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Sample records for autophagy ubiquitination inflammation

  1. Ubiquitin and Autophagy%泛素与自噬

    Institute of Scientific and Technical Information of China (English)

    冯梅; 王莉新; 王易

    2011-01-01

    Protein degradation mediated by ubiquitin and autophagy are the basic mechanisms involved in cellular self-regulation. Ubiquitin may be involved in the process of autophagy by serving as a umversal recognition signal. Induction of autophagy can promote ubiquitination, thereby enhancing the degradation of substrate. This paper mainly focuses on the relation and the potential mutual regulation between ubiquitination and autophagy, as well as the phenomenon of programmed cell death that is associated with both ubiquitination and autophagy processes.%泛素调节的蛋白质降解过程和细胞的自噬现象都是细胞自我调节的基本机制.其中,泛素可能作为一种普遍的识别信号参与了自噬过程;而自噬的诱导又能促进泛素化作用,从而增强对底物的降解.本文着重探讨这两者间的关系及可能存在的相互调节作用,并兼及两者共同涉及的细胞程序性死亡现象.

  2. Autophagy is an inflammation-related defensive mechanism against disease.

    Science.gov (United States)

    Joven, Jorge; Guirro, Maria; Mariné-Casadó, Roger; Rodríguez-Gallego, Esther; Menéndez, Javier A

    2014-01-01

    The inflammatory response is an energy-intensive process. Consequently, metabolism is closely associated with immune function. The autophagy machinery plays a role in metabolism by providing energy but may also be used to attack invading pathogens (xenophagy). The autophagy machinery may function to protect against not only the threats of infection but also the threats of the host's own response acting on the central immunological tolerance and the negative regulation of innate and inflammatory signaling. The balance between too little and too much autophagy is critical for the survival of immune cells because autophagy is linked to type 2-cell death programmed necrosis and apoptosis. Changes in inflammatory cells are driven by extracellular signals; however, the mechanisms by which cytokines mediate autophagy regulation and govern immune cell function remain unknown. Certain cytokines increase autophagy, whereas others inhibit autophagy. The relationship between autophagy and inflammation is also important in the pathogenesis of metabolic, non-communicable diseases. Inflammation per se is not the cause of obesity-associated diseases, but it is secondary to both the positive energy balance and the specific cellular responses. In metabolic tissues, the suppression of autophagy increases inflammation with the overexpression of cytokines, resulting in an activation of autophagy. The physiological role of these apparently contradictory findings remains uncertain but exemplifies future challenges in the therapeutic modulation of autophagy in the management of disease.

  3. RNF2 is recruited by WASH to ubiquitinate AMBRA1 leading to downregulation of autophagy

    OpenAIRE

    Xia, Pengyan; Wang, Shuo; Huang, Guanling; Du, Ying; Zhu, Pingping; Li, Man; Fan, Zusen

    2014-01-01

    WASH (Wiskott-Aldrich syndrome protein (WASP) and SCAR homolog) was identified to function in endosomal sorting via Arp2/3 activation. We previously demonstrated that WASH is a new interactor of BECN1 and present in the BECN1-PIK3C3 complex with AMBRA1. The AMBRA1-DDB1-CUL4A complex is an E3 ligase for K63-linked ubiquitination of BECN1, which is required for starvation-induced autophagy. WASH suppresses autophagy by inhibition of BECN1 ubiquitination. However, how AMBRA1 is regulated during ...

