C-1,2 and C-2,3 dioxygenation routes degrade fluoranthene via fluorene-type metabolites, whereas the C-7,8 routes oxidize fluoranthene via acenaphthylene-type metabolites. The major site of dioxygenation is the C-2,3 dioxygenation route, which consists of 18 enzymatic steps via 9-fluorenone-1-carboxylic acid and phthalate with the initial ring-hydroxylating oxygenase, NidA3B3, oxidizing fluoranthene to fluoranthene cis-2,3-dihydrodiol. Nonspecific monooxygenation of fluoranthene with subsequent O methylation of dihydroxyfluoranthene also occurs as a detoxification reaction. PMID:17449607

Kweon, Ohgew; Kim, Seong-Jae; Jones, Richard C; Freeman, James P; Adjei, Michael D; Edmondson, Ricky D; Cerniglia, Carl E

2007-07-01

296

Transfer of the high GC cyclohexane carboxylate degradation pathway from Rhodopseudomonas palustris to Escherichia coli for production of biotin  

PubMed Central

This work demonstrates the transfer of the five gene cyclohexane carboxylate (CHC) degradation pathway from the high GC alphaproteobacterium Rhodopseudomonas palustris to Escherichia coli, a gammaproteobacterium. The degradation product of this pathway is pimeloyl-CoA, a key metabolite in E. coli's biotin biosynthetic pathway. This pathway is useful for biotin overproduction in E. coli, however, the expression of GC-rich genes is troublesome in this host. When the native R. palustris CHC degradation pathway is transferred to a ?bioH pimeloyl-CoA auxotroph of E. coli, it is unable to complement growth in the presence of CHC. To overcome this expression problem we redesigned the operon with decreased GC content and removed stretches of high GC intergenic DNA which comprise the 5' untranslated region of each gene, replacing these features with shorter low GC sequences. We show this synthetic construct enables growth of the ?bioH strain in the presence of CHC. When the synthetic degradation pathway is overexpressed in conjunction with the downstream genes for biotin biosynthesis, we measured significant accumulation of biotin in the growth medium, showing that the pathway transfer is successfully integrated with the host metabolism. PMID:18396082

Bernstein, Jeffrey R.; Bulter, Thomas; Liao, James C.

2008-01-01

297

Sequential RNA degradation pathways provide a fail-safe mechanism to limit the accumulation of unspliced transcripts in Saccharomyces cerevisiae  

PubMed Central

The nuclear exosome and the nonsense-mediated mRNA decay (NMD) pathways have been implicated in the degradation of distinct unspliced transcripts in Saccharomyces cerevisiae. In this study we show that these two systems can act sequentially on specific unspliced pre-mRNAs to limit their accumulation. Using steady-state and decay analyses, we show that while specific unspliced transcripts rely mostly on NMD or on the nuclear exosome for their degradation, some unspliced RNAs are stabilized only when both the nuclear exosome and NMD are inactivated. We found that the mechanism of degradation of these unspliced pre-mRNAs is not influenced by promoter identity. However, the specificity in the pre-mRNAs degradation pathways can be manipulated by changing the rate of export or retention of these mRNAs. For instance, reducing the nuclear export of pre-mRNAs mostly degraded by NMD results in a higher fraction of unspliced transcripts degraded by the nuclear exosome. Reciprocally, inactivating the Mlp retention factors results in a higher fraction of unspliced transcripts degraded by NMD for precursors normally targeted by the nuclear exosome. Overall, these results demonstrate that a functional redundancy exists between nuclear and cytoplasmic degradation pathways for unspliced pre-mRNAs, and suggest that the degradation routes of these species are mainly determined by the efficiency of their nuclear export rates. The presence of these two sequential degradation pathways for unspliced pre-mRNAs underscores the importance of limiting their accumulation and might serve as a fail-safe mechanism to prevent the expression of these nonfunctional RNAs. PMID:22753783

Sayani, Shakir; Chanfreau, Guillaume F.

2012-01-01

298

Ubiquitin ligase Nedd4 promotes alpha-synuclein degradation by the endosomal-lysosomal pathway.  

PubMed

?-Synuclein is an abundant brain protein that binds to lipid membranes and is involved in the recycling of presynaptic vesicles. In Parkinson disease, ?-synuclein accumulates in intraneuronal inclusions often containing ubiquitin chains. Here we show that the ubiquitin ligase Nedd4, which functions in the endosomal-lysosomal pathway, robustly ubiquitinates ?-synuclein, unlike ligases previously implicated in its degradation. Purified Nedd4 recognizes the carboxyl terminus of ?-synuclein (residues 120-133) and attaches K63-linked ubiquitin chains. In human cells, Nedd4 overexpression enhances ?-synuclein ubiquitination and clearance by a lysosomal process requiring components of the endosomal-sorting complex required for transport. Conversely, Nedd4 down-regulation increases ?-synuclein content. In yeast, disruption of the Nedd4 ortholog Rsp5p decreases ?-synuclein degradation and enhances inclusion formation and ?-synuclein toxicity. In human brains, Nedd4 is present in pigmented neurons and is expressed especially strongly in neurons containing Lewy bodies. Thus, ubiquitination by Nedd4 targets ?-synuclein to the endosomal-lysosomal pathway and, by reducing ?-synuclein content, may help protect against the pathogenesis of Parkinson disease and other ?-synucleinopathies. PMID:21953697

Tofaris, George K; Kim, Hyoung Tae; Hourez, Raphael; Jung, Jin-Woo; Kim, Kwang Pyo; Goldberg, Alfred L

2011-10-11

299

Degradation pathway of quinolines in a biofilm system under denitrifying conditions  

SciTech Connect

This article reports for the first time the degradation pathways of quinoline, isoquinoline, and methylquinolines by a mixed culture in a biofilm under nitrate-reducing conditions. A simple reverse-phase high-performance liquid chromatography method using ultraviolet detection at 223 nm for determination of seven quinoline analogues and 15 metabolites was developed, and gas chromatography--mass spectrometry and thin-layer chromatography analyses were used for identification. The inhibition of nitrification by the parent compounds and their degradation products was assessed by a nitrification toxicity test called MINNTOX. Quinoline and 3-, 4-, 6-, and 8-methylquinoline were all transformed by hydroxylation into their 2-hydroxyquinoline analogues (2-quinolinones), and isoquinoline was transformed into 1-hydroxyisoquinoline. 2-Methylquinoline was not transformed by this microcosm, likely due to the blockage at position 2 by the methyl group. The hydroxylated metabolites of isoquinoline and quinolines methylated at the heterocyclic ring were not transformed further, whereas metabolites of quinoline and quinolines methylated at the homocyclic ring were hydrogenated at position 3 and 4, and the resulting 3,4-dihydro-2-quinolinone analogues accumulated. Of these metabolites, only 3,4-dihydro-2-quinolinone from the degradation of quinoline was further transformed into unidentified products. All quinolines and their metabolites had inhibiting effects on the nitrifying bacteria at the same level (ppm) in the applied bioassay, indicating that the inhibition of the compounds was not influenced by the initial transformation reactions.

Johansen, S.S.; Arvin, E.; Mosbaek, H. [Technical Univ. of Denmark, Lyngby (Denmark). Dept. of Environmental Science and Engineering; Hansen, A.B. [National Environmental Research Inst., Roskilde (Denmark). Dept. of Environmental Chemistry

1997-09-01

300

Endothelin-converting Enzymes Degrade Intracellular ?-Amyloid Produced within the Endosomal/Lysosomal Pathway and Autophagosomes*  

PubMed Central

Impairments in A? removal are increasingly being considered as a possible cause for the abnormal A? build-up typical of Alzheimer disease. Of particular interest is a pool of A? that accumulates intraneuronally and may contribute to neuronal toxicity. The mechanism for intraneuronal accumulation, however, is not well understood and is commonly attributed to impaired removal of extracellular A? by neurons. Based on the intracellular distribution of the well established A? degrading enzymes, ECE-1 and ECE-2, we tested whether impairments in their catalytic activity could lead to intracellular A? accumulation. Using SH-SY5Y cells overexpressing wild-type amyloid precursor protein and pharmacological inhibition of endogenous ECE activity, we found that ECEs participate in the degradation of at least two distinct pools of A?; one destined for secretion and the other being produced and degraded within the endosomal-autophagic-lysosomal pathways. Although ECE-1 regulates both pools of A?, ECE-2 regulates mainly the intracellular pool of the peptide. Consistent with this result, ECE-2 was found to co-localize with markers of the endosomal/lysosomal pathway but not with a trans-Golgi network marker. Furthermore, ECE-2 was detected in autophagic vesicles in cells treated with chloroquine. Under these conditions, ECE inhibition produced significantly higher elevations in intracellular A? than chloroquine treatment alone. This study highlights the existence of A? clearance mechanisms by ECEs at intracellular sites of production. Alterations in ECE activity may be considered as a cause for increased intraneuronal A? in Alzheimer disease. PMID:23283972

Pacheco-Quinto, Javier; Eckman, Elizabeth A.

2013-01-01

301

Identification of Genes and Pathways Related to Phenol Degradation in Metagenomic Libraries from Petroleum Refinery Wastewater  

PubMed Central

Two fosmid libraries, totaling 13,200 clones, were obtained from bioreactor sludge of petroleum refinery wastewater treatment system. The library screening based on PCR and biological activity assays revealed more than 400 positive clones for phenol degradation. From these, 100 clones were randomly selected for pyrosequencing in order to evaluate the genetic potential of the microorganisms present in wastewater treatment plant for biodegradation, focusing mainly on novel genes and pathways of phenol and aromatic compound degradation. The sequence analysis of selected clones yielded 129,635 reads at an estimated 17-fold coverage. The phylogenetic analysis showed Burkholderiales and Rhodocyclales as the most abundant orders among the selected fosmid clones. The MG-RAST analysis revealed a broad metabolic profile with important functions for wastewater treatment, including metabolism of aromatic compounds, nitrogen, sulphur and phosphorus. The predicted 2,276 proteins included phenol hydroxylases and cathecol 2,3- dioxygenases, involved in the catabolism of aromatic compounds, such as phenol, byphenol, benzoate and phenylpropanoid. The sequencing of one fosmid insert of 33 kb unraveled the gene that permitted the host, Escherichia coli EPI300, to grow in the presence of aromatic compounds. Additionally, the comparison of the whole fosmid sequence against bacterial genomes deposited in GenBank showed that about 90% of sequence showed no identity to known sequences of Proteobacteria deposited in the NCBI database. This study surveyed the functional potential of fosmid clones for aromatic compound degradation and contributed to our knowledge of the biodegradative capacity and pathways of microbial assemblages present in refinery wastewater treatment system. PMID:23637911

Silva, Cynthia C.; Hayden, Helen; Sawbridge, Tim; Mele, Pauline; De Paula, Sérgio O.; Silva, Lívia C. F.; Vidigal, Pedro M. P.; Vicentini, Renato; Sousa, Maíra P.; Torres, Ana Paula R.; Santiago, Vânia M. J.; Oliveira, Valéria M.

2013-01-01

302

Opine utilization by Agrobacterium spp.: octopine-type Ti plasmids encode two pathways for mannopinic acid degradation.  

PubMed Central

Octopine-type strains of Agrobacterium tumefaciens degrade the opine mannopinic acid through a specific pathway which involves cleavage of the molecule at the C--N bond between the amino acid and the sugar moieties. Mannose was identified as a product of the reaction. This pathway was inducible by mannopinic and agropinic acids, but not by mannopine or agropine, the two other mannityl opines. The transport system for this pathway appeared to be specific for mannopinic acid. A second, nonspecific pathway for mannopinic acid degradation was also identified. This involved some of the catabolic functions associated with the metabolism of mannopine and agropine. This second pathway was inducible by mannopine and agropine but not by mannopinic or agropinic acids. The transport system for this pathway appeared to have a broad specificity. Transposon Tn5 insertion mutants affected in the specific catabolic pathway were isolated and analyzed. These mutants continued to catabolize mannopine and agropine. Both mapped to a region of the Ti plasmid previously shown to be associated with the catabolism of mannopinic acid. Restriction enzyme analysis of the Ti plasmid from strain 89.10, an octopine strain that is naturally unable to utilize mannopinic acid, showed a deletion in this same region encoding the specific mannopinic acid degradation pathway. Analysis of recombinant clones showed that the second, nonspecific pathway was encoded in a region of the Ti plasmid associated with mannopine and agropine catabolism. This region shared no structural overlap with the segment of the plasmid encoding the specific mannopinic acid degradative pathway. Images PMID:2838452

Dessaux, Y; Guyon, P; Petit, A; Tempé, J; Demarez, M; Legrain, C; Tate, M E; Farrand, S K

1988-01-01

303

Degradation of 4-Chlorophenol via the meta Cleavage Pathway by Comamonas testosteroni JH5  

PubMed Central

Comamonas testosteroni JH5 used 4-chlorophenol (4-CP) as its sole source of energy and carbon up to a concentration of 1.8 mM, accompanied by the stoichiometric release of chloride. The degradation of 4-CP mixed with the isomeric 2-CP by resting cells led to the accumulation of 3-chlorocatechol (3-CC), which inactivated the catechol 2,3-dioxygenase. As a result, further 4-CP breakdown was inhibited and 4-CC accumulated as a metabolite. In the crude extract of 4-CP-grown cells, catechol 1,2-dioxygenase and muconate cycloisomerase activities were not detected, whereas the activities of catechol 2,3-dioxygenase, 2-hydroxymuconic semialdehyde dehydrogenase, 2-hydroxymuconic semialdehyde hydrolase, and 2-oxopent-4-enoate hydratase were detected. These enzymes of the meta cleavage pathway showed activity with 4-CC and with 5-chloro-2-hydroxymuconic semialdehyde. The activities of the dioxygenase and semialdehyde dehydrogenase were constitutive. Two key metabolites of the meta cleavage pathway, the meta cleavage product (5-chloro-2-hydroxymuconic semialdehyde) and 5-chloro-2-hydroxymuconic acid, were detected. Thus, our previous postulation that C. testosteroni JH5 uses the meta cleavage pathway for the complete mineralization of 4-CP was confirmed. PMID:16535738

Hollender, J.; Hopp, J.; Dott, W.

1997-01-01

304

CHIP: A Co-chaperone for Degradation by the Proteasome.  