  4. NBR1-mediated selective autophagy targets insoluble ubiquitinated protein aggregates in plant stress responses.

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    Jie Zhou

    Full Text Available Plant autophagy plays an important role in delaying senescence, nutrient recycling, and stress responses. Functional analysis of plant autophagy has almost exclusively focused on the proteins required for the core process of autophagosome assembly, but little is known about the proteins involved in other important processes of autophagy, including autophagy cargo recognition and sequestration. In this study, we report functional genetic analysis of Arabidopsis NBR1, a homolog of mammalian autophagy cargo adaptors P62 and NBR1. We isolated two nbr1 knockout mutants and discovered that they displayed some but not all of the phenotypes of autophagy-deficient atg5 and atg7 mutants. Like ATG5 and ATG7, NBR1 is important for plant tolerance to heat, oxidative, salt, and drought stresses. The role of NBR1 in plant tolerance to these abiotic stresses is dependent on its interaction with ATG8. Unlike ATG5 and ATG7, however, NBR1 is dispensable in age- and darkness-induced senescence and in resistance to a necrotrophic pathogen. A selective role of NBR1 in plant responses to specific abiotic stresses suggest that plant autophagy in diverse biological processes operates through multiple cargo recognition and delivery systems. The compromised heat tolerance of atg5, atg7, and nbr1 mutants was associated with increased accumulation of insoluble, detergent-resistant proteins that were highly ubiquitinated under heat stress. NBR1, which contains an ubiquitin-binding domain, also accumulated to high levels with an increasing enrichment in the insoluble protein fraction in the autophagy-deficient mutants under heat stress. These results suggest that NBR1-mediated autophagy targets ubiquitinated protein aggregates most likely derived from denatured or otherwise damaged nonnative proteins generated under stress conditions.

  5. WASH inhibits autophagy through suppression of Beclin 1 ubiquitination

    OpenAIRE

    Xia, Pengyan; Wang, Shuo; Du, Ying; Zhao, Zhenao; Shi, Lei; Sun, Lei; Huang, Guanling; Ye, Buqing; Li, Chong; Dai, Zhonghua; Hou, Ning; Cheng, Xuan; Sun, Qingyuan; Li, Lei(Beijing Institute of Petrochemical Technology, Beijing, 102617, People's Republic of China); Yang, Xiao

    2013-01-01

    Autophagy degrades cytoplasmic proteins and organelles to recycle cellular components that are required for cell survival and tissue homeostasis. However, it is not clear how autophagy is regulated in mammalian cells. WASH (Wiskott–Aldrich syndrome protein (WASP) and SCAR homologue) plays an essential role in endosomal sorting through facilitating tubule fission via Arp2/3 activation. Here, we demonstrate a novel function of WASH in modulation of autophagy. We show that WASH deficiency causes...

  6. Cellular and Molecular Connections between Autophagy and Inflammation

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    Pierre Lapaquette

    2015-01-01

    Full Text Available Autophagy is an intracellular catabolic pathway essential for the recycling of proteins and larger substrates such as aggregates, apoptotic corpses, or long-lived and superfluous organelles whose accumulation could be toxic for cells. Because of its unique feature to engulf part of cytoplasm in double-membrane cup-shaped structures, which further fuses with lysosomes, autophagy is also involved in the elimination of host cell invaders and takes an active part of the innate and adaptive immune response. Its pivotal role in maintenance of the inflammatory balance makes dysfunctions of the autophagy process having important pathological consequences. Indeed, defects in autophagy are associated with a wide range of human diseases including metabolic disorders (diabetes and obesity, inflammatory bowel disease (IBD, and cancer. In this review, we will focus on interrelations that exist between inflammation and autophagy. We will discuss in particular how mediators of inflammation can regulate autophagy activity and, conversely, how autophagy shapes the inflammatory response. Impact of genetic polymorphisms in autophagy-related gene on inflammatory bowel disease will be also discussed.

  7. Diabetes and the Brain: Oxidative Stress, Inflammation, and Autophagy

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    María Muriach

    2014-01-01

    Full Text Available Diabetes mellitus is a common metabolic disorder associated with chronic complications including a state of mild to moderate cognitive impairment, in particular psychomotor slowing and reduced mental flexibility, not attributable to other causes, and shares many symptoms that are best described as accelerated brain ageing. A common theory for aging and for the pathogenesis of this cerebral dysfunctioning in diabetes relates cell death to oxidative stress in strong association to inflammation, and in fact nuclear factor κB (NFκB, a master regulator of inflammation and also a sensor of oxidative stress, has a strategic position at the crossroad between oxidative stress and inflammation. Moreover, metabolic inflammation is, in turn, related to the induction of various intracellular stresses such as mitochondrial oxidative stress, endoplasmic reticulum (ER stress, and autophagy defect. In parallel, blockade of autophagy can relate to proinflammatory signaling via oxidative stress pathway and NFκB-mediated inflammation.

  8. Inflammation, Autophagy, and Obesity: Common Features in the Pathogenesis of Pancreatitis and Pancreatic Cancer

    OpenAIRE

    Gukovsky, Ilya; Ning LI; Todoric, Jelena; Gukovskaya, Anna; Karin, Michael

    2013-01-01

    Inflammation and autophagy are cellular defense mechanisms. When these processes are deregulated (deficient or overactivated) they produce pathologic effects, such as oxidative stress, metabolic impairments, and cell death. Unresolved inflammation and disrupted regulation of autophagy are common features of pancreatitis and pancreatic cancer. Furthermore, obesity, a risk factor for pancreatitis and pancreatic cancer, promotes inflammation and inhibits or deregulates autophagy, creating an env...

  9. Protein breakdown in muscle wasting: Role of autophagy-lysosome and ubiquitin-proteasome

    OpenAIRE

    Sandri, Marco

    2013-01-01

    Skeletal muscle adapts its mass as consequence of physical activity, metabolism and hormones. Catabolic conditions or inactivity induce signaling pathways that regulate the process of muscle loss. Muscle atrophy in adult tissue occurs when protein degradation rates exceed protein synthesis. Two major protein degradation pathways, the ubiquitin-proteasome and the autophagy-lysosome systems, are activated during muscle atrophy and variably contribute to the loss of muscle mass. These degradatio...

  10. B cell autophagy mediates TLR7-dependent autoimmunity and inflammation.

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    Weindel, Chi G; Richey, Lauren J; Bolland, Silvia; Mehta, Abhiruchi J; Kearney, John F; Huber, Brigitte T

    2015-01-01

    Systemic lupus erythematosus (SLE) is a heterogeneous autoimmune disease, defined by loss of B cell self-tolerance that results in production of antinuclear antibodies (ANA) and chronic inflammation. While the initiating events in lupus development are not well defined, overexpression of the RNA-recognizing toll-like receptor (TLR)7 has been linked to SLE in humans and mice. We postulated that autophagy plays an essential role in TLR7 activation of B cells for the induction of SLE by delivering RNA ligands to the endosomes, where this innate immune receptor resides. To test this hypothesis, we compared SLE development in Tlr7 transgenic (Tg) mice with or without B cell-specific ablation of autophagy (Cd19-Cre Atg5(f/f)). We observed that in the absence of B cell autophagy the 2 hallmarks of SLE, ANA and inflammation, were eliminated, thus curing these mice of lupus. This was also evident in the significantly extended survival of the autophagy-deficient mice compared to Tlr7.1 Tg mice. Furthermore, glomerulonephritis was ameliorated, and the serum levels of inflammatory cytokines in the knockout (KO) mice were indistinguishable from those of control mice. These data provide direct evidence that B cells require TLR7-dependent priming through an autophagy-dependent mechanism before autoimmunity is induced, thereafter involving many cell types. Surprisingly, hyper-IgM production persisted in Tlr7.1 Tg mice in the absence of autophagy, likely involving a different activation pathway than the production of autoantibodies. Furthermore, these mice still presented with anemia, but responded with a striking increase in extramedullary hematopoiesis (EMH), possibly due to the absence of pro-inflammatory cytokines.

  11. The Ubiquitin Ligase Siah2 Regulates Obesity-induced Adipose Tissue Inflammation

    OpenAIRE

    Kilroy, Gail; Carter, Lauren E; Newman, Susan; Burk, David H.; Manuel, Justin; Möller, Andreas; Bowtell, David D.; Mynatt, Randall L.; Ghosh, Sujoy; Floyd, Z. Elizabeth

    2015-01-01

    Objective Chronic, low-grade adipose tissue inflammation associated with adipocyte hypertrophy is an important link in the relationship between obesity and insulin resistance. Although ubiquitin ligases regulate inflammatory processes, the role of these enzymes in metabolically driven adipose tissue inflammation is relatively unexplored. Herein, we examined the effect of the ubiquitin ligase Siah2 on obesity-related adipose tissue inflammation. Methods Wild-type and Siah2KO mice were fed a lo...