PubMed

Protein homeostasis relies on a balance between protein folding and protein degradation. Molecular chaperones like Hsp70 and Hsp90 fulfil well-defined roles in protein folding and conformational stability via ATP dependent reaction cycles. These folding cycles are controlled by associations with a cohort of non-client protein co-chaperones, such as Hop, p23 and Aha1. Pro-folding co-chaperones facilitate the transit of the client protein through the chaperone mediated folding process. However, chaperones are also involved in ubiquitin-mediated proteasomal degradation of client proteins. Similar to folding complexes, the ability of chaperones to mediate protein degradation is regulated by co-chaperones, such as the C terminal Hsp70 binding protein (CHIP). CHIP binds to Hsp70 and Hsp90 chaperones through its tetratricopeptide repeat (TPR) domain and functions as an E3 ubiquitin ligase using a modified RING finger domain (U-box). This unique combination of domains effectively allows CHIP to network chaperone complexes to the ubiquitin-proteasome system. This chapter reviews the current understanding of CHIP as a co-chaperone that switches Hsp70/Hsp90 chaperone complexes from protein folding to protein degradation. PMID:25487024

Edkins, Adrienne L

2015-01-01

305

Dexamethasone-induced selenoprotein S degradation is required for adipogenesis  

PubMed Central

Although adipogenesis is associated with induction of endoplasmic reticulum (ER) stress, the role of selenoprotein S (SEPS1), an ER resident selenoprotein known to regulate ER stress and ER-associated protein degradation, is unknown. We found an inverse relationship between SEPS1 level in adipose tissue and adiposity in mice. While SEPS1 expression was increased during adipogenesis, a markedly reduced SEPS1 protein level was found in the early phase of adipogenesis due to dexamethasone (DEX)-induced proteosomal degradation of SEPS1. Overexpression of SEPS1 in the early phase of cell differentiation resulted in impairment of adipogenesis with reduced levels of CCAAT/enhancer binding protein ? and other adipocyte marker genes during the course of adipogenesis. Conversely, knockdown of SEPS1 resulted in the promotion of adipogenesis. Additionally, altered SEPS1 expression was associated with changes in expression of ER stress marker genes in the early phase of adipogenesis, and ubiquitin-proteasome system (UPS)-related ubiquitination and proteasome function. Our study reveals that SEPS1 is a novel anti-adipogenic selenoprotein that modulates ER stress- and UPS-dependent adipogenesis. Our results also identifies a novel function of DEX in the regulation of adipogenesis through induction of SEPS1 degradation. Taken together, DEX-dependent degradation of SEPS1 in the early phase of adipogenesis is necessary for initiating ER stress- and UPS-dependent maturation of adipocytes. PMID:23687306

Kim, Choon Young; Kim, Kee-Hong

2013-01-01

306

Physiology of deletion mutants in the anaerobic ?-myrcene degradation pathway in Castellaniella defragrans  

PubMed Central

Background Monoterpenes present a large and versatile group of unsaturated hydrocarbons of plant origin with widespread use in the fragrance as well as food industry. The anaerobic ?-myrcene degradation pathway in Castellaniella defragrans strain 65Phen differs from well known aerobic, monooxygenase-containing pathways. The initial enzyme linalool dehydratase-isomerase ldi/LDI catalyzes the hydration of ?-myrcene to (S)-(+)-linalool and its isomerization to geraniol. A high-affinity geraniol dehydrogenase geoA/GeDH and a geranial dehydrogenase geoB/GaDH contribute to the formation of geranic acid. A genetic system was for the first time applied for the betaproteobacterium to prove in vivo the relevance of the linalool dehydratase-isomerase and the geraniol dehydrogenase. In-frame deletion cassettes were introduced by conjugation and two homologous recombination events. Results Polar effects were absent in the in-frame deletion mutants C. defragrans ?ldi and C. defragrans ?geoA. The physiological characterization of the strains demonstrated a requirement of the linalool dehydratase-isomerase for growth on acyclic monoterpenes, but not on cyclic monoterpenes. The deletion of geoA resulted in a phenotype with hampered growth rate on monoterpenes as sole carbon and energy source as well as reduced biomass yields. Enzyme assays revealed the presence of a second geraniol dehydrogenase. The deletion mutants were in trans complemented with the broad-host range expression vector pBBR1MCS-4ldi and pBBR1MCS-2geoA, restoring in both cases the wild type phenotype. Conclusions In-frame deletion mutants of genes in the anaerobic ?-myrcene degradation revealed novel insights in the in vivo function. The deletion of a high-affinity geraniol dehydrogenase hampered, but did not preclude growth on monoterpenes. A second geraniol dehydrogenase activity was present that contributes to the ?-myrcene degradation pathway. Growth on cyclic monoterpenes independent of the initial enzyme LDI suggests the presence of a second enzyme system activating unsaturated hydrocarbons. PMID:22947208

2012-01-01

307

Carbon and chlorine isotope analysis to identify abiotic degradation pathways of 1,1,1-trichloroethane.  

PubMed

This study investigates dual C-Cl isotope fractionation during 1,1,1-TCA transformation by heat-activated persulfate (PS), hydrolysis/dehydrohalogenation (HY/DH) and Fe(0). Compound-specific chlorine isotope analysis of 1,1,1-TCA was performed for the first time, and transformation-associated isotope fractionation ? bulk C and ? bulk Cl values were -4.0 ± 0.2‰ and no chlorine isotope fractionation with PS, -1.6 ± 0.2‰ and -4.7 ± 0.1‰ for HY/DH, -7.8 ± 0.4‰ and -5.2 ± 0.2‰ with Fe(0). Distinctly different dual isotope slopes (??13C/??37Cl): ? with PS, 0.33 ± 0.04 for HY/DH and 1.5 ± 0.1 with Fe(0) highlight the potential of this approach to identify abiotic degradation pathways of 1,1,1-TCA in the field. The trend observed with PS agreed with a C-H bond oxidation mechanism in the first reaction step. For HY/DH and Fe(0) pathways, different slopes were obtained although both pathways involve cleavage of a C-Cl bond in their initial reaction step. In contrast to the expected larger primary carbon isotope effects relative to chlorine for C-Cl bond cleavage, ? bulk C < ? bulk Cl was observed for HY/DH and in a similar range for reduction by Fe(0), suggesting the contribution of secondary chlorine isotope effects. Therefore, different magnitude of secondary chlorine isotope effects could at least be partly responsible for the distinct slopes between HY/DH and Fe(0) pathways. Following this dual isotope approach, abiotic transformation processes can unambiguously be identified and quantified. PMID:25379605

Palau, Jordi; Shouakar-Stash, Orfan; Hunkeler, Daniel

2014-12-16

308

SCF(Slmb) E3 ligase-mediated degradation of Expanded is inhibited by the Hippo pathway in Drosophila.  

PubMed

Deregulation of the evolutionarily conserved Hippo pathway has been implicated in abnormal development of animals and in several types of cancer. One mechanism of Hippo pathway regulation is achieved by controlling the stability of its regulatory components. However, the executive E3 ligases that are involved in this process, and how the process is regulated, remain poorly defined. In this study, we identify, through a genetic candidate screen, the SCF(Slmb) E3 ligase as a novel negative regulator of the Hippo pathway in Drosophila imaginal tissues via mediation of the degradation of Expanded (Ex). Mechanistic study shows that Slmb-mediated degradation of Ex is inhibited by the Hippo signaling. Considering the fact that Hippo signaling suppresses the transcription of ex, we propose that the Hippo pathway employs a double security mechanism to ensure fine-tuned homeostasis during development. PMID:25522691

Zhang, Hongtao; Li, Changqing; Chen, Hanqing; Wei, Chuanxian; Dai, Fei; Wu, Honggang; Dui, Wen; Deng, Wu-Min; Jiao, Renjie

2015-01-01

309

Unusual Starch Degradation Pathway via Cyclodextrins in the Hyperthermophilic Sulfate-Reducing Archaeon Archaeoglobus fulgidus Strain 7324?  

PubMed Central

The hyperthermophilic archaeon Archaeoglobus fulgidus strain 7324 has been shown to grow on starch and sulfate and thus represents the first sulfate reducer able to degrade polymeric sugars. The enzymes involved in starch degradation to glucose 6-phosphate were studied. In extracts of starch-grown cells the activities of the classical starch degradation enzymes, ?-amylase and amylopullulanase, could not be detected. Instead, evidence is presented here that A. fulgidus utilizes an unusual pathway of starch degradation involving cyclodextrins as intermediates. The pathway comprises the combined action of an extracellular cyclodextrin glucanotransferase (CGTase) converting starch to cyclodextrins and the intracellular conversion of cyclodextrins to glucose 6-phosphate via cyclodextrinase (CDase), maltodextrin phosphorylase (Mal-P), and phosphoglucomutase (PGM). These enzymes, which are all induced after growth on starch, were characterized. CGTase catalyzed the conversion of starch to mainly ?-cyclodextrin. The gene encoding CGTase was cloned and sequenced and showed highest similarity to a glucanotransferase from Thermococcus litoralis. After transport of the cyclodextrins into the cell by a transport system to be defined, these molecules are linearized via a CDase, catalyzing exclusively the ring opening of the cyclodextrins to the respective maltooligodextrins. These are degraded by a Mal-P to glucose 1-phosphate. Finally, PGM catalyzes the conversion of glucose 1-phosphate to glucose 6-phosphate, which is further degraded to pyruvate via the modified Embden-Meyerhof pathway. PMID:17921308

Labes, Antje; Schönheit, Peter

2007-01-01

310

The histamine degradative uptake pathway in human vascular endothelial cells and skin fibroblasts is dependent on extracellular Na+ and Cl-  

SciTech Connect

We have previously reported that human vascular endothelial cells and skin fibroblasts carry out degradation of (3H)histamine by a mechanism involving two successive enzymatic steps: imidazole ring tele-methylation by the cells' endogenous methyltransferase and subsequent amine oxidation by an exogenous diamine oxidase. Both histamine and the exogenous second enzyme in the pathway associate with the cells via separate binding sites or receptors. The enzymatic degradation process results in cellular accumulation of the proximal and distal metabolites tele-methylhistamine and 1-methyl-4-imidazoleacetic acid (MIAA). We have now demonstrated that this two-stage histamine degradative pathway is dependent on Na+ and Cl- in the extracellular environment. Accumulation of (3H) histamine-derived products is partially inhibited under conditions of Na+ deprivation and more substantially when Cl- is also withdrawn. The individual tele-methylation and amine oxidation enzymatic reactions themselves are unaffected or actually facilitated under these conditions. This indicates that it is the cellular mechanism for uptake coupled to the degradative pathway which reflects the cation and anion dependency. Restoration of degradative uptake displays a biphasic Na+ concentration curve, suggesting that the uptake process may be driven by multiple components. These findings indicate a role for both inward Na+ and Cl- ion movement in this cellular degradative uptake mechanism.

Haddock, R.C.; Mack, P.; Leal, S.; Baenziger, N.L. (Washington Univ. School of Medicine, St. Louis, MO (USA))

1990-08-25

311

Morpholine Degradation Pathway of Mycobacterium aurum MO1: Direct Evidence of Intermediates by In Situ 1H Nuclear Magnetic Resonance  

PubMed Central

Resting Mycobacterium aurum MO1 cells were incubated with morpholine, a waste from the chemical industry. The kinetics of biodegradation was monitored by using in situ nuclear magnetic resonance (NMR). The incubation medium was directly analyzed by 1H NMR. This technique allowed the unambiguous identification of two intermediates of the metabolic pathway involved in the biodegradation process, glycolate and 2-(2-aminoethoxy)acetate. The latter compound, which was not commercially available, was synthesized, in three steps, from 2-(2-aminoethoxy)ethanol. Quantitative analysis of the kinetics of degradation of morpholine was performed by integrating the signals of the different metabolites in 1H-NMR spectra. Morpholine was degraded within 10 h. The intermediates increased during the first 10 h and finally disappeared after 20 h incubation. Assays of degradation were also carried out with glycolate and ethanolamine, hypothetical intermediates of the morpholine degradation pathway. They were degraded within 4 and 8 h, respectively. Until now, no tool for direct detection of intermediates or even morpholine has been available, consequently, only hypothetical pathways have been proposed. The approach described here gives both qualitative and quantitative information about the metabolic routes used in morpholine degradation by M. aurum MO1. It could be used to investigate many biodegradative processes. PMID:9435073

Combourieu, B.; Besse, P.; Sancelme, M.; Veschambre, H.; Delort, A. M.; Poupin, P.; Truffaut, N.

1998-01-01

312

Degradation pathways of lamotrigine under advanced treatment by direct UV photolysis, hydroxyl radicals, and ozone.  

PubMed

Lamotrigine is recently recognized as a persistent pharmaceutical in the water environment and wastewater effluents. Its degradation was studied under UV and ozone advanced oxidation treatments with reaction kinetics of lamotrigine with ozone (?4 M(-1)s(-1)), hydroxyl radical [(2.1 ± 0.3) × 10(9)M(-1)s(-1)] and by UV photolysis with low and medium pressure mercury vapor lamps [quantum yields ?0 and (2.7 ± 0.4)× 10(-4) respectively] determined. All constants were measured at pH 6 and at temperature ?20°C. The results indicate that lamotrigine is slow to respond to direct photolysis or oxidation by ozone and no attenuation of the contaminant is expected in UV or ozone disinfection applications. The compound reacts rapidly with hydroxyl radicals indicating that advanced oxidation processes would be effective for its treatment. Degradation products were identified under each treatment process using accurate mass time-of-flight spectrometry and pathways of decay were proposed. The main transformation pathways in each process were: dechlorination of the benzene ring during direct photolysis; hydroxyl group addition to the benzene ring during the reaction with hydroxyl radicals; and triazine ring opening after reaction with ozone. Different products that form in each process may be to a varying degree less environmentally stable than the parent lamotrigine. In addition, a novel method of ozone quenching without addition of salts is presented. The new quenching method would allow subsequent mass spectrometry analysis without a solid phase extraction clean-up step. The method involves raising the pH of the sample to approximately 10 for a few seconds and lowering it back and is therefore limited to applications for which temporary pH change is not expected to affect the outcome of the analysis. PMID:25150682

Keen, Olya S; Ferrer, Imma; Michael Thurman, E; Linden, Karl G

2014-12-01

313

Structural basis of lentiviral subversion of a cellular protein degradation pathway  

PubMed Central

Lentiviruses contain accessory genes that have evolved to counteract the effects of host cellular defence proteins that inhibit productive infection. One such restriction factor, SAMHD1, inhibits HIV-1 infection of myeloid-lineage cells 1,2 as well as resting CD4+ T cells 3,4 by reducing the cellular dNTP concentration to a level where the viral reverse transcriptase cannot function 5,6. In other lentiviruses, including HIV-2 and related SIVs, SAMHD1 restriction is overcome by the action of viral accessory protein x (Vpx) or the related viral protein r (Vpr) that target and recruit SAMHD1 for proteasomal degradation 7,8. The molecular mechanism by which these viral proteins are able to usurp the host cell’s ubiquitination machinery to destroy the cell’s protection against these viruses has not been defined. We present here the crystal structure of a ternary complex of Vpx with the host cell’s E3 ligase substrate adaptor DCAF1 and the C-terminal region of SAMHD1. Vpx is made up of a three-helical bundle, stabilised by a zinc finger motif and wraps tightly around the disc-shaped DCAF1 molecule to present a new molecular surface. This adapted surface is then able to recruit SAMHD1 via its C-terminus making it a competent substrate for the E3 ligase to mark for proteasomal degradation. The structure provides the first description of how a lentiviral accessory protein is able to subvert the cell’s normal protein degradation pathway to inactivate the cellular viral defence system. PMID:24336198

Schwefel, David; Groom, Harriet C. T.; Boucherit, Virginie C.; Christodoulou, Evangelos; Walker, Philip A.; Stoye, Jonathan P.; Bishop, Kate N.; Taylor, Ian A.

2013-01-01

314

Structural basis of lentiviral subversion of a cellular protein degradation pathway  

NASA Astrophysics Data System (ADS)

Lentiviruses contain accessory genes that have evolved to counteract the effects of host cellular defence proteins that inhibit productive infection. One such restriction factor, SAMHD1, inhibits human immunodeficiency virus (HIV)-1 infection of myeloid-lineage cells as well as resting CD4+ T cells by reducing the cellular deoxynucleoside 5'-triphosphate (dNTP) concentration to a level at which the viral reverse transcriptase cannot function. In other lentiviruses, including HIV-2 and related simian immunodeficiency viruses (SIVs), SAMHD1 restriction is overcome by the action of viral accessory protein x (Vpx) or the related viral protein r (Vpr) that target and recruit SAMHD1 for proteasomal degradation. The molecular mechanism by which these viral proteins are able to usurp the host cell's ubiquitination machinery to destroy the cell's protection against these viruses has not been defined. Here we present the crystal structure of a ternary complex of Vpx with the human E3 ligase substrate adaptor DCAF1 and the carboxy-terminal region of human SAMHD1. Vpx is made up of a three-helical bundle stabilized by a zinc finger motif, and wraps tightly around the disc-shaped DCAF1 molecule to present a new molecular surface. This adapted surface is then able to recruit SAMHD1 via its C terminus, making it a competent substrate for the E3 ligase to mark for proteasomal degradation. The structure reported here provides a molecular description of how a lentiviral accessory protein is able to subvert the cell's normal protein degradation pathway to inactivate the cellular viral defence system.

Schwefel, David; Groom, Harriet C. T.; Boucherit, Virginie C.; Christodoulou, Evangelos; Walker, Philip A.; Stoye, Jonathan P.; Bishop, Kate N.; Taylor, Ian A.

2014-01-01

315

Genetic investigation of the catabolic pathway for degradation of abietane diterpenoids by Pseudomonas abietaniphila BKME-9.  

PubMed

We have cloned and sequenced the dit gene cluster encoding enzymes of the catabolic pathway for abietane diterpenoid degradation by Pseudomonas abietaniphila BKME-9. The dit gene cluster is located on a 16.7-kb DNA fragment containing 13 complete open reading frames (ORFs) and 1 partial ORF. The genes ditA1A2A3 encode the alpha and beta subunits and the ferredoxin of the dioxygenase which hydroxylates 7-oxodehydroabietic acid to 7-oxo-11,12-dihydroxy-8, 13-abietadien acid. The dioxygenase mutant strain BKME-941 (ditA1::Tn5) did not grow on nonaromatic abietanes, and transformed palustric and abietic acids to 7-oxodehydroabietic acid in cell suspension assays. Thus, nonaromatic abietanes are aromatized prior to further degradation. Catechol 2,3-dioxygenase activity of xylE transcriptional fusion strains showed induction of ditA1 and ditA3 by abietic, dehydroabietic, and 7-oxodehydroabietic acids, which support the growth of strain BKME-9, as well as by isopimaric and 12, 14-dichlorodehydroabietic acids, which are diterpenoids that do not support the growth of strain BKME-9. In addition to the aromatic-ring-hydroxylating dioxygenase genes, the dit cluster includes ditC, encoding an extradiol ring cleavage dioxygenase, and ditR, encoding an IclR-type transcriptional regulator. Although ditR is not strictly required for the growth of strain BKME-9 on abietanes, a ditR::Km(r) mutation in a ditA3::xylE reporter strain demonstrated that it encodes an inducer-dependent transcriptional activator of ditA3. An ORF with sequence similarity to genes encoding permeases (ditE) is linked with genes involved in abietane degradation. PMID:10850995

Martin, V J; Mohn, W W

2000-07-01

316

Reactions Involved in the Lower Pathway for Degradation of 4-Nitrotoluene by Mycobacterium Strain HL 4-NT-1  

PubMed Central

In spite of the variety of initial reactions, the aerobic biodegradation of aromatic compounds generally yields dihydroxy intermediates for ring cleavage. Recent investigation of the degradation of nitroaromatic compounds revealed that some nitroaromatic compounds are initially converted to 2-aminophenol rather than dihydroxy intermediates by a number of microorganisms. The complete pathway for the metabolism of 2-aminophenol during the degradation of nitrobenzene by Pseudomonas pseudoalcaligenes JS45 has been elucidated previously. The pathway is parallel to the catechol extradiol ring cleavage pathway, except that 2-aminophenol is the ring cleavage substrate. Here we report the elucidation of the pathway of 2-amino-4-methylphenol (6-amino-m-cresol) metabolism during the degradation of 4-nitrotoluene by Mycobacterium strain HL 4-NT-1 and the comparison of the substrate specificities of the relevant enzymes in strains JS45 and HL 4-NT-1. The results indicate that the 2-aminophenol ring cleavage pathway in strain JS45 is not unique but is representative of the pathways of metabolism of other o-aminophenolic compounds. PMID:10877799

He, Zhongqi; Spain, Jim C.

2000-01-01

317

Autophagic degradation contributes to muscle wasting in cancer cachexia.  

PubMed

Muscle protein wasting in cancer cachexia is a critical problem. The underlying mechanisms are still unclear, although the ubiquitin-proteasome system has been involved in the degradation of bulk myofibrillar proteins. The present work has been aimed to investigate whether autophagic degradation also plays a role in the onset of muscle depletion in cancer-bearing animals and in glucocorticoid-induced atrophy and sarcopenia of aging. The results show that autophagy is induced in muscle in three different models of cancer cachexia and in glucocorticoid-treated mice. In contrast, autophagic degradation in the muscle of sarcopenic animals is impaired but can be reactivated by calorie restriction. These results further demonstrate that different mechanisms are involved in pathologic muscle wasting and that autophagy, either excessive or defective, contributes to the complicated network that leads to muscle atrophy. In this regard, particularly intriguing is the observation that in cancer hosts and tumor necrosis factor ?-treated C2C12 myotubes, insulin can only partially blunt autophagy induction. This finding suggests that autophagy is triggered through mechanisms that cannot be circumvented by using classic upstream modulators, prompting us to identify more effective approaches to target this proteolytic system. PMID:23395093

Penna, Fabio; Costamagna, Domiziana; Pin, Fabrizio; Camperi, Andrea; Fanzani, Alessandro; Chiarpotto, Elena M; Cavallini, Gabriella; Bonelli, Gabriella; Baccino, Francesco M; Costelli, Paola

2013-04-01

318

Protacs for Treatment of Cancer  

PubMed Central

Protein degradation is the cell’s mechanism of eliminating misfolded or unwanted proteins. The pathway by which proteins are degraded occurs through the ubiquitin-proteasome system. Ubiquitin is a small 9-kilodalton (kDa) protein that is attached to proteins. A minimum of four ubiquitins is required for proteins to be recognized by the degradation machinery, known as the 26S proteasome. Defects in ubiquitination have been identified in a number of diseases, including cancer, neurodegenerative diseases, and metabolic disorders. We sought to exploit the delicate balance between protein synthesis and degradation to treat cancer by designing a chimeric molecule, known as Protac (Proteolysis Targeting Chimeric molecule). Protacs are heterobifunctional nanomolecules that are approximately 10 nanometers (nm) in size and can recruit proteins that cause cancer to the ubiquitin-proteasome machinery for degradation. In this review, we discuss the development of this novel technology for the treatment of cancer. PMID:20075761

Sakamoto, Kathleen M.

2010-01-01

319

The Whole Genome Sequence of Sphingobium chlorophenolicum L-1: Insights into the Evolution of the Pentachlorophenol Degradation Pathway  

SciTech Connect

Sphingobium chlorophenolicum Strain L-1 can mineralize the toxic pesticide pentachlorophenol (PCP). We have sequenced the genome of S. chlorophenolicum Strain L-1. The genome consists of a primary chromosome that encodes most of the genes for core processes, a secondary chromosome that encodes primarily genes that appear to be involved in environmental adaptation, and a small plasmid. The genes responsible for degradation of PCP are found on chromosome 2. We have compared the genomes of S. chlorophenolicum Strain L-1 and Sphingobium japonicum, a closely related Sphingomonad that degrades lindane. Our analysis suggests that the genes encoding the first three enzymes in the PCP degradation pathway were acquired via two different horizontal gene transfer events, and the genes encoding the final two enzymes in the pathway were acquired from the most recent common ancestor of these two bacteria.

Copley, Shelley D. [University of Colorado; Rokicki, Joseph [University of Colorado; Turner, Pernilla [University of Colorado; Daligault, Hajnalka E. [Los Alamos National Laboratory (LANL); Nolan, Matt [U.S. Department of Energy, Joint Genome Institute; Land, Miriam L [ORNL

2012-01-01

320

GADD45b mediates p53 protein degradation via Src/PP2A/MDM2 pathway upon arsenite treatment  

E-print Network

OPEN GADD45b mediates p53 protein degradation via Src/PP2A/MDM2 pathway upon arsenite treatment Y protein degradation via targeting Src/protein phosphatase 2A (PP2A)/murine double minute 2 (MDM2) pathway) activation. We demonstrated here that GADD45b mediated its anti-apoptotic effect via promoting p53 protein

Cai, Long

321

Reaction pathway of the degradation of the p-hydroxybenzoic acid by sulfate radical generated by ionizing radiations  

NASA Astrophysics Data System (ADS)

The degradation of p-hydroxybenzoic acid (HBA) in aqueous solutions by ionizing radiation was studied. The phenolic pollutant was easily removed by the electron beam irradiation, as more than 80% of the initial 100 ?M introduced was degraded for a dose of 600 Gy. It was shown that the addition of persulfate, producing the sulfate radical as additional reactive species, induced a change in the reaction pathway. LC-MS analyses were performed in order to identify the different by-products formed. In the absence of persulfate, the main by-product formed was 3,4-dihydroxybenzoic acid, while in presence of persulfate, 1,4-benzoquinone was detected and the hydroxylated by-products were not present. A reaction pathway of HBA degradation by hydroxyl and sulfate radicals was proposed from the identification of the chemical structure of the different by-products detected. The influences of pH and dissolved oxygen were also studied. A high decline of HBA degradation was observed at pH 11 compared to pH 4.5, this decrease was minimized in the presence of persulfate. The dissolved oxygen concentration was found to be a limiting parameter of HBA degradation, however an excess of dissolved oxygen in solution did not improve the degradation to a large extent.

Criquet, Justine; Leitner, Nathalie Karpel Vel

2015-01-01

322

Connecting lignin-degradation pathway with pre-treatment inhibitor sensitivity of Cupriavidus necator.  

PubMed

To produce lignocellulosic biofuels economically, the complete release of monomers from the plant cell wall components, cellulose, hemicellulose, and lignin, through pre-treatment and hydrolysis (both enzymatic and chemical), and the efficient utilization of these monomers as carbon sources, is crucial. In addition, the identification and development of robust microbial biofuel production strains that can tolerate the toxic compounds generated during pre-treatment and hydrolysis is also essential. In this work, Cupriavidus necator was selected due to its capabilities for utilizing lignin monomers and producing polyhydroxylbutyrate (PHB), a bioplastic as well as an advanced biofuel intermediate. We characterized the growth kinetics of C. necator in pre-treated corn stover slurry as well as individually in the pre-sence of 11 potentially toxic compounds in the saccharified slurry. We found that C. necator was sensitive to the saccharified slurry produced from dilute acid pre-treated corn stover. Five out of 11 compounds within the slurry were characterized as toxic to C. necator, namely ammonium acetate, furfural, hydroxymethylfurfural (HMF), benzoic acid, and p-coumaric acid. Aldehydes (e.g., furfural and HMF) were more toxic than the acetate and the lignin degradation products benzoic acid and p-coumaric acid; furfural was identified as the most toxic compound. Although toxic to C. necator at high concentration, ammonium acetate, benzoic acid, and p-coumaric acid could be utilized by C. necator with a stimulating effect on C. necator growth. Consequently, the lignin degradation pathway of C. necator was reconstructed based on genomic information and literature. The efficient conversion of intermediate catechol to downstream products of cis,cis-muconate or 2-hydroxymuconate-6-semialdehyde may help improve the robustness of C. necator to benzoic acid and p-coumaric acid as well as improve PHB productivity. PMID:24904560

Wang, Wei; Yang, Shihui; Hunsinger, Glendon B; Pienkos, Philip T; Johnson, David K

2014-01-01

323

Connecting lignin-degradation pathway with pre-treatment inhibitor sensitivity of Cupriavidus necator  

PubMed Central

To produce lignocellulosic biofuels economically, the complete release of monomers from the plant cell wall components, cellulose, hemicellulose, and lignin, through pre-treatment and hydrolysis (both enzymatic and chemical), and the efficient utilization of these monomers as carbon sources, is crucial. In addition, the identification and development of robust microbial biofuel production strains that can tolerate the toxic compounds generated during pre-treatment and hydrolysis is also essential. In this work, Cupriavidus necator was selected due to its capabilities for utilizing lignin monomers and producing polyhydroxylbutyrate (PHB), a bioplastic as well as an advanced biofuel intermediate. We characterized the growth kinetics of C. necator in pre-treated corn stover slurry as well as individually in the pre-sence of 11 potentially toxic compounds in the saccharified slurry. We found that C. necator was sensitive to the saccharified slurry produced from dilute acid pre-treated corn stover. Five out of 11 compounds within the slurry were characterized as toxic to C. necator, namely ammonium acetate, furfural, hydroxymethylfurfural (HMF), benzoic acid, and p-coumaric acid. Aldehydes (e.g., furfural and HMF) were more toxic than the acetate and the lignin degradation products benzoic acid and p-coumaric acid; furfural was identified as the most toxic compound. Although toxic to C. necator at high concentration, ammonium acetate, benzoic acid, and p-coumaric acid could be utilized by C. necator with a stimulating effect on C. necator growth. Consequently, the lignin degradation pathway of C. necator was reconstructed based on genomic information and literature. The efficient conversion of intermediate catechol to downstream products of cis,cis-muconate or 2-hydroxymuconate-6-semialdehyde may help improve the robustness of C. necator to benzoic acid and p-coumaric acid as well as improve PHB productivity. PMID:24904560

Wang, Wei; Yang, Shihui; Hunsinger, Glendon B.; Pienkos, Philip T.; Johnson, David K.

2014-01-01

324

Role of the Endoplasmic Reticulum-associated Degradation (ERAD) Pathway in Degradation of Hepatitis C Virus Envelope Proteins and Production of Virus Particles*  

PubMed Central

Viral infections frequently cause endoplasmic reticulum (ER) stress in host cells leading to stimulation of the ER-associated degradation (ERAD) pathway, which subsequently targets unassembled glycoproteins for ubiquitylation and proteasomal degradation. However, the role of the ERAD pathway in the viral life cycle is poorly defined. In this paper, we demonstrate that hepatitis C virus (HCV) infection activates the ERAD pathway, which in turn controls the fate of viral glycoproteins and modulates virus production. ERAD proteins, such as EDEM1 and EDEM3, were found to increase ubiquitylation of HCV envelope proteins via direct physical interaction. Knocking down of EDEM1 and EDEM3 increased the half-life of HCV E2, as well as virus production, whereas exogenous expression of these proteins reduced the production of infectious virus particles. Further investigation revealed that only EDEM1 and EDEM3 bind with SEL1L, an ER membrane adaptor protein involved in translocation of ERAD substrates from the ER to the cytoplasm. When HCV-infected cells were treated with kifunensine, a potent inhibitor of the ERAD pathway, the half-life of HCV E2 increased and so did virus production. Kifunensine inhibited the binding of EDEM1 and EDEM3 with SEL1L, thus blocking the ubiquitylation of HCV E2 protein. Chemical inhibition of the ERAD pathway neither affected production of the Japanese encephalitis virus (JEV) nor stability of the JEV envelope protein. A co-immunoprecipitation assay showed that EDEM orthologs do not bind with JEV envelope protein. These findings highlight the crucial role of the ERAD pathway in the life cycle of specific viruses. PMID:21878646

Saeed, Mohsan; Suzuki, Ryosuke; Watanabe, Noriyuki; Masaki, Takahiro; Tomonaga, Mitsunori; Muhammad, Amir; Kato, Takanobu; Matsuura, Yoshiharu; Watanabe, Haruo; Wakita, Takaji; Suzuki, Tetsuro

2011-01-01

325

Cytolethal Distending Toxins Require Components of the ER-Associated Degradation Pathway for Host Cell Entry  

PubMed Central

Intracellular acting protein exotoxins produced by bacteria and plants are important molecular determinants that drive numerous human diseases. A subset of these toxins, the cytolethal distending toxins (CDTs), are encoded by several Gram-negative pathogens and have been proposed to enhance virulence by allowing evasion of the immune system. CDTs are trafficked in a retrograde manner from the cell surface through the Golgi apparatus and into the endoplasmic reticulum (ER) before ultimately reaching the host cell nucleus. However, the mechanism by which CDTs exit the ER is not known. Here we show that three central components of the host ER associated degradation (ERAD) machinery, Derlin-2 (Derl2), the E3 ubiquitin-protein ligase Hrd1, and the AAA ATPase p97, are required for intoxication by some CDTs. Complementation of Derl2-deficient cells with Derl2:Derl1 chimeras identified two previously uncharacterized functional domains in Derl2, the N-terminal 88 amino acids and the second ER-luminal loop, as required for intoxication by the CDT encoded by Haemophilus ducreyi (Hd-CDT). In contrast, two motifs required for Derlin-dependent retrotranslocation of ERAD substrates, a conserved WR motif and an SHP box that mediates interaction with the AAA ATPase p97, were found to be dispensable for Hd-CDT intoxication. Interestingly, this previously undescribed mechanism is shared with the plant toxin ricin. These data reveal a requirement for multiple components of the ERAD pathway for CDT intoxication and provide insight into a Derl2-dependent pathway exploited by retrograde trafficking toxins. PMID:25078082

Eshraghi, Aria; Dixon, Shandee D.; Tamilselvam, Batcha; Kim, Emily Jin-Kyung; Gargi, Amandeep; Kulik, Julia C.; Damoiseaux, Robert; Blanke, Steven R.; Bradley, Kenneth A.