  12. MARCH2 regulates autophagy by promoting CFTR ubiquitination and degradation and PIK3CA-AKT-MTOR signaling.

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    Xia, Dan; Qu, Liujing; Li, Ge; Hongdu, Beiqi; Xu, Chentong; Lin, Xin; Lou, Yaxin; He, Qihua; Ma, Dalong; Chen, Yingyu

    2016-09-01

    MARCH2 (membrane-associated RING-CH protein 2), an E3 ubiquitin ligase, is mainly associated with the vesicle trafficking. In the present study, for the first time, we demonstrated that MARCH2 negatively regulates autophagy. Our data indicated that overexpression of MARCH2 impaired autophagy, as evidenced by attenuated levels of LC3B-II and impaired degradation of endogenous and exogenous autophagic substrates. By contrast, loss of MARCH2 expression had the opposite effects. In vivo experiments demonstrate that MARCH2 knockout mediated autophagy results in an inhibition of tumorigenicity. Further investigation revealed that the induction of autophagy by MARCH2 deficiency was mediated through the PIK3CA-AKT-MTOR signaling pathway. Additionally, we found that MARCH2 interacts with CFTR (cystic fibrosis transmembrane conductance regulator), promotes the ubiquitination and degradation of CFTR, and inhibits CFTR-mediated autophagy in tumor cells. The functional PDZ domain of MARCH2 is required for the association with CFTR. Thus, our study identified a novel negative regulator of autophagy and suggested that the physical and functional connection between the MARCH2 and CFTR in different conditions will be elucidated in the further experiments. PMID:27308891

  13. Modification by Ubiquitin-Like Proteins: Significance in Apoptosis and Autophagy Pathways

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    Monde Ntwasa

    2012-09-01

    Full Text Available Ubiquitin-like proteins (Ubls confer diverse functions on their target proteins. The modified proteins are involved in various biological processes, including DNA replication, signal transduction, cell cycle control, embryogenesis, cytoskeletal regulation, metabolism, stress response, homeostasis and mRNA processing. Modifiers such as SUMO, ATG12, ISG15, FAT10, URM1, and UFM have been shown to modify proteins thus conferring functions related to programmed cell death, autophagy and regulation of the immune system. Putative modifiers such as Domain With No Name (DWNN have been identified in recent times but not fully characterized. In this review, we focus on cellular processes involving human Ubls and their targets. We review current progress in targeting these modifiers for drug design strategies.

  14. Autophagy resolves early retinal inflammation in Igf1-deficient mice

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    Rodríguez-de la Rosa, Lourdes; Murillo-Cuesta, Silvia; Vaquero-Villanueva, Laura; Hurlé, Juan M.; Varela-Nieto, Isabel; Valverde, Ángela M.

    2016-01-01

    ABSTRACT Insulin-like growth factor-1 (IGF-1) is a growth factor with differentiating, anti-apoptotic and metabolic functions in the periphery, and anti-inflammatory properties in the nervous system. Mice that have mutations in the Igf1 gene, rendering the gene product inactive (Igf1−/−), present with age-related visual loss accompanied by structural alterations in the first synapses of the retinal pathway. Recent advances have revealed a crucial role of autophagy in immunity and inflammation. Keeping in mind this close relationship, we aimed to decipher these processes in the context of the defects that occur during ageing in the retina of Igf1−/− mice. Tnfa and Il1b mRNAs, and phosphorylation of JNK and p38 MAPK were elevated in the retinas of 6- and 12-month old Igf1−/− mice compared to those in age-matched Igf1+/+ controls. In 6-month-old Igf1−/− retinas, increased mRNA levels of the autophagy mediators Becn1, Atg9, Atg5 and Atg4, decreased p62 (also known as SQSTM1) protein expression together with an increased LC3-II:LC3-I ratio reflected active autophagic flux. However, in retinas from 12-month-old Igf1−/− mice, Nlrp3 mRNA, processing of the IL1β pro-form and immunostaining of active caspase-1 were elevated compared to those in age-matched Igf1+/+ controls, suggesting activation of the inflammasome. This effect concurred with accumulation of autophagosomes and decreased autophagic flux in the retina. Microglia localization and status of activation in the retinas of 12-month-old Igf1+/+ and Igf1−/− mice, analyzed by immunostaining of Cd11b and Iba-1, showed a specific distribution pattern in the outer plexiform layer (OPL), inner plexiform layer (IPL) and inner nuclear layer (INL), and revealed an increased number of activated microglia cells in the retina of 12-month-old blind Igf1−/− mice. Moreover, reactive gliosis was exclusively detected in the retinas from 12-month-old blind Igf1−/− mice. In conclusion, this study