2014-01-01

326

Cytolethal distending toxins require components of the ER-associated degradation pathway for host cell entry.  

PubMed

Intracellular acting protein exotoxins produced by bacteria and plants are important molecular determinants that drive numerous human diseases. A subset of these toxins, the cytolethal distending toxins (CDTs), are encoded by several Gram-negative pathogens and have been proposed to enhance virulence by allowing evasion of the immune system. CDTs are trafficked in a retrograde manner from the cell surface through the Golgi apparatus and into the endoplasmic reticulum (ER) before ultimately reaching the host cell nucleus. However, the mechanism by which CDTs exit the ER is not known. Here we show that three central components of the host ER associated degradation (ERAD) machinery, Derlin-2 (Derl2), the E3 ubiquitin-protein ligase Hrd1, and the AAA ATPase p97, are required for intoxication by some CDTs. Complementation of Derl2-deficient cells with Derl2:Derl1 chimeras identified two previously uncharacterized functional domains in Derl2, the N-terminal 88 amino acids and the second ER-luminal loop, as required for intoxication by the CDT encoded by Haemophilus ducreyi (Hd-CDT). In contrast, two motifs required for Derlin-dependent retrotranslocation of ERAD substrates, a conserved WR motif and an SHP box that mediates interaction with the AAA ATPase p97, were found to be dispensable for Hd-CDT intoxication. Interestingly, this previously undescribed mechanism is shared with the plant toxin ricin. These data reveal a requirement for multiple components of the ERAD pathway for CDT intoxication and provide insight into a Derl2-dependent pathway exploited by retrograde trafficking toxins. PMID:25078082

Eshraghi, Aria; Dixon, Shandee D; Tamilselvam, Batcha; Kim, Emily Jin-Kyung; Gargi, Amandeep; Kulik, Julia C; Damoiseaux, Robert; Blanke, Steven R; Bradley, Kenneth A

2014-07-01

327

Discovery of a novel L-lyxonate degradation pathway in Pseudomonas aeruginosa PAO1.  

PubMed

The l-lyxonate dehydratase (LyxD) in vitro enzymatic activity and in vivo metabolic function were assigned to members of an isofunctional family within the mandelate racemase (MR) subgroup of the enolase superfamily. This study combined in vitro and in vivo data to confirm that the dehydration of l-lyxonate is the biological role of the members of this family. In vitro kinetic experiments revealed catalytic efficiencies of ?10(4) M(-1) s(-1) as previously observed for members of other families in the MR subgroup. Growth studies revealed that l-lyxonate is a carbon source for Pseudomonas aeruginosa PAO1; transcriptomics using qRT-PCR established that the gene encoding LyxD as well as several other conserved proximal genes were upregulated in cells grown on l-lyxonate. The proximal genes were shown to be involved in a pathway for the degradation of l-lyxonate, in which the first step is dehydration by LyxD followed by dehydration of the 2-keto-3-deoxy-l-lyxonate product by 2-keto-3-deoxy-l-lyxonate dehydratase to yield ?-ketoglutarate semialdehyde. In the final step, ?-ketoglutarate semialdehyde is oxidized by a dehydrogenase to ?-ketoglutarate, an intermediate in the citric acid cycle. An X-ray structure for the LyxD from Labrenzia aggregata IAM 12614 with Mg(2+) in the active site was determined that confirmed the expectation based on sequence alignments that LyxDs possess a conserved catalytic His-Asp dyad at the end of seventh and sixth ?-strands of the (?/?)7?-barrel domain as well as a conserved KxR motif at the end of second ?-strand; substitutions for His 316 or Arg 179 inactivated the enzyme. This is the first example of both the LyxD function in the enolase superfamily and a pathway for the catabolism of l-lyxonate. PMID:24831290

Ghasempur, Salehe; Eswaramoorthy, Subramaniam; Hillerich, Brandan S; Seidel, Ronald D; Swaminathan, Subramanyam; Almo, Steven C; Gerlt, John A

2014-05-27

328

2-Hydroxycyclohexanecarboxyl Coenzyme A Dehydrogenase, an Enzyme Characteristic of the Anaerobic Benzoate Degradation Pathway Used by Rhodopseudomonas palustris  

PubMed Central

A gene, badH, whose predicted product is a member of the short-chain dehydrogenase/reductase family of enzymes, was recently discovered during studies of anaerobic benzoate degradation by the photoheterotrophic bacterium Rhodopseudomonas palustris. Purified histidine-tagged BadH protein catalyzed the oxidation of 2-hydroxycyclohexanecarboxyl coenzyme A (2-hydroxychc-CoA) to 2-ketocyclohexanecarboxyl-CoA. These compounds are proposed intermediates of a series of three reactions that are shared by the pathways of cyclohexanecarboxylate and benzoate degradation used by R. palustris. The 2-hydroxychc-CoA dehydrogenase activity encoded by badH was dependent on the presence of NAD+; no activity was detected with NADP+ as a cofactor. The dehydrogenase activity was not sensitive to oxygen. The enzyme has apparent Km values of 10 and 200 ?M for 2-hydroxychc-CoA and NAD+, respectively. Western blot analysis with antisera raised against purified His-BadH identified a 27-kDa protein that was present in benzoate- and cyclohexanecarboxylate-grown but not in succinate-grown R. palustris cell extracts. The active form of the enzyme is a homotetramer. badH was determined to be the first gene in an operon, termed the cyclohexanecarboxylate degradation operon, containing genes required for both benzoate and cyclohexanecarboxylate degradation. A nonpolar R. palustris badH mutant was unable to grow on benzoate or cyclohexanecarboxylate but had wild-type growth rates on succinate. Cells blocked in expression of the entire cyclohexanecarboxylate degradation operon excreted cyclohex-1-ene-1-carboxylate into the growth medium when given benzoate. This confirms that cyclohex-1-ene-1-carboxyl-CoA is an intermediate of anaerobic benzoate degradation by R. palustris. This compound had previously been shown not to be formed by Thauera aromatica, a denitrifying bacterium that degrades benzoate by a pathway that is slightly different from the R. palustris pathway. 2-Hydroxychc-CoA dehydrogenase does not participate in anaerobic benzoate degradation by T. aromatica and thus may serve as a useful indicator of an R. palustris-type benzoate degradation pathway. PMID:10781543

Pelletier, Dale A.; Harwood, Caroline S.

2000-01-01

329

Ozonation degradation of microcystin-LR in aqueous solution: intermediates, byproducts and pathways.  

PubMed

The intermediates and byproducts formed during the ozonation of microcystin-LR (MC-LR, m/z = 995.5) and the probable degradation pathway were investigated at different initial molar ratios of ozone to MC-LR ([O3]0/[MC-LR]0). Seven reaction intermediates with m/z ? 795.4 were observed by LC/MS, and four of them (m/z = 815.4, 827.3, 853.3 and 855.3) have not been previously reported. Meanwhile, six aldehyde-based byproducts with molecular weights of 30-160 were detected for the first time. Intermediates structures demonstrated that ozone reacted with two sites of MC-LR: the diene bonds in the Adda side chain and the Mdha amino acid in the cyclic structure. The fragment from the Adda side chain oxidative cleavage could be further oxidized to an aldehyde with a molecular weight of 160 at low [O3]0/[MC-LR]0. Meanwhile, the polypeptide structure of MC-LR was difficult to be further oxidized, unless [O3]0/[MC-LR]0 > 10. After further oxidation of the intermediates, five other aldehyde-based byproducts were detected by GC/MS: formaldehyde, acetaldehyde, isovaleraldehyde, glyoxal and methylglyoxal. Formaldehyde, isovaleraldehyde and methylglyoxal were the dominant species. The yields of the aldehydes varied greatly, depending on the value of [O3]0/[MC-LR]0. PMID:24981743

Chang, Jing; Chen, Zhong-lin; Wang, Zhe; Shen, Ji-min; Chen, Qian; Kang, Jing; Yang, Lei; Liu, Xiao-wei; Nie, Chang-xin

2014-10-15

330

A signalling pathway controlling c-Myc degradation that impacts oncogenic transformation of human cells.  

PubMed

The stability of c-Myc is regulated by multiple Ras effector pathways. Phosphorylation at Ser 62 stabilizes c-Myc, whereas subsequent phosphorylation at Thr 58 is required for its degradation. Here we show that Ser 62 is dephosphorylated by protein phosphatase 2A (PP2A) before ubiquitination of c-Myc, and that PP2A activity is regulated by the Pin1 prolyl isomerase. Furthermore, the absence of Pin1 or inhibition of PP2A stabilizes c-Myc. A stable c-Myc(T58A) mutant that cannot bind Pin1 or be dephosphorylated by PP2A replaces SV40 small T antigen in human cell transformation and tumorigenesis assays. Therefore, small T antigen, which inactivates PP2A, exerts its oncogenic potential by preventing dephosphorylation of c-Myc, resulting in c-Myc stabilization. Thus, Ras-dependent signalling cascades ensure transient and self-limiting accumulation of c-Myc, disruption of which contributes to human cell oncogenesis. PMID:15048125

Yeh, Elizabeth; Cunningham, Melissa; Arnold, Hugh; Chasse, Dawn; Monteith, Teresa; Ivaldi, Giovanni; Hahn, William C; Stukenberg, P Todd; Shenolikar, Shirish; Uchida, Takafumi; Counter, Christopher M; Nevins, Joseph R; Means, Anthony R; Sears, Rosalie

2004-04-01

331

The molecular components of the extracellular protein-degradation pathways of the ectomycorrhizal fungus Paxillus involutus  

PubMed Central

Proteins contribute to a major part of the organic nitrogen (N) in forest soils. This N is mobilized and becomes available to trees as a result of the depolymerizing activities of symbiotic ectomycorrhizal fungi. The mechanisms by which these fungi depolymerize proteins and assimilate the released N are poorly characterized. Biochemical analysis and transcriptome profiling were performed to examine the proteolytic machinery and the uptake system of the ectomycorrhizal basidiomycete Paxillus involutus during the assimilation of organic N from various protein sources and extracts of organic matter. All substrates induced secretion of peptidase activity with an acidic pH optimum, mostly contributed by aspartic peptidases. The peptidase activity was transiently repressed by ammonium. Transcriptional analysis revealed a large number of extracellular endo- and exopeptidases. The expression levels of these peptidases were regulated in parallel with transporters and enzymes involved in the assimilation and metabolism of the released peptides and amino acids. For the first time the molecular components of the protein degradation pathways of an ectomycorrhizal fungus are described. The data suggest that the transcripts encoding these components are regulated in response to the chemical properties and the availability of the protein substrates. PMID:23902518

Shah, Firoz; Rineau, Francois; Canbäck, Björn; Johansson, Tomas; Tunlid, Anders

2013-01-01

332

TRAF3 negatively regulates calcineurin-NFAT pathway by targeting calcineurin B subunit for degradation.  

PubMed

Calcineurin (CN) is the only serine/threonine specific protein phosphatase regulated by Ca(2+) /calmodulin (CaM), which is composed of catalytic A subunit (CNA) and regulatory B subunit (CNB). Tumor necrosis factor (TNF) receptor associated factor 3 (TRAF3) is an essential component in the Toll like receptors and TNF receptors (TNFRs) pathways. The TRAF domain of TRAF3 interacts with a large range of proteins, which share consensus sequences known as TRAF interacting motifs (TIMs). By sequence alignment, we identified two potential TIMs in CNB. However, the relation between TRAF3 and CN has not been reported before. To explore this, we highly expressed the former insoluble TRAF domain of TRAF3 in soluble form by using CaM fusion system for the first time. We demonstrated that the TRAF domain of TRAF3 interacted with CNB. On further investigation, over-expression of TRAF3 inhibited endogenous CN's activity, which decreased NFAT reporter activity and IL-2 production. Knock-down of TRAF3 partially enhanced CN's activity. The possible mechanism was that TRAF3 functioned as ubiquitin E3 ligase for CN and promoted its degradation. PMID:22715070

Wang, Xinyu; Huang, Yi; Li, Li; Wei, Qun

2012-09-01

333

Endocytosis and ligand dissociation and degradation mediated by the hepatic galactosyl receptor occur via two different pathways  

SciTech Connect

Isolated rat hepatocytes express two distinct populations of surface Galactosyl receptor activity, measured by the binding of /sup 125/I-asialo-orosomucoid (ASOR), which they designate State 1 and State 2. Freshly isolated cells express only state 1 receptors. Cells equilibrated at 37/sup 0/C also express State 2 receptors, which represent 50-80% of the total surface activity. In the absence of ligand, State 2 receptor activity is reversibly decreased by metabolic energy poisons, low temperature and microtubule drugs, whereas State 1 receptor activity is unaffected. Endocytosis of /sup 125/I-ASOR by State 1 receptors is followed by a slow dissociation of /sup 125/I-ASOR from receptor but the immediate release of acid soluble degradation products. In contrast, State 2 receptors mediate endocytosis which involves a rapid dissociation step but a 20 min lag, prior to the release of degradation products. Both pathways follow first order kinetics and are functional under steady state conditions indicating coordinated receptor recycling. Degradation mediated by both pathways is inhibited by leupeptin and chloroquine. The State 1 and 2 pathways can be further differentiated by the greater sensitivity of the latter to microtubule drugs. These results suggest that there are either structurally different native receptors or that identical receptors are directed into two different functional pathways, for example by interaction with different types of coated pits.

Weigel, P.H.; Clarke, B.L.; Oka, J.A.