  15. The Ubiquitin Ligase XIAP Recruits LUBAC for NOD2 Signaling in Inflammation and Innate Immunity

    DEFF Research Database (Denmark)

    Damgaard, Rune Busk; Nachbur, Ueli; Yabal, Monica;

    2012-01-01

    -linked lymphoproliferative syndrome type-2 (XLP-2). Here, we demonstrate that the RING domain of XIAP is essential for NOD2 signaling and that XIAP contributes to exacerbation of inflammation-induced hepatitis in experimental mice. We find that XIAP ubiquitylates RIPK2 and recruits the linear ubiquitin chain assembly...... signaling. We conclude that XIAP and LUBAC constitute essential ubiquitin ligases in NOD2-mediated inflammatory signaling and propose that deregulation of NOD2 signaling contributes to XLP-2 pathogenesis....

  16. Critical role for IL-18 in spontaneous lung inflammation caused by autophagy deficiency.

    Science.gov (United States)

    Abdel Fattah, Elmoataz; Bhattacharya, Abhisek; Herron, Alan; Safdar, Zeenat; Eissa, N Tony

    2015-06-01

    Autophagy is an important component of the immune response. However, the functions of autophagy in human diseases are much less understood. We studied biological consequences of autophagy deficiency in mice lacking the essential autophagy gene Atg7 or Atg5 in myeloid cells. Surprisingly, these mice presented with spontaneous sterile lung inflammation, characterized by marked recruitment of inflammatory cells, submucosal thickening, goblet cell metaplasia, and increased collagen content. Lung inflammation was associated with increase in several proinflammatory cytokines in the bronchoalveolar lavage and in serum. This inflammation was largely driven by IL-18 as a result of constitutive inflammasome activation. Following i.p. LPS injection, autophagy-deficient mice had higher levels of proinflammatory cytokines in lungs and in serum, as well as increased mortality, than control mice. Intranasal bleomycin challenge exacerbated lung inflammation in autophagy-deficient mice and produced more severe fibrotic changes than in control mice. These results uncover a new and important role for autophagy as negative regulator of lung inflammation.

  17. Modulation of inflammation by autophagy: consequences for Crohn's disease.

    NARCIS (Netherlands)

    Plantinga, T.S.; Joosten, L.A.B.; Meer, J.W.M. van der; Netea, M.G.

    2012-01-01

    Autophagy, the cellular machinery for targeting intracellular components for lysosomal degradation, is critically involved in the host defence to pathogenic microorganisms. Recent studies have unveiled several aspects of the immune response that are regulated by autophagy, including antigen presenta

  18. SIRT1 inactivation induces inflammation through the dysregulation of autophagy in human THP-1 cells

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    Takeda-Watanabe, Ai; Kitada, Munehiro; Kanasaki, Keizo [Diabetology and Endocrinology, Kanazawa Medical University, Kahoku-Gun, Ishikawa (Japan); Koya, Daisuke, E-mail: koya0516@kanazawa-med.ac.jp [Diabetology and Endocrinology, Kanazawa Medical University, Kahoku-Gun, Ishikawa (Japan)