1986-05-01

334

Analysis of Hydroxycinnamic Acid Degradation in Agrobacterium fabrum Reveals a Coenzyme A-Dependent, Beta-Oxidative Deacetylation Pathway  

PubMed Central

The soil- and rhizosphere-inhabiting bacterium Agrobacterium fabrum (genomospecies G8 of the Agrobacterium tumefaciens species complex) is known to have species-specific genes involved in ferulic acid degradation. Here, we characterized, by genetic and analytical means, intermediates of degradation as feruloyl coenzyme A (feruloyl-CoA), 4-hydroxy-3-methoxyphenyl-?-hydroxypropionyl–CoA, 4-hydroxy-3-methoxyphenyl-?-ketopropionyl–CoA, vanillic acid, and protocatechuic acid. The genes atu1416, atu1417, and atu1420 have been experimentally shown to be necessary for the degradation of ferulic acid. Moreover, the genes atu1415 and atu1421 have been experimentally demonstrated to be essential for this degradation and are proposed to encode a phenylhydroxypropionyl-CoA dehydrogenase and a 4-hydroxy-3-methoxyphenyl-?-ketopropionic acid (HMPKP)–CoA ?-keto-thiolase, respectively. We thus demonstrated that the A. fabrum hydroxycinnamic degradation pathway is an original coenzyme A-dependent ?-oxidative deacetylation that could also transform p-coumaric and caffeic acids. Finally, we showed that this pathway enables the metabolism of toxic compounds from plants and their use for growth, likely providing the species an ecological advantage in hydroxycinnamic-rich environments, such as plant roots or decaying plant materials. PMID:24657856

Campillo, Tony; Renoud, Sébastien; Kerzaon, Isabelle; Vial, Ludovic; Baude, Jessica; Gaillard, Vincent; Bellvert, Floriant; Chamignon, Cécile; Comte, Gilles; Lavire, Céline; Hommais, Florence

2014-01-01

335

Characterization of a new catechol branch of the ?-ketoadipate pathway induced for benzoate degradation in Acinetobacter lwoffii K24  

Microsoft Academic Search

Acinetobacter lwoffii K24 is a known aniline-degrading bacterium. In previous studies, two catechol branches of the ?-ketoadipate pathway were reported to be induced for aniline degradation, and related enzymes (CatA1 and CatA2) were identified from the aniline-induced proteome of A. lwoffii K24 [S.I. Kim, S.H. Leem, J.S. Choi, Y.H. Chung, S. Kim, Y.M. Park, Y.K. Park, Y.N. Lee, K.S. Ha,

Young-Ho Yoon; Sung-Ho Yun; Soon-Ho Park; So-Young Seol; Sun-Hee Leem; Seung Il Kim

2007-01-01

336

Listeriolysin O Secreted by Listeria monocytogenes into the Host Cell Cytosol Is Degraded by the N-End Rule Pathway  

Microsoft Academic Search

The intracellular pathogen Listeria monocytogenes escapes from a phagosomal compartment into the cytosol by secreting the pore-forming cytolysin listeriolysin O (LLO). During the proliferation of L. monocytogenes bacteria in the mammalian cell cytosol, the secreted LLO is targeted for degradation by the ubiquitin system. We report here that LLO is a substrate of the ubiquitin-dependent N-end rule pathway, which recognizes

Pamela Schnupf; Jianmin Zhou; Alexander Varshavsky; Daniel A. Portnoy

2007-01-01

337

Mutation of the E6AP Ubiquitin Ligase Reduces Nuclear Inclusion Frequency While Accelerating Polyglutamine-Induced Pathology in SCA1 Mice  

Microsoft Academic Search

Mutant ataxin-1, the expanded polyglutamine protein causing spinocerebellar ataxia type 1 (SCA1), aggregates in ubiquitin-positive nuclear inclusions (NI) that alter proteasome distribution in affected SCA1 patient neurons. Here, we observed that ataxin-1 is degraded by the ubiquitin-proteasome pathway. While ataxin-1 [2Q] and mutant ataxin-1 [92Q] are polyubiquitinated equally well in vitro, the mutant form is three times more resistant to

Christopher J. Cummings; Eyal Reinstein; Yaling Sun; Barbara Antalffy; Yong-hui Jiang; Aaron Ciechanover; Harry T. Orr; Arthur L. Beaudet; Huda Y Zoghbi

1999-01-01

338

Mechanism of induction of muscle protein loss by hyperglycaemia  

Microsoft Academic Search

Treatment of murine myotubes with high glucose concentrations (10 and 25 mM) stimulated protein degradation through the ubiquitin–proteasome pathway, and also caused activation (autophosphorylation) of PKR (double-stranded-RNA-dependent protein kinase) and eIF2? (eukaryotic initiation factor 2?). Phosphorylation of PKR and eIF2? was also seen in the gastrocnemius muscle of diabetic ob\\/ob mice. High glucose levels also inhibited protein synthesis. The effect of

Steven T. Russell; Seema Rajani; Raj S. Dhadda; Michael J. Tisdale

2009-01-01

339

Aging perturbs 26S proteasome assembly in Drosophila melanogaster  

Microsoft Academic Search

Aging is associated with loss of quality control in protein turnover. The ubiquitin-proteasome pathway is critical to this quality control process as it degrades mutated and damaged proteins. We identified a unique aging-dependent mechanism that contributes to proteasome dysfunction in Drosophila melanogaster. Our studies are the first to show that the major protea- some form in old (43-47 days old)

Vita A. Vernace; Lisette Arnaud; Thomas Schmidt-Glenewinkel; Maria E. Figueiredo-Pereira

2007-01-01

340

Proteasome inhibitors: from research tools to drug candidates.  

PubMed

The 26S proteasome is a 2.4 MDa multifunctional ATP-dependent proteolytic complex, which degrades the majority of cellular polypeptides by an unusual enzyme mechanism. Several groups of proteasome inhibitors have been developed and are now widely used as research tools to study the role of the ubiquitin-proteasome pathway in various cellular processes, and two inhibitors are now in clinical trials for treatment of multiple cancers and stroke. PMID:11514224

Kisselev, A F; Goldberg, A L

2001-08-01

341

Inhibition of autophagy induced by proteasome inhibition increases cell death in human SHG-44 glioma cells  

Microsoft Academic Search

Aim:The ubiquitin-proteasome system (UPS) and lysosome-dependent macroautophagy (autophagy) are two major intracellular pathways for protein degradation. Recent studies suggest that proteasome inhibitors may reduce tumor growth and activate autophagy. Due to the dual roles of autophagy in tumor cell survival and death, the effect of autophagy on the destiny of glioma cells remains unclear. In this study, we sought to

Peng-fei Ge; Ji-zhou Zhang; Xiao-fei Wang; Fan-kai Meng; Wen-chen Li; Yong-xin Luan; Feng Ling; Yi-nan Luo

2009-01-01

342

Blocking Autophagy Prevents Bortezomib-Induced NF-?B Activation by Reducing I-?B? Degradation in Lymphoma Cells  

PubMed Central

Here we show that bortezomib induces effective proteasome inhibition and accumulation of poly-ubiquitinated proteins in diffuse large B-cell lymphoma (DLBCL) cells. This leads to induction of endoplasmic reticulum (ER) stress as demonstrated by accumulation of the protein CHOP, as well as autophagy, as demonstrated by accumulation of LC3-II proteins. Our data suggest that recruitment of both ubiquitinated proteins and LC3-II by p62 directs ubiquitinated proteins, including I-?B?, to the autophagosome. Degradation of I-?B? results in increased NF-?B nuclear translocation and transcription activity. Since bortezomib treatment promoted I-?B? phosphorylation, ubiquitination and degradation, this suggests that the route of I-?B? degradation was not via the ubiquitin-proteasome degradation system. The autophagy inhibitor chloroquine (CQ) significantly inhibited bortezomib-induced I-?B? degradation, increased complex formation with NF-?B and reduced NF-?B nuclear translocation and DNA binding activity. Importantly, the combination of proteasome and autophagy inhibitors showed synergy in killing DLBCL cells. In summary, bortezomib-induced autophagy confers relative DLBCL cell drug resistance by eliminating I-?B?. Inhibition of both autophagy and the proteasome has great potential to kill apoptosis-resistant lymphoma cells. PMID:22393418

Jia, Li; Gopinathan, Ganga; Sukumar, Johanna T.; Gribben, John G.

2012-01-01

343

Mitochondrial dismissal in mammals, from protein degradation to mitophagy.  

PubMed

Mitochondria are double-membraned highly dynamic organelles; the shape, location and function of which are determined by a constant balance between opposing fusion and fission events. A fine modulation of mitochondrial structure is crucial for their correct functionality and for many physiological cell processes, the status of these organelles, being thus a key aspect in a cell's fate. Indeed, the homeostasis of mitochondria needs to be highly regulated for the above mentioned reasons, and since a) they are the major source of energy; b) they participate in various signaling pathways; albeit at the same time c) they are also the major source of reactive oxygen species (ROS, the main damaging detrimental players for all cell components). Elaborate mechanisms of mitochondrial quality control have evolved for maintaining a functional mitochondrial network and avoiding cell damage. The first mechanism is the removal of damaged mitochondrial proteins within the organelle via chaperones and protease; the second is the cytosolic ubiquitin-proteasome system (UPS), able to eliminate proteins embedded in the outer mitochondrial membrane; the third is the removal of the entire mitochondria through mitophagy, in the case of extensive organelle damage and dysfunction. In this review, we provide an overview of these mitochondria stability and quality control mechanisms, highlighting mitophagy, and emphasizing the central role of mitochondrial dynamics in this context. This article is part of a Special Issue entitled: Dynamic and ultrastructure of bioenergetic membranes and their components. PMID:24275087

Campello, Silvia; Strappazzon, Flavie; Cecconi, Francesco

2014-04-01

344

Study of Biochemical Pathways and Enzymes Involved in Pyrene Degradation by Mycobacterium sp. Strain KMS  

Technology Transfer Automated Retrieval System (TEKTRAN)

Pyrene degradation is known in bacteria. In this study, Mycobacterium sp. Strain KMS was used to study the metabolites produced during, and enzymes involved in, pyrene degradation. Several key metabolites, including pyrene-4,5-dione, cis-4,5-pyrene-dihydrodiol, phenanthrene-4,5-dicarboxylic acid, ...

345

HUWE1 ubiquitinates MyoD and targets it for proteasomal degradation  

SciTech Connect

Highlights: Black-Right-Pointing-Pointer HUWE1 ubiquitinates MyoD in vitro and in cells. Black-Right-Pointing-Pointer The ubiquitination by HUWE1 targets MyoD for proteasomal degradation. Black-Right-Pointing-Pointer HUWE1 can modify MyoD on its N-terminal residue. -- Abstract: MyoD is a tissue-specific transcriptional activator that acts as a master switch for muscle development. It activates a broad array of muscle-specific genes, which leads to conversion of proliferating myoblasts into mature myotubes. The ubiquitin proteasome system (UPS) plays an important role in controlling MyoD. Both its N-terminal residue and internal lysines can be targeted by ubiquitin, and both modifications appear to direct it for proteasomal degradation. The protein is short-lived and has a half-life of {approx}45 min in different cells. It was reported that MyoD can be ubiquitinated by MAFbx/AT-1, but accumulating lines of experimental evidence showed that other ligase(s) may also participate in its targeting. Here we describe the involvement of HUWE1 in the ubiquitination and proteasomal degradation of MyoD. Furthermore, we show that the ligase can ubiquitinate the protein in its N-terminal residue.

Noy, Tahel; Suad, Oded [Cancer and Vascular Biology Research Center, The Rappaport Faculty of Medicine and Research Institute, Technion-Israel Institute of Technology, Haifa 31096 (Israel)] [Cancer and Vascular Biology Research Center, The Rappaport Faculty of Medicine and Research Institute, Technion-Israel Institute of Technology, Haifa 31096 (Israel); Taglicht, Daniel [Proteologics Ltd., Weizmann Science Park, Rehovot 76704 (Israel)] [Proteologics Ltd., Weizmann Science Park, Rehovot 76704 (Israel); Ciechanover, Aaron, E-mail: c_tzachy@netvision.net.il [Cancer and Vascular Biology Research Center, The Rappaport Faculty of Medicine and Research Institute, Technion-Israel Institute of Technology, Haifa 31096 (Israel)] [Cancer and Vascular Biology Research Center, The Rappaport Faculty of Medicine and Research Institute, Technion-Israel Institute of Technology, Haifa 31096 (Israel)

2012-02-10

346

Degradation pathway and kinetics of 1-alkyl-3-methylimidazolium bromides oxidation in an ultrasonic nanoscale zero-valent iron/hydrogen peroxide system.  

PubMed

Fenton and Fenton-like oxidation has been already demonstrated to be efficient for the degradation of imidazolium ionic liquids (ILs), but little is known for their degradation pathway and kinetics in such systems. In this work, degradation pathway and kinetics of 1-alkyl-3-methylimidazolium bromides ([Cnmim]Br, n=2, 4, 6, 8, and 10) were investigated in an ultrasound nanoscale zero-valent iron/hydrogen peroxide (US-nZVI/H2O2) system. For this purpose, 1-butyl-3-methylimidazolium bromide ([C4mim]Br) was used as a representative ionic liquid to optimize pH value, nZVI dose, and H2O2 concentration for the degradation reaction. Then, the degradation kinetics of [Cnmim]Br was investigated under optimal conditions, and their degradation intermediates were monitored by gas chromatography-mass spectrometry (GC-MS). It was shown that the degradation of [Cnmim]Br in such a heterogeneous Fenton-like system could be described by a second order kinetic model, and a number of intermediate products were detected. Based on these intermediate products, detailed pathways were proposed for the degradation of [Cnmim]Br in the ultrasound-assisted nZVI/H2O2 system. These findings may be useful for the better understanding of degradation mechanism of the imidazolium ILs in aqueous solutions. PMID:25463239

Zhou, Haimei; Shen, Yuanyuan; Lv, Ping; Wang, Jianji; Li, Pu

2015-03-01

347

Fragmentation pathways of polycyclic polyisoprenylated benzophenones and degradation profile of nemorosone by multiple-stage tandem mass spectrometry.  

PubMed

Nemorosone is a polycyclic polyisoprenylated benzophenone (PPBs) with strong cytotoxic activity. It is the major constituent of Clusia rosea floral resin and brown Cuban propolis. Other PPBs found in Cuban propolis are oxidized and cyclized derivatives of nemorosone. The instability of PPBs carrying an enolizable 1,3-diketone system has been suggested, and the elucidation of this aspect is very fundamental for the evaluation of their biologic activity. Electrospray ionization multistage tandem mass spectrometry (ESI-MS(n)) was employed to shed light on the origin of these derivatives of nemorosone and to define the stability of this natural product. For this purpose, we initially performed MS(n) experiments on seven related PPBs to obtain useful information for structural characterization of this class of compounds and to identify the degradation products of nemorosone. The proposed fragmentation pathways, supported by exact mass measurements, allowed the nature of side chains on the bicyclo core and the type and position of their modifications to be established. In a second part, the degradation profile of nemorosone under different conditions was investigated to assess the possible effects of isolation procedures, climatic, and storage conditions on its stability. Our results reveal that nemorosone undergoes rapid degradation in n-hexane and chloroform solutions. The degradation products, identified by HPLC-ESI/MS(n) and NMR, are identical to derivatives of nemorosone previously isolated from propolis and plants. Thus, these PPBs are artefacts formed predominantly during the extraction and purification procedures. PMID:19540134

Piccinelli, Anna Lisa; Campone, Luca; Dal Piaz, Fabrizio; Cuesta-Rubio, Osmany; Rastrelli, Luca

2009-09-01

348

Hemoglobin degradation in the human malaria pathogen Plasmodium falciparum: a catabolic pathway initiated by a specific aspartic protease  

PubMed Central

Hemoglobin is an important nutrient source for intraerythrocytic malaria organisms. Its catabolism occurs in an acidic digestive vacuole. Our previous studies suggested that an aspartic protease plays a key role in the degradative process. We have now isolated this enzyme and defined its role in the hemoglobinolytic pathway. Laser desorption mass spectrometry was used to analyze the proteolytic action of the purified protease. The enzyme has a remarkably stringent specificity towards native hemoglobin, making a single cleavage between alpha 33Phe and 34Leu. This scission is in the hemoglobin hinge region, unraveling the molecule and exposing other sites for proteolysis. The protease is inhibited by pepstatin and has NH2-terminal homology to mammalian aspartic proteases. Isolated digestive vacuoles make a pepstatin- inhibitable cleavage identical to that of the purified enzyme. The pivotal role of this aspartic hemoglobinase in initiating hemoglobin degradation in the malaria parasite digestive vacuoles is demonstrated. PMID:2007860

1991-01-01

349

Anaerobic Degradation of p-Ethylphenol by “Aromatoleum aromaticum” Strain EbN1: Pathway, Regulation, and Involved Proteins? †  

PubMed Central

The denitrifying “Aromatoleum aromaticum” strain EbN1 was demonstrated to utilize p-ethylphenol under anoxic conditions and was suggested to employ a degradation pathway which is reminiscent of known anaerobic ethylbenzene degradation in the same bacterium: initial hydroxylation of p-ethylphenol to 1-(4-hydroxyphenyl)-ethanol followed by dehydrogenation to p-hydroxyacetophenone. Possibly, subsequent carboxylation and thiolytic cleavage yield p-hydroxybenzoyl-coenzyme A (CoA), which is channeled into the central benzoyl-CoA pathway. Substrate-specific formation of three of the four proposed intermediates was confirmed by gas chromatographic-mass spectrometric analysis and also by applying deuterated p-ethylphenol. Proteins suggested to be involved in this degradation pathway are encoded in a single large operon-like structure (?15 kb). Among them are a p-cresol methylhydroxylase-like protein (PchCF), two predicted alcohol dehydrogenases (ChnA and EbA309), a biotin-dependent carboxylase (XccABC), and a thiolase (TioL). Proteomic analysis (two-dimensional difference gel electrophoresis) revealed their specific and coordinated upregulation in cells adapted to anaerobic growth with p-ethylphenol and p-hydroxyacetophenone (e.g., PchF up to 29-fold). Coregulated proteins of currently unknown function (e.g., EbA329) are possibly involved in p-ethylphenol- and p-hydroxyacetophenone-specific solvent stress responses and related to other aromatic solvent-induced proteins of strain EbN1. PMID:18539747

Wöhlbrand, Lars; Wilkes, Heinz; Halder, Thomas; Rabus, Ralf

2008-01-01

350

Novel pathway for the degradation of 2-chloro-4-nitrobenzoic acid by Acinetobacter sp. strain RKJ12.  

PubMed

The organism Acinetobacter sp. RKJ12 is capable of utilizing 2-chloro-4-nitrobenzoic acid (2C4NBA) as a sole source of carbon, nitrogen, and energy. In the degradation of 2C4NBA by strain RKJ12, various metabolites were isolated and identified by a combination of chromatographic, spectroscopic, and enzymatic activities, revealing a novel assimilation pathway involving both oxidative and reductive catabolic mechanisms. The metabolism of 2C4NBA was initiated by oxidative ortho dehalogenation, leading to the formation of 2-hydroxy-4-nitrobenzoic acid (2H4NBA), which subsequently was metabolized into 2,4-dihydroxybenzoic acid (2,4-DHBA) by a mono-oxygenase with the concomitant release of chloride and nitrite ions. Stoichiometric analysis indicated the consumption of 1 mol O(2) per conversion of 2C4NBA to 2,4-DHBA, ruling out the possibility of two oxidative reactions. Experiments with labeled H(2)(18)O and (18)O(2) indicated the involvement of mono-oxygenase-catalyzed initial hydrolytic dechlorination and oxidative denitration mechanisms. The further degradation of 2,4-DHBA then proceeds via reductive dehydroxylation involving the formation of salicylic acid. In the lower pathway, the organism transformed salicylic acid into catechol, which was mineralized by the ortho ring cleavage catechol-1,2-dioxygenase to cis, cis-muconic acid, ultimately forming tricarboxylic acid cycle intermediates. Furthermore, the studies carried out on a 2C4NBA(-) derivative and a 2C4NBA(+) transconjugant demonstrated that the catabolic genes for the 2C4NBA degradation pathway possibly reside on the ?55-kb transmissible plasmid present in RKJ12. PMID:21803909

Prakash, Dhan; Kumar, Ravi; Jain, R K; Tiwary, B N

2011-09-01

351

Isolation of a novel beta-cypermethrin degrading strain Bacillus subtilis BSF01 and its biodegradation pathway.  

PubMed

Continuous use of the pyrethroid insecticide beta-cypermethrin (beta-cp) has resulted in serious environmental contamination problems. We report here that a novel bacterial strain BSF01, which was isolated from activated sludge and identified as Bacillus subtilis (collection number: CCTCC AB 2014103), showed high efficiency in degrading beta-cp. Strain BSF01 was able to utilize beta-cp as the sole carbon source for growth and degraded 89.4 % of 50 mg L(-1) beta-cp within 7 days. The optimal conditions for beta-cp degradation were determined to be 34.5 °C, pH 6.7, and inocula amount 0.11 g dry wt L(-1) using response surface methodology. The kinetic parameters q max, K s, and K i were established to be 2.19 day(-1), 76.37 mg L(-1), and 54.14 mg L(-1), respectively. The critical inhibitor concentration was determined to be 64.30 mg L(-1). Seven metabolites were identified by gas chromatography-mass spectrometry. Furthermore, a novel biodegradation pathway for beta-cp was proposed on the basis of analysis of the metabolites. This strain was also capable of degrading a wide range of pyrethroid insecticides including cypermethrin, deltamethrin, cyhalothrin, and beta-cyfluthrin, which similar to beta-cp are hazardous chemicals. Taken together, our results depict the biodegradation pathway of beta-cp and highlight the promising potentials of strain BSF01 in bioremediation of pyrethroid-contaminated environments. PMID:25398281

Xiao, Ying; Chen, Shaohua; Gao, Yuanqi; Hu, Wei; Hu, Meiying; Zhong, Guohua

2014-11-15

352

Degradation pathway of 2,5-di- tert-butyl-1,4-dimethoxybenzene at high potential  

Microsoft Academic Search

2,5-Di-tert-butyl-1,4-dimethoxybenzene is a redox shuttle for overcharge protection of LiFePO4-based lithium-ion batteries with a working potential lower than 4.2V versus Li+\\/Li. However, an irreversible decomposition occurs when the working potential is higher than 4.2V versus Li+\\/Li. 2,3,5,6-Tetrafluoro-1,4-di-tert-butoxybenzene was also investigated as a model molecule to further understand the possible degradation pathway of 2,5-di-tert-butyl-1,4-dimethxybenzene. The experimental results strongly suggest that the

Zonghai Chen; K. Amine

2007-01-01

353

Identification and characterization of the furfural and 5-(hydroxymethyl)furfural degradation pathways of Cupriavidus basilensis HMF14  

PubMed Central

The toxic fermentation inhibitors in lignocellulosic hydrolysates pose significant problems for the production of second-generation biofuels and biochemicals. Among these inhibitors, 5-(hydroxymethyl)furfural (HMF) and furfural are specifically notorious. In this study, we describe the complete molecular identification and characterization of the pathway by which Cupriavidus basilensis HMF14 metabolizes HMF and furfural. The identification of this pathway enabled the construction of an HMF and furfural-metabolizing Pseudomonas putida. The genetic information obtained furthermore enabled us to predict the HMF and furfural degrading capabilities of sequenced bacterial species that had not previously been connected to furanic aldehyde metabolism. These results pave the way for in situ detoxification of lignocellulosic hydrolysates, which is a major step toward improved efficiency of utilization of lignocellulosic feedstock. PMID:20194784

Koopman, Frank; Wierckx, Nick; de Winde, Johannes H.; Ruijssenaars, Harald J.

2010-01-01

354

Molecular pathways: translational potential of deubiquitinases as drug targets.  

PubMed

The ubiquitin proteasome system (UPS) is the main system for controlled protein degradation and a key regulator of fundamental cellular processes. The dependency of cancer cells on a functioning UPS coupled with the clinical success of bortezomib for the treatment of multiple myeloma have made the UPS an obvious target for drug development. Deubiquitinases (DUB) are components of the UPS that encompass a diverse family of ubiquitin isopeptidases that catalyze the removal of ubiquitin moieties from target proteins or from polyubiquitin chains, resulting in altered signaling or changes in protein stability. Increasing evidence has implicated deregulation of DUB activity in the initiation and progression of cancer. The altered pattern of DUB expression observed in many tumors can potentially serve as a clinical marker for predicting disease outcome and therapy response. The finding of DUB overexpression in tumor cells suggests that they may serve as novel targets for the development of anticancer therapies. Several specific and broad-spectrum DUB inhibitors are shown to have antitumor activity in preclinical in vivo models with low levels of systemic toxicity. Future studies will hopefully establish the clinical potential for DUB inhibitors as a strategy to treat cancer. PMID:25085788

D'Arcy, Pádraig; Linder, Stig

2014-08-01

355

c-myc RNA degradation in growing and differentiating cells: Possible alternate pathways  

SciTech Connect

Transcripts of the proto-oncogene c-myc are composed of a rapidly degraded polyadenylated RNA species and an apparently much more stable, nonadenylated RNA species. In this report, the extended kinetics of c-myc RNA turnover have been examined in rapidly growing cells and in cells induced to differentiate. When transcription was blocked with actinomycin D in rapidly growing cells, poly(A)/sup +/ c-myc was rapidly degraded (t/sub 1/2/ = 12 min). c-myc RNA lacking poly (A) initially remained at or near control levels; however, after 80 to 90 min it was degraded with kinetics similar to those of poly (A)/sup +/ c-myc RNA. These bizarre kinetics are due to the deadenylation of poly (A)/sup +/ c-myc RNA to form poly (A)/sup -/ c-myc, thereby initially maintaining the poly (A)/sup -/ c-myc RNA pool when transcription is blocked. In contrast to growing cells, cells induced to differentiate degraded both poly (A)/sup +/ and poly (A)/sup -/ c-myc RNA rapidly. The rapid disappearance of both RNA species in differentiating cells suggests that a large proportion of the poly (A)/sup +/ c-myc RNA was directly degraded without first being converted to poly (A)/sup -/ c-myc RNA. Others have shown that transcriptional elongation of the c-myc gene is rapidly blocked in differentiating cells. The authors therefore hypothesize that in differentiating cells a direct, rapid degradation of poly (A)/sup +/ c-myc RNA may act as a backup or fail-safe system to ensure that c-myc protein is not synthesized.

Swartwout, S.G. (Roswell Park Memorial Inst., Buffalo, NY (USA)); Kinniburgh, A.J. (Roswell Park Memorial Inst., Buffalo, NY (USA). Dept. of Hematology Research)

1989-01-01

356

Targeting C4-demethylating genes in the cholesterol pathway sensitizes cancer cells to EGFR inhibitors via increased EGFR degradation  

PubMed Central

Persistent signaling by the oncogenic epidermal growth factor receptor (EGFR) is a major source of cancer resistance to EGFR targeting. We established that inactivation of two sterol biosynthesis pathway genes, SC4MOL (sterol C4-methyl oxidase-like) and its partner NSDHL (NADP-dependent steroid dehydrogenase-like), sensitized tumor cells to EGFR inhibitors. Bioinformatics modeling of interactions for the sterol pathway genes in eukaryotes allowed us to hypothesize, and then extensively validate an unexpected role for SC4MOL and NSDHL in controlling the signaling, vesicular trafficking and degradation of EGFR and its dimerization partners, ERBB2 and ERBB3. Metabolic block upstream of SC4MOL with ketoconazole or CYP51A1 siRNA rescued cancer cell viability and EGFR degradation. Inactivation of SC4MOL markedly sensitized A431 xenografts to cetuximab, a therapeutic anti-EGFR antibody. Analysis of Nsdhl-deficient Bpa1H/+ mice confirmed dramatic and selective loss of internalized PDGFR in fibroblasts, and reduced activation of EGFR and its effectors in regions of skin lacking NSDHL. PMID:23125191

Sukhanova, Anna; Gorin, Andrey; Serebriiskii, Ilya G.; Gabitova, Linara; Zheng, Hui; Restifo, Diana; Egleston, Brian L.; Cunningham, David; Bagnyukova, Tetyana; Liu, Hanqing; Nikonova, Anna; Adams, Gregory P.; Zhou, Yan; Yang, Dong-Hua; Mehra, Ranee; Burtness, Barbara; Cai, Kathy Q.; Klein-Szanto, Andres; Kratz, Lisa E.; Kelley, Richard I.; Weiner, Louis M.; Herman, Gail E.; Golemis, Erica A.; Astsaturov, Igor

2012-01-01

357

Bifurcated Degradative Pathway of 3-Sulfolactate in Roseovarius nubinhibens ISM via Sulfoacetaldehyde Acetyltransferase and (S)-Cysteate Sulfolyase ? †  

PubMed Central

Data from the genome sequence of the aerobic, marine bacterium Roseovarius nubinhibens ISM were interpreted such that 3-sulfolactate would be degraded as a sole source of carbon and energy for growth via a novel bifurcated pathway including two known desulfonative enzymes, sulfoacetaldehyde acetyltransferase (EC 2.3.3.15) (Xsc) and cysteate sulfo-lyase (EC 4.4.1.25) (CuyA). Strain ISM utilized sulfolactate quantitatively with stoichiometric excretion of the sulfonate sulfur as sulfate. A combination of enzyme assays, analytical chemistry, enzyme purification, peptide mass fingerprinting, and reverse transcription-PCR data supported the presence of an inducible, tripartite sulfolactate uptake system (SlcHFG), and a membrane-bound sulfolactate dehydrogenase (SlcD) which generated 3-sulfopyruvate, the point of bifurcation. 3-Sulfopyruvate was in part decarboxylated by 3-sulfopyruvate decarboxylase (EC 4.1.1.79) (ComDE), which was purified. The sulfoacetaldehyde that was formed was desulfonated by Xsc, which was identified, and the acetyl phosphate was converted to acetyl-coenzyme A by phosphate acetyltransferase (Pta). The other portion of the 3-sulfopyruvate was transaminated to (S)-cysteate, which was desulfonated by CuyA, which was identified. The sulfite that was formed was presumably exported by CuyZ (TC 9.B.7.1.1 in the transport classification system), and a periplasmic sulfite dehydrogenase is presumed. Bioinformatic analyses indicated that transporter SlcHFG is rare but that SlcD is involved in three different combinations of pathways, the bifurcated pathway shown here, via CuyA alone, and via Xsc alone. This novel pathway involves ComDE in biodegradation, whereas it was discovered in the biosynthesis of coenzyme M. The different pathways of desulfonation of sulfolactate presumably represent final steps in the biodegradation of sulfoquinovose (and exudates derived from it) in marine and aquatic environments. PMID:19581363

Denger, Karin; Mayer, Jutta; Buhmann, Matthias; Weinitschke, Sonja; Smits, Theo H. M.; Cook, Alasdair M.

2009-01-01

358

Direct entry by RNase E is a major pathway for the degradation and processing of RNA in Escherichia coli.  

PubMed

Escherichia coli endoribonuclease E has a major influence on gene expression. It is essential for the maturation of ribosomal and transfer RNA as well as the rapid degradation of messenger RNA. The latter ensures that translation closely follows programming at the level of transcription. Recently, one of the hallmarks of RNase E, i.e. its ability to bind via a 5'-monophosphorylated end, was shown to be unnecessary for the initial cleavage of some polycistronic tRNA precursors. Here we show using RNA-seq analyses of ribonuclease-deficient strains in vivo and a 5'-sensor mutant of RNase E in vitro that, contrary to current models, 5'-monophosphate-independent, 'direct entry' cleavage is a major pathway for degrading and processing RNA. Moreover, we present further evidence that direct entry is facilitated by RNase E binding simultaneously to multiple unpaired regions. These simple requirements may maximize the rate of degradation and processing by permitting multiple sites to be surveyed directly without being constrained by 5'-end tethering. Cleavage was detected at a multitude of sites previously undescribed for RNase E, including ones that regulate the activity and specificity of ribosomes. A potentially broad role for RNase G, an RNase E paralogue, in the trimming of 5'-monophosphorylated ends was also revealed. PMID:25237058

Clarke, Justin E; Kime, Louise; Romero A, David; McDowall, Kenneth J

2015-01-01

359

The Alkyl tert-Butyl Ether Intermediate 2-Hydroxyisobutyrate Is Degraded via a Novel Cobalamin-Dependent Mutase Pathway  

PubMed Central

Fuel oxygenates such as methyl and ethyl tert-butyl ether (MTBE and ETBE, respectively) are degraded only by a limited number of bacterial strains. The aerobic pathway is generally thought to run via tert-butyl alcohol (TBA) and 2-hydroxyisobutyrate (2-HIBA), whereas further steps are unclear. We have now demonstrated for the newly isolated ?-proteobacterial strains L108 and L10, as well as for the closely related strain CIP I-2052, that 2-HIBA was degraded by a cobalamin-dependent enzymatic step. In these strains, growth on substrates containing the tert-butyl moiety, such as MTBE, TBA, and 2-HIBA, was strictly dependent on cobalt, which could be replaced by cobalamin. Tandem mass spectrometry identified a 2-HIBA-induced protein with high similarity to a peptide whose gene sequence was found in the finished genome of the MTBE-degrading strain Methylibium petroleiphilum PM1. Alignment analysis identified it as the small subunit of isobutyryl-coenzyme A (CoA) mutase (ICM; EC 5.4.99.13), which is a cobalamin-containing carbon skeleton-rearranging enzyme, originally described only in Streptomyces spp. Sequencing of the genes of both ICM subunits from strain L108 revealed nearly 100% identity with the corresponding peptide sequences from M. petroleiphilum PM1, suggesting a horizontal gene transfer event to have occurred between these strains. Enzyme activity was demonstrated in crude extracts of induced cells of strains L108 and L10, transforming 2-HIBA into 3-hydroxybutyrate in the presence of CoA and ATP. The physiological and evolutionary aspects of this novel pathway involved in MTBE and ETBE metabolism are discussed. PMID:16751524

Rohwerder, Thore; Breuer, Uta; Benndorf, Dirk; Lechner, Ute; Müller, Roland H.