    2012-10-12

    Highlights: Black-Right-Pointing-Pointer SIRT1 inactivation decreases autophagy in THP-1 cell. Black-Right-Pointing-Pointer Inhibition of autophagy induces inflammation. Black-Right-Pointing-Pointer SIRT1 inactivation induces inflammation through NF-{kappa}B activation. Black-Right-Pointing-Pointer The p62/Sqstm1 accumulation by impairment of autophagy is related to NF-{kappa}B activation. Black-Right-Pointing-Pointer SIRT1 inactivation is involved in the activation of mTOR and decreased AMPK activation. -- Abstract: Inflammation plays a crucial role in atherosclerosis. Monocytes/macrophages are some of the cells involved in the inflammatory process in atherogenesis. Autophagy exerts a protective effect against cellular stresses like inflammation, and it is regulated by nutrient-sensing pathways. The nutrient-sensing pathway includes SIRT1, a NAD{sup +}-dependent histone deacetylase, which is implicated in the regulation of a variety of cellular processes including inflammation and autophagy. The mechanism through which the dysfunction of SIRT1 contributes to the regulation of inflammation in relation to autophagy in monocytes/macrophages is unclear. In the present study, we demonstrate that treatment with 2-[(2-Hydroxynaphthalen-1-ylmethylene)amino]-N-(1-phenethyl)benzamide (Sirtinol), a chemical inhibitor of SIRT1, induces the overexpression of inflammation-related genes such as tumor necrosis factor (TNF)-{alpha} and interleukin (IL)-6 through nuclear factor (NF)-{kappa}B signaling activation, which is associated with autophagy dysfunction, as shown through p62/Sqstm1 accumulation and decreased expression of light chain (LC) 3 II in THP-1 cells. The autophagy inhibitor, 3-methyladenine, also induces inflammation-related NF-{kappa}B activation. In p62/Sqstm1 knockdown cells, Sirtinol-induced inflammation through NF-{kappa}B activation is blocked. In addition, inhibition of SIRT1 is involved in the activation of the mammalian target of rapamycin (mTOR) pathway and

  19. Autophagy in periodontitis patients and gingival fibroblasts: unraveling the link between chronic diseases and inflammation

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    Bullon Pedro

    2012-10-01

    Full Text Available Abstract Background Periodontitis, the most prevalent chronic inflammatory disease, has been related to cardiovascular diseases. Autophagy provides a mechanism for the turnover of cellular organelles and proteins through a lysosome-dependent degradation pathway. The aim of this research was to study the role of autophagy in peripheral blood mononuclear cells from patients with periodontitis and gingival fibroblasts treated with a lipopolysaccharide of Porphyromonas gingivalis. Autophagy-dependent mechanisms have been proposed in the pathogenesis of inflammatory disorders and in other diseases related to periodontitis, such as cardiovascular disease and diabetes. Thus it is important to study the role of autophagy in the pathophysiology of periodontitis. Methods Peripheral blood mononuclear cells from patients with periodontitis (n = 38 and without periodontitis (n = 20 were used to study autophagy. To investigate the mechanism of autophagy, we evaluated the influence of a lipopolysaccharide from P. gingivalis in human gingival fibroblasts, and autophagy was monitored morphologically and biochemically. Autophagosomes were observed by immunofluorescence and electron microscopy. Results We found increased levels of autophagy gene expression and high levels of mitochondrial reactive oxygen species production in peripheral blood mononuclear cells from patients with periodontitis compared with controls. A significantly positive correlation between both was observed. In human gingival fibroblasts treated with lipopolysaccharide from P. gingivalis, there was an increase of protein and transcript of autophagy-related protein 12 (ATG12 and microtubule-associated protein 1 light chain 3 alpha LC3. A reduction of mitochondrial reactive oxygen species induced a decrease in autophagy whereas inhibition of autophagy in infected cells increased apoptosis, showing the protective role of autophagy. Conclusion Results from the present study suggest that autophagy

  20. The ubiquitin signal and autophagy: an orchestrated dance leading to mitochondrial degradation.

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    Yamano, Koji; Matsuda, Noriyuki; Tanaka, Keiji

    2016-03-01

    The quality of mitochondria, essential organelles that produce ATP and regulate numerous metabolic pathways, must be strictly monitored to maintain cell homeostasis. The loss of mitochondrial quality control systems is acknowledged as a determinant for many types of neurodegenerative diseases including Parkinson's disease (PD). The two gene products mutated in the autosomal recessive forms of familial early-onset PD, Parkin and PINK1, have been identified as essential proteins in the clearance of damaged mitochondria via an autophagic pathway termed mitophagy. Recently, significant progress has been made in understanding how the mitochondrial serine/threonine kinase PINK1 and the E3 ligase Parkin work together through a novel stepwise cascade to identify and eliminate damaged mitochondria, a process that relies on the orchestrated crosstalk between ubiquitin/phosphorylation signaling and autophagy. In this review, we highlight our current understanding of the detailed molecular mechanisms governing Parkin-/PINK1-mediated mitophagy and the evidences connecting Parkin/PINK1 function and mitochondrial clearance in neurons. PMID:26882551

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

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    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); Kang, Seok-Seong, E-mail: sskang@ivi.int [Laboratory Sciences Division, International Vaccine Institute, Seoul 151-919 (Korea, Republic of)

    2011-01-14

    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.