2006-01-01

360

The alkyl tert-butyl ether intermediate 2-hydroxyisobutyrate is degraded via a novel cobalamin-dependent mutase pathway.  

PubMed

Fuel oxygenates such as methyl and ethyl tert-butyl ether (MTBE and ETBE, respectively) are degraded only by a limited number of bacterial strains. The aerobic pathway is generally thought to run via tert-butyl alcohol (TBA) and 2-hydroxyisobutyrate (2-HIBA), whereas further steps are unclear. We have now demonstrated for the newly isolated beta-proteobacterial strains L108 and L10, as well as for the closely related strain CIP I-2052, that 2-HIBA was degraded by a cobalamin-dependent enzymatic step. In these strains, growth on substrates containing the tert-butyl moiety, such as MTBE, TBA, and 2-HIBA, was strictly dependent on cobalt, which could be replaced by cobalamin. Tandem mass spectrometry identified a 2-HIBA-induced protein with high similarity to a peptide whose gene sequence was found in the finished genome of the MTBE-degrading strain Methylibium petroleiphilum PM1. Alignment analysis identified it as the small subunit of isobutyryl-coenzyme A (CoA) mutase (ICM; EC 5.4.99.13), which is a cobalamin-containing carbon skeleton-rearranging enzyme, originally described only in Streptomyces spp. Sequencing of the genes of both ICM subunits from strain L108 revealed nearly 100% identity with the corresponding peptide sequences from M. petroleiphilum PM1, suggesting a horizontal gene transfer event to have occurred between these strains. Enzyme activity was demonstrated in crude extracts of induced cells of strains L108 and L10, transforming 2-HIBA into 3-hydroxybutyrate in the presence of CoA and ATP. The physiological and evolutionary aspects of this novel pathway involved in MTBE and ETBE metabolism are discussed. PMID:16751524

Rohwerder, Thore; Breuer, Uta; Benndorf, Dirk; Lechner, Ute; Müller, Roland H

2006-06-01

361

Stability of 6:2 fluorotelomer sulfonate in advanced oxidation processes: degradation kinetics and pathway.  

PubMed

Perfluorooctane sulfonate (PFOS), a widely used mist suppressant in hard chrome electroplating industry, has been listed in the Stockholm Convention for global ban. 6:2 Fluorotelomer sulfonate (6:2 FTS) acid and salts have been adopted as alternative products in the market, but no data about their abiotic degradation has been reported. In the present study, the degradability of 6:2 FTS potassium salt (6:2 FTS-K) was evaluated under various advanced oxidation processes, including ultraviolet (UV) irradiation, UV with hydrogen peroxide (H2O2), alkaline ozonation (O3, pH = 11), peroxone (O3/H2O2), and Fenton reagent oxidation (Fe(2+)/H2O2). UV/H2O2 was found to be the most effective approach, where the degradation of 6:2 FTS-K followed the pseudo-first-order kinetics. The intermediates were mainly shorter chain perfluoroalkyl carboxylic acid (C7 to C2), while sulfate (SO4 (2-)) and fluoride (F(-)) were found to be the final products. The high yields of SO4 (2-) and F(-) indicate that 6:2 FTS-K can be nearly completely desulfonated and defluorinated under UV/H2O2 condition. The degradation should firstly begin with the substitution of hydrogen atom by hydroxyl radicals, followed by desulfonation, carboxylation, and sequential "flake off" of CF2 unit. Compared with PFOS which is inert in most advanced oxidation processes, 6:2 FTS-K is more degradable as the alternative. PMID:24352540

Yang, Xiaoling; Huang, Jun; Zhang, Kunlun; Yu, Gang; Deng, Shubo; Wang, Bin

2014-03-01

362

Organellar oligopeptidase (OOP) provides a complementary pathway for targeting peptide degradation in mitochondria and chloroplasts  

PubMed Central

Both mitochondria and chloroplasts contain distinct proteolytic systems for precursor protein processing catalyzed by the mitochondrial and stromal processing peptidases and for the degradation of targeting peptides catalyzed by presequence protease. Here, we have identified and characterized a component of the organellar proteolytic systems in Arabidopsis thaliana, the organellar oligopeptidase, OOP (At5g65620). OOP belongs to the M3A family of peptide-degrading metalloproteases. Using two independent in vivo methods, we show that the protease is dually localized to mitochondria and chloroplasts. Furthermore, we localized the OPP homolog At5g10540 to the cytosol. Analysis of peptide degradation by OOP revealed substrate size restriction from 8 to 23 aa residues. Short mitochondrial targeting peptides (presequence of the ribosomal protein L29 and presequence of 1-aminocyclopropane-1-carboxylic acid deaminase 1) and N- and C-terminal fragments derived from the presequence of the ATPase beta subunit ranging in size from 11 to 20 aa could be degraded. MS analysis showed that OOP does not exhibit a strict cleavage pattern but shows a weak preference for hydrophobic residues (F/L) at the P1 position. The crystal structures of OOP, at 1.8–1.9 Å, exhibit an ellipsoidal shape consisting of two major domains enclosing the catalytic cavity of 3,000 Å3. The structural and biochemical data suggest that the protein undergoes conformational changes to allow peptide binding and proteolysis. Our results demonstrate the complementary role of OOP in targeting-peptide degradation in mitochondria and chloroplasts. PMID:24043784

Kmiec, Beata; Teixeira, Pedro F.; Berntsson, Ronnie P.-A.; Murcha, Monika W.; Branca, Rui M. M.; Radomiljac, Jordan D.; Regberg, Jakob; Svensson, Linda M.; Bakali, Amin; Langel, Ülo; Lehtiö, Janne; Whelan, James; Stenmark, Pål; Glaser, Elzbieta

2013-01-01

363

Comparison of EJC-enhanced and EJC-independent NMD in human cells reveals two partially redundant degradation pathways  

PubMed Central

Nonsense-mediated mRNA decay (NMD) is a eukaryotic post-transcriptional gene regulation mechanism that eliminates mRNAs with the termination codon (TC) located in an unfavorable environment for efficient translation termination. The best-studied NMD-targeted mRNAs contain premature termination codons (PTCs); however, NMD regulates even many physiological mRNAs. An exon-junction complex (EJC) located downstream from a TC acts as an NMD-enhancing signal, but is not generally required for NMD. Here, we compared these “EJC-enhanced” and “EJC-independent” modes of NMD with regard to their requirement for seven known NMD factors in human cells using two well-characterized NMD reporter genes (immunoglobulin ? and ?-Globin) with or without an intron downstream from the PTC. We show that both NMD modes depend on UPF1 and SMG1, but detected transcript-specific differences with respect to the requirement for UPF2 and UPF3b, consistent with previously reported UPF2- and UPF3-independent branches of NMD. In addition and contrary to expectation, a higher sensitivity of EJC-independent NMD to reduced UPF2 and UPF3b concentrations was observed. Our data further revealed a redundancy of the endo- and exonucleolytic mRNA degradation pathways in both modes of NMD. Moreover, the relative contributions of both decay pathways differed between the reporters, with PTC-containing immunoglobulin ? transcripts being preferentially subjected to SMG6-mediated endonucleolytic cleavage, whereas ?-Globin transcripts were predominantly degraded by the SMG5/SMG7-dependent pathway. Overall, the surprising heterogeneity observed with only two NMD reporter pairs suggests the existence of several mechanistically distinct branches of NMD in human cells. PMID:23962664

Metze, Stefanie; Herzog, Veronika A.; Ruepp, Marc-David; Mühlemann, Oliver

2013-01-01

364

Caspase-3 triggers a TPCK-sensitive protease pathway leading to degradation of the BH3-only protein puma.  

PubMed

The protein Puma (p53-upregulated modulator of apoptosis) belongs to the BH3-only group of the Bcl-2 family and is a major regulator of apoptosis. Although the transcriptional regulation of Puma is clearly established, little is known about the regulation of its expression at the protein levels. We show here that various signals--including the cytokine TGF?, the death effector TRAIL or chemical drugs such as anisomycin--downregulate Puma protein levels via a novel pathway based on the sequential activation of caspase-3 and a protease inhibited by the serpase inhibitor N-tosyl-L-phenylalanine chloromethyl ketone. This pathway is specific for Puma because (1) the levels of other BH3-only proteins, such as Bim and Noxa were not modified by these stimuli and (2) this caspase-mediated degradation was dependent on both the BH3 and C-terminal domains of Puma. Our data also show that Puma is regulated during the caspase-3-dependent differentiation of murine embryonic stem cells and suggest that this pathway may be relevant and important during caspase-mediated cell differentiation not associated with apoptosis. PMID:20640889

Hadji, Abbas; Clybouw, Cyril; Auffredou, Marie-Thérèse; Alexia, Catherine; Poalas, Konstantinos; Burlion, Aude; Feraud, Olivier; Leca, Gérald; Vazquez, Aimé

2010-12-01

365

Influences of light and oxygen conditions on photosynthetic bacteria macromolecule degradation: different metabolic pathways.  

PubMed

Direct degradation of macromolecules by photosynthetic bacteria (PSB) is important for the industrial application of PSB wastewater treatment. Light and oxygen are the most important parameters in PSB growth. This paper studied the PSB macromolecule degradation process under three different light and oxygen conditions: light-anaerobic, natural light-microaerobic and dark-aerobic. The results showed that under three different light-oxygen conditions, PSB degradation of macromolecules was higher than 90%; the removal ratios of COD, TN, TP, total sugar and protein were also high; and the biomass yield reached nearly 0.5 mg-biomass/mg-COD-removal. Light and oxygen significantly influenced the efficiency. Macromolecules and pollutants removals were higher under oxygen condition than those under light-anaerobic condition. Theoretical analysis showed that under aerobic condition, PSB carried out oxidative phosphorylation, in which pollutants were sufficiently utilized with high mineralization degree. Under light-anaerobic condition, PSB carried out photophosphorylation and fermentation, which led to low pollutants removal efficiency. PMID:21871794

Lu, Haifeng; Zhang, Guangming; Wan, Tian; Lu, Yufeng

2011-10-01

366

Protein degradation and protein synthesis in long-term memory formation  

PubMed Central

Long-term memory (LTM) formation requires transient changes in the activity of intracellular signaling cascades that are thought to regulate new gene transcription and de novo protein synthesis in the brain. Consistent with this, protein synthesis inhibitors impair LTM for a variety of behavioral tasks when infused into the brain around the time of training or following memory retrieval, suggesting that protein synthesis is a critical step in LTM storage in the brain. However, evidence suggests that protein degradation mediated by the ubiquitin-proteasome system (UPS) may also be a critical regulator of LTM formation and stability following retrieval. This requirement for increased protein degradation has been shown in the same brain regions in which protein synthesis is required for LTM storage. Additionally, increases in the phosphorylation of proteins involved in translational control parallel increases in protein polyubiquitination and the increased demand for protein degradation is regulated by intracellular signaling molecules thought to regulate protein synthesis during LTM formation. In some cases inhibiting proteasome activity can rescue memory impairments that result from pharmacological blockade of protein synthesis, suggesting that protein degradation may control the requirement for protein synthesis during the memory storage process. Results such as these suggest that protein degradation and synthesis are both critical for LTM formation and may interact to properly “consolidate” and store memories in the brain. Here, we review the evidence implicating protein synthesis and degradation in LTM storage and highlight the areas of overlap between these two opposing processes. We also discuss evidence suggesting these two processes may interact to properly form and store memories. LTM storage likely requires a coordinated regulation between protein degradation and synthesis at multiple sites in the mammalian brain. PMID:25018696

Jarome, Timothy J.; Helmstetter, Fred J.

2014-01-01

367

An NMD Pathway in Yeast Involving Accelerated Deadenylation and Exosome-Mediated 3??5? Degradation  

Microsoft Academic Search

Eukaryotic mRNAs containing premature termination codons are subjected to accelerated turnover, known as nonsense-mediated decay (NMD). Recognition of translation termination events as premature requires a surveillance complex, which includes the RNA helicase Upf1p. In Saccharomyces cerevisiae, NMD provokes rapid decapping followed by 5??3? exonucleolytic decay. Here we report an alternative, decapping-independent NMD pathway involving deadenylation and subsequent 3??5? exonucleolytic decay.

Philip Mitchell; David Tollervey

2003-01-01

368

Relationship between the proteasomal system and autophagy  

PubMed Central

Two major pathways degrade most cellular proteins in eukaryotic cells: the ubiquitin–proteasome system (UPS), which usually degrades the majority of proteins, and autophagy, primarily responsible for the degradation of most long-lived or aggregated proteins and cellular organelles. Disruption of these processes can contribute to pathology of a variety of diseases. Further, both pathways are critical for the maintenance of several aspects of cellular homeostasis, but, until recently, were thought to be largely distinct. Recent advances in this field, however, now strongly suggest that their activities are carefully orchestrated through several interfacing elements that are presented and discussed in this review. PMID:23638318

Lilienbaum, Alain

2013-01-01

369

Pathways and metabolites of microbial degradation of selected acidic pharmaceutical and their occurrence in municipal wastewater treated by a membrane bioreactor  

Microsoft Academic Search

Laboratory degradation tests with five acidic pharmaceuticals using activated sludge as inocculum under aerobic conditions were performed and microbial metabolites were analysed by liquid chromatography-mass spectrometry (LC-MS). Ketoprofen was partly mineralized as a sole source of carbon and energy and the metabolites determined by LC-MS suggest microbial ketoprofen degradation to proceed along the pathway known for biphenyls and related compounds.

José Benito Quintana; Stefan Weiss; Thorsten Reemtsma

2005-01-01

370

Oxidation of microcystin-LR by ferrate(VI): kinetics, degradation pathways, and toxicity assessments.  

PubMed

The presence of the potent cyanotoxin, microcystin-LR (MC-LR), in drinking water sources poses a serious risk to public health. The kinetics of the reactivity of ferrate(VI) (Fe(VI)O4(2-), Fe(VI)) with MC-LR and model compounds (sorbic acid, sorbic alcohol, and glycine anhydride) are reported over a range of solution pH. The degradation of MC-LR followed second-order kinetics with the bimolecular rate constant (kMCLR+Fe(VI)) decreasing from 1.3 ± 0.1 × 10(2) M(-1) s(-1) at pH 7.5 to 8.1 ± 0.08 M(-1) s(-1) at pH 10.0. The specific rate constants for the individual ferrate species were determined and compared with a number of common chemical oxidants employed for water treatment. Detailed product studies using liquid chromatography-mass spectrometry/mass spectrometry (LC-MS/MS) indicated the oxidized products (OPs) were primarily the result of hydroxylation of the aromatic ring, double bond of the methyldehydroalanine (Mdha) amino acid residue, and diene functionality. Products studies also indicate fragmentation of the cyclic MC-LR structure occurs under the reaction conditions. The analysis of protein phosphatase (PP1) activity suggested that the degradation byproducts of MC-LR did not possess significant biological toxicity. Fe(VI) was effective for the degradation MC-LR in water containing carbonate ions and fulvic acid (FA) and in lake water samples, but higher Fe(VI) dosages would be needed to completely remove MC-LR in lake water compared to deionized water. PMID:25215438

Jiang, Wenjun; Chen, Long; Batchu, Sudha Rani; Gardinali, Piero R; Jasa, Libor; Marsalek, Blahoslav; Zboril, Radek; Dionysiou, Dionysios D; O'Shea, Kevin E; Sharma, Virender K

2014-10-21

371

New pathway for degradation of sulfonated azo dyes by microbial peroxidases of Phanerochaete chrysosporium and Streptomyces chromofuscus.  