  2. Itch inhibits IL-17-mediated colon inflammation and tumorigenesis by ROR-γt ubiquitination.

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    Kathania, Mahesh; Khare, Prashant; Zeng, Minghui; Cantarel, Brandi; Zhang, Haiying; Ueno, Hideki; Venuprasad, K

    2016-08-01

    Dysregulated expression of interleukin 17 (IL-17) in the colonic mucosa is associated with colonic inflammation and cancer. However, the cell-intrinsic molecular mechanisms by which IL-17 expression is regulated remain unclear. We found that deficiency in the ubiquitin ligase Itch led to spontaneous colitis and increased susceptibility to colon cancer. Itch deficiency in the TH17 subset of helper T cells, innate lymphoid cells and γδ T cells resulted in the production of elevated amounts of IL-17 in the colonic mucosa. Mechanistically, Itch bound to the transcription factor ROR-γt and targeted ROR-γt for ubiquitination. Inhibition or genetic inactivation of ROR-γt attenuated IL-17 expression and reduced spontaneous colonic inflammation in Itch(-/-) mice. Thus, we have identified a previously unknown role for Itch in regulating IL-17-mediated colonic inflammation and carcinogenesis. PMID:27322655

  3. Autophagy is essential for ultrafine particle-induced inflammation and mucus hyperproduction in airway epithelium.

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    Chen, Zhi-Hua; Wu, Yin-Fang; Wang, Ping-Li; Wu, Yan-Ping; Li, Zhou-Yang; Zhao, Yun; Zhou, Jie-Sen; Zhu, Chen; Cao, Chao; Mao, Yuan-Yuan; Xu, Feng; Wang, Bei-Bei; Cormier, Stephania A; Ying, Song-Min; Li, Wen; Shen, Hua-Hao

    2016-01-01

    Environmental ultrafine particulate matter (PM) is capable of inducing airway injury, while the detailed molecular mechanisms remain largely unclear. Here, we demonstrate pivotal roles of autophagy in regulation of inflammation and mucus hyperproduction induced by PM containing environmentally persistent free radicals in human bronchial epithelial (HBE) cells and in mouse airways. PM was endocytosed by HBE cells and simultaneously triggered autophagosomes, which then engulfed the invading particles to form amphisomes and subsequent autolysosomes. Genetic blockage of autophagy markedly reduced PM-induced expression of inflammatory cytokines, e.g. IL8 and IL6, and MUC5AC in HBE cells. Mice with impaired autophagy due to knockdown of autophagy-related gene Becn1 or Lc3b displayed significantly reduced airway inflammation and mucus hyperproduction in response to PM exposure in vivo. Interference of the autophagic flux by lysosomal inhibition resulted in accumulated autophagosomes/amphisomes, and intriguingly, this process significantly aggravated the IL8 production through NFKB1, and markedly attenuated MUC5AC expression via activator protein 1. These data indicate that autophagy is required for PM-induced airway epithelial injury, and that inhibition of autophagy exerts therapeutic benefits for PM-induced airway inflammation and mucus hyperproduction, although they are differentially orchestrated by the autophagic flux.

  4. The Differential Profiling of Ubiquitin-Proteasome and Autophagy Systems in Different Tissues before the Onset of Huntington's Disease Models.