PubMed Central

Pathways for the degradation of 3,5-dimethyl-4-hydroxy-azobenzene-4'-sulfonic acid (I) and 3-methoxy-4-hydroxyazobenzene-4'-sulfonamide (II) by the manganese peroxidase and ligninase of Phanerochaete chrysosporium and by the peroxidase of Streptomyces chromofuscus have been proposed. Twelve metabolic products were found, and their mechanisms of formation were explained. Preliminary oxidative activation of the dyes resulted in the formation of cationic species, making the molecules vulnerable to the nucleophilic attack of water. Two types of hydrolytic cleavage were observed. Asymmetric splitting gave rise to quinone and diazene derivatives, while symmetric splitting resulted in the formation of quinone monoimine and nitroso derivatives. These unstable intermediates underwent further redox, oxidation, and hydrolytic transformation, eventually furnishing 11 organic products and ammonia. PMID:8113173

Goszczynski, S; Paszczynski, A; Pasti-Grigsby, M B; Crawford, R L; Crawford, D L

1994-01-01

372

The unique degradation pathway of the PTS2 receptor, Pex7, is dependent on the PTS receptor/coreceptor, Pex5 and Pex20  

PubMed Central

Peroxisomal matrix protein import uses two peroxisomal targeting signals (PTSs). Most matrix proteins use the PTS1 pathway and its cargo receptor, Pex5. The PTS2 pathway is dependent on another receptor, Pex7, and its coreceptor, Pex20. We found that during the matrix protein import cycle, the stability and dynamics of Pex7 differ from those of Pex5 and Pex20. In Pichia pastoris, unlike Pex5 and Pex20, Pex7 is constitutively degraded in wild-type cells but is stabilized in pex mutants affecting matrix protein import. Degradation of Pex7 is more prevalent in cells grown in methanol, in which the PTS2 pathway is nonessential, in comparison with oleate, suggesting regulation of Pex7 turnover. Pex7 must shuttle into and out of peroxisomes before it is polyubiquitinated and degraded by the proteasome. The shuttling of Pex7, and consequently its degradation, is dependent on the receptor recycling pathways of Pex5 and Pex20 and relies on an interaction between Pex7 and Pex20. We also found that blocking the export of Pex20 from peroxisomes inhibits PTS1-mediated import, suggesting sharing of limited components in the export of PTS receptors/coreceptors. The shuttling and stability of Pex7 are divergent from those of Pex5 and Pex20, exemplifying a novel interdependence of the PTS1 and PTS2 pathways. PMID:25009284

Hagstrom, Danielle; Ma, Changle; Guha-Polley, Soumi; Subramani, Suresh

2014-01-01

373

The atzB gene of Pseudomonas sp. strain ADP encodes the second enzyme of a novel atrazine degradation pathway.  

PubMed Central

We previously reported the isolation of a 21.5-kb genomic DNA fragment from Pseudomonas sp. strain ADP, which contains the atzA gene, encoding the first metabolic step for the degradation of the herbicide atrazine (M. de Souza, L. P. Wackett, K. L. Boundy-Mills, R. T. Mandelbaum, and M. J. Sadowsky, Appl. Environ. Microbiol. 61:3373-3378, 1995). In this study, we show that this fragment also contained the second gene of the atrazine metabolic pathway, atzB. AtzB catalyzed the transformation of hydroxyatrazine to N-isopropylammelide. The product was identified by use of high-performance liquid chromatography, mass spectrometery, and nuclear magnetic resonance spectroscopy. Tn5 mutagenesis of pMD1 was used to determine that atzB was located 8 kb downstream of atzA. Hydroxyatrazine degradation activity was localized to a 4.0-kb ClaI fragment, which was subcloned into the vector pACYC184 to produce plasmid pATZB-2. The DNA sequence of this region was determined and found to contain two large overlapping divergent open reading frames, ORF1 and ORF2. ORF1 was identified as the coding region of atzB by demonstrating that (i) only ORF1 was transcribed in Pseudomonas sp. strain ADP, (ii) a Tn5 insertion in ORF2 did not disrupt function, and (iii) codon usage was consistent with ORF1 being translated. AtzB had 25% amino acid identity with TrzA, a protein that catalyzes a hydrolytic deamination of the s-triazine substrate melamine. The atzA and atzB genes catalyze the first two steps of the metabolic pathway in a bacterium that rapidly metabolizes atrazine to carbon dioxide, ammonia, and chloride. PMID:9055410

Boundy-Mills, K L; de Souza, M L; Mandelbaum, R T; Wackett, L P; Sadowsky, M J

1997-01-01

374

Electrochemical degradation of sulfonamides at BDD electrode: kinetics, reaction pathway and eco-toxicity evaluation.  

PubMed

The investigation dealt with electrochemical oxidation of five sulfonamides (SNs): sulfadiazine (SDZ), sulfathiazole (STZ), sulfamerazine (SMR), sulfamethazine (SMN) and sulfadimethoxine (SDM) in aqueous solution at boron-doped diamond (BDD) electrode. All studied sulfonamides were degraded according to a pseudo first order kinetics. The structure of SNs had no significant effect on the values of pseudo first order rate constants. Increased degradation efficiency was observed in higher temperature and in acidic pH. Due to the presence of chlorine and nitrate SNs were more effectively oxidized from municipal wastewater treatment plant (WWTP) effluents than from pure supporting electrolyte Na2SO4. The intermediates identified by LC-MS and GC-MS analysis suggested that the hydroxyl radicals attack mainly the SN bond, but also the aromatic ring systems (aniline, pyrimidine or triazole) of SNs. Finally, the toxicity of the SNs solutions and effluents after electrochemical treatment was assessed through the measurement of growth inhibition of green algae (Scenedesmus vacualatus) and duckweed (Lemna minor). Toxicity of SMR, STZ, SMN solutions before and after electrochemical oxidation and SDM solution after the process in L. minor test was observed. No significant toxicity of studied SNs was observed in algae test. PMID:25215656

Fabia?ska, Aleksandra; Bia?k-Bieli?ska, Anna; Stepnowski, Piotr; Stolte, Stefan; Siedlecka, Ewa Maria

2014-09-15

375

A Shared Endoplasmic Reticulum-associated Degradation Pathway Involving the EDEM1 Protein for Glycosylated and Nonglycosylated Proteins*  

PubMed Central

Studies of misfolded protein targeting to endoplasmic reticulum-associated degradation (ERAD) have largely focused on glycoproteins, which include the bulk of the secretory proteins. Mechanisms of targeting of nonglycosylated proteins are less clear. Here, we studied three nonglycosylated proteins and analyzed their use of known glycoprotein quality control and ERAD components. Similar to an established glycosylated ERAD substrate, the uncleaved precursor of asialoglycoprotein receptor H2a, its nonglycosylated mutant, makes use of calnexin, EDEM1, and HRD1, but only glycosylated H2a is a substrate for the cytosolic SCFFbs2 E3 ubiquitin ligase with lectin activity. Two nonglycosylated BiP substrates, NS-1? light chain and truncated Ig? heavy chain, interact with the ERAD complex lectins OS-9 and XTP3-B and require EDEM1 for degradation. EDEM1 associates through a region outside of its mannosidase-like domain with the nonglycosylated proteins. Similar to glycosylated substrates, proteasomal inhibition induced accumulation of the nonglycosylated proteins and ERAD machinery in the endoplasmic reticulum-derived quality control compartment. Our results suggest a shared ERAD pathway for glycosylated and nonglycosylated proteins composed of luminal lectin machinery components also capable of protein-protein interactions. PMID:23233672

Shenkman, Marina; Groisman, Bella; Ron, Efrat; Avezov, Edward; Hendershot, Linda M.; Lederkremer, Gerardo Z.

2013-01-01

376

Biochemical, Transcriptional and Translational Evidences of the Phenol-meta-Degradation Pathway by the Hyperthermophilic Sulfolobus solfataricus 98/2  

PubMed Central

Phenol is a widespread pollutant and a model molecule to study the biodegradation of monoaromatic compounds. After a first oxidation step leading to catechol in mesophilic and thermophilic microorganisms, two main routes have been identified depending on the cleavage of the aromatic ring: ortho involving a catechol 1,2 dioxygenase (C12D) and meta involving a catechol 2,3 dioxygenase (C23D). Our work aimed at elucidating the phenol-degradation pathway in the hyperthermophilic archaea Sulfolobus solfataricus 98/2. For this purpose, the strain was cultivated in a fermentor under different substrate and oxygenation conditions. Indeed, reducing dissolved-oxygen concentration allowed slowing down phenol catabolism (specific growth and phenol-consumption rates dropped 55% and 39%, respectively) and thus, evidencing intermediate accumulations in the broth. HPLC/Diode Array Detector and LC-MS analyses on culture samples at low dissolved-oxygen concentration (DOC ?=? 0.06 mg.L?1) suggested, apart for catechol, the presence of 2-hydroxymuconic acid, 4-oxalocrotonate and 4-hydroxy-2-oxovalerate, three intermediates of the meta route. RT-PCR analysis on oxygenase-coding genes of S. solfataricus 98/2 showed that the gene coding for the C23D was expressed only on phenol. In 2D-DIGE/MALDI-TOF analysis, the C23D was found and identified only on phenol. This set of results allowed us concluding that S. solfataricus 98/2 degrade phenol through the meta route. PMID:24349276

Davidson, Sylvain; Pophillat, Matthieu; Lorquin, Jean; Auria, Richard; Simon, Gwenola; Casalot, Laurence

2013-01-01

377

Biochemical, transcriptional and translational evidences of the phenol-meta-degradation pathway by the hyperthermophilic Sulfolobus solfataricus 98/2.  

PubMed

Phenol is a widespread pollutant and a model molecule to study the biodegradation of monoaromatic compounds. After a first oxidation step leading to catechol in mesophilic and thermophilic microorganisms, two main routes have been identified depending on the cleavage of the aromatic ring: ortho involving a catechol 1,2 dioxygenase (C12D) and meta involving a catechol 2,3 dioxygenase (C23D). Our work aimed at elucidating the phenol-degradation pathway in the hyperthermophilic archaea Sulfolobus solfataricus 98/2. For this purpose, the strain was cultivated in a fermentor under different substrate and oxygenation conditions. Indeed, reducing dissolved-oxygen concentration allowed slowing down phenol catabolism (specific growth and phenol-consumption rates dropped 55% and 39%, respectively) and thus, evidencing intermediate accumulations in the broth. HPLC/Diode Array Detector and LC-MS analyses on culture samples at low dissolved-oxygen concentration (DOC ?=? 0.06 mg x L(-1)) suggested, apart for catechol, the presence of 2-hydroxymuconic acid, 4-oxalocrotonate and 4-hydroxy-2-oxovalerate, three intermediates of the meta route. RT-PCR analysis on oxygenase-coding genes of S. solfataricus 98/2 showed that the gene coding for the C23D was expressed only on phenol. In 2D-DIGE/MALDI-TOF analysis, the C23D was found and identified only on phenol. This set of results allowed us concluding that S. solfataricus 98/2 degrade phenol through the meta route. PMID:24349276

Comte, Alexia; Christen, Pierre; Davidson, Sylvain; Pophillat, Matthieu; Lorquin, Jean; Auria, Richard; Simon, Gwenola; Casalot, Laurence

2013-01-01

378

Generation of Dopamine Neurons with Improved Cell Survival and Phenotype Maintenance Using a Degradation-Resistant Nurr1 Mutant  

PubMed Central

Nurr1 is a transcription factor specific for the development and maintenance of the midbrain dopamine (DA) neurons. Exogenous Nurr1 in neural precursor (NP) cells induces the differentiation of DA neurons in vitro that are capable of reversing motor dysfunctions in a rodent model for Parkinson disease. The promise of this therapeutic approach, however, is unclear due to poor cell survival and phenotype loss of DA cells after transplantation. We herein demonstrate that Nurr1 proteins undergo ubiquitin-proteasome-system-mediated degradation in differentiating NP cells. The degradation process is activated by a direct Akt-mediated phosphorylation of Nurr1 proteins and can be prevented by abolishing the Akt-target sequence in Nurr1 (Nurr1Akt). Overexpression of Nurr1Akt in NP cells yielded DA neurons in which Nurr1 protein levels were maintained for prolonged periods. The sustained Nurr1 expression endowed the Nurr1Akt-induced DA neurons with resistance to toxic stimuli, enhanced survival, and sustained DA phenotypes in vitro and in vivo after transplantation. PMID:19522012

Jo, A-Young; Kim, Mi-Young; Lee, Hyun-Seob; Rhee, Yong-Hee; Lee, Jeong-Eun; Baek, Kwang-Hyun; Park, Chang-Hwan; Koh, Hyun-Chul; Shin, Incheol; Lee, Yong-Sung; Lee, Sang-Hun

2009-01-01

379

FBXL5-mediated degradation of single-stranded DNA-binding protein hSSB1 controls DNA damage response.  

PubMed

Human single-strand (ss) DNA binding proteins 1 (hSSB1) has been shown to participate in DNA damage response and maintenance of genome stability by regulating the initiation of ATM-dependent signaling. ATM phosphorylates hSSB1 and prevents hSSB1 from ubiquitin-proteasome-mediated degradation. However, the E3 ligase that targets hSSB1 for destruction is still unknown. Here, we report that hSSB1 is the bona fide substrate for an Fbxl5-containing SCF (Skp1-Cul1-F box) E3 ligase. Fbxl5 interacts with and targets hSSB1 for ubiquitination and degradation, which could be prevented by ATM-mediated hSSB1 T117 phosphorylation. Furthermore, cells overexpression of Fbxl5 abrogated the cellular response to DSBs, including activation of ATM and phosphorylation of ATM targets and exhibited increased radiosensitivity, chemosensitivity and defective checkpoint activation after genotoxic stress stimuli. Moreover, the protein levels of hSSB1 and Fbxl5 showed an inverse correlation in lung cancer cells lines and clinical lung cancer samples. Therefore, Fbxl5 may negatively modulate hSSB1 to regulate DNA damage response, implicating Fbxl5 as a novel, promising therapeutic target for lung cancers. PMID:25249620

Chen, Zhi-Wei; Liu, Bin; Tang, Nai-Wang; Xu, Yun-Hua; Ye, Xiang-Yun; Li, Zi-Ming; Niu, Xiao-Min; Shen, Sheng-Ping; Lu, Shun; Xu, Ling

2015-01-01

380

Coupled activation and degradation of eEF2K regulates protein synthesis in response to genotoxic stress.  

PubMed

The kinase eEF2K [eukaryotic elongation factor 2 (eEF2) kinase] controls the rate of peptide chain elongation by phosphorylating eEF2, the protein that mediates the movement of the ribosome along the mRNA by promoting translocation of the transfer RNA from the A to the P site in the ribosome. eEF2K-mediated phosphorylation of eEF2 on threonine 56 (Thr??) decreases its affinity for the ribosome, thereby inhibiting elongation. Here, we show that in response to genotoxic stress, eEF2K was activated by AMPK (adenosine monophosphate-activated protein kinase)-mediated phosphorylation on serine 398. Activated eEF2K phosphorylated eEF2 and induced a temporary ribosomal slowdown at the stage of elongation. Subsequently, during DNA damage checkpoint silencing, a process required to allow cell cycle reentry, eEF2K was degraded by the ubiquitin-proteasome system through the ubiquitin ligase SCF(?TrCP) (Skp1-Cul1-F-box protein, ?-transducin repeat-containing protein) to enable rapid resumption of translation elongation. This event required autophosphorylation of eEF2K on a canonical ?TrCP-binding domain. The inability to degrade eEF2K during checkpoint silencing caused sustained phosphorylation of eEF2 on Thr?? and delayed the resumption of translation elongation. Our study therefore establishes a link between DNA damage signaling and translation elongation. PMID:22669845

Kruiswijk, Flore; Yuniati, Laurensia; Magliozzi, Roberto; Low, Teck Yew; Lim, Ratna; Bolder, Renske; Mohammed, Shabaz; Proud, Christopher G; Heck, Albert J R; Pagano, Michele; Guardavaccaro, Daniele

2012-06-01