    Science.gov (United States)

    Her, Lu-Shiun; Lin, Jian-Yu; Fu, Mu-Hui; Chang, Yu-Fan; Li, Chia-Ling; Tang, Ting-Yu; Jhang, Yu-Ling; Chang, Chih-Yi; Shih, Meng-Chi; Cheng, Pei-Hsun; Yang, Shang-Hsun

    2015-07-01

    Huntington's disease (HD) is a genetic and neurodegenerative disease, leading to motor and cognitive dysfunction in HD patients. At cellular level, this disease is caused by the accumulation of mutant huntingtin (HTT) in different cells, and finally results in the dysfunction of different cells. To clean these mutant proteins, ubiquitin-proteasome system (UPS) and autophagy system are two critical pathways in the brain; however, little is known in other peripheral tissues. As mutant HTT affects different tissues progressively and might influence the UPS and autophagy pathways at early stages, we attempted to examine two clearance systems in HD models before the onset. Here, in vitro results showed that the accumulation of UPS signals with time was observed obviously in neuroblastoma and kidney cells, not in other cells. In HD transgenic mice, we observed the impairment of UPS, but not autophagy, over time in the cortex and striatum. In heart and muscle tissues, disturbance of autophagy was observed, whereas dysfunction of UPS was displayed in liver and lung. These results suggest that two protein clearance pathways are disturbed differentially in different tissues before the onset of HD, and enhancement of protein clearance at early stages might provide a potential stratagem to alleviate the progression of HD. PMID:25178567

  5. The Effect of Autophagy on Inflammation Cytokines in Renal Ischemia/Reperfusion Injury.

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    Ling, Haibin; Chen, Hongguang; Wei, Miao; Meng, Xiaoyin; Yu, Yonghao; Xie, Keliang

    2016-02-01

    Acute kidney injury (AKI) is characterized by a rapid loss of kidney function and an antigen-independent inflammatory process that causes tissue damage, which was one of the main manifestations of kidney ischemia/reperfusion (I/R). Recent studies have demonstrated autophagy participated in the pathological process of acute kidney injury. In this study, we discuss how autophagy regulated inflammation response in the kidney I/R. AKI was performed by renal I/R. Autophagy activator rapamycin (Rap) and inhibitor 3-methyladenine (MA) were used to investigate the role of autophagy on kidney function and inflammation response. After the experiment, kidney tissues were obtained for the detection of autophagy-related protein microtubule-associated protein light chain 3(LC3)II, Beclin1, and Rab7 and lysosome-associated membrane protein type (LAMP)2 protein by reverse transcription-polymerase chain reaction (PT-PCR) and Western blotting, and histopathology and tissue injury scores also. The blood was harvested to measure kidney function (creatinine (Cr) and blood urea nitrogen (BUN) levels) after I/R. Cytokines TNF-α, IL-6, HMGB1, and IL-10 were measured after I/R. I/R induced the expression of LC3II, Beclin1, LAMP2, and Rab7. The activation and inhibition of autophagy by rapamycin and 3-MA were promoted and attenuated histological and renal function in renal I/R rats, respectively. Cytokines TNF-α, IL-6, and HMGB1 were decreased, and IL-10 was further increased after activation of autophagy treated in I/R rats, while 3-MA exacerbated the pro-inflammatory cytokines TNF-α, IL-6, HMGB1, and anti-inflammatory cytokine IL-10 in renal I/R. I/R can activated the autophagy, and autophagy increase mitigated the renal injury by decreasing kidney injury score, levels of Cr and BUN after renal I/R, and inflammation response via regulating the balance of pro-inflammation and anti-inflammation cytokines.

  6. Autophagy in infection.

    Science.gov (United States)

    Deretic, Vojo

    2010-04-01

    Autophagy is a ubiquitous eukaryotic cytoplasmic quality and quantity control pathway. The role of autophagy in cytoplasmic homeostasis seamlessly extends to cell-autonomous defense against intracellular microbes. Recent studies also point to fully integrated, multitiered regulatory and effector connections between autophagy and nearly all facets of innate and adaptive immunity. Autophagy in the immune system as a whole confers measured immune responses; on the flip side, suppression of autophagy can lead to inflammation and tissue damage, as evidenced by Crohn's disease predisposition polymorphisms in autophagy basal apparatus (Atg16L) and regulatory (IRGM) genes. Polymorphisms in the IRGM gene in human populations have also been linked to predisposition to tuberculosis. There are several areas of most recent growth: first, links between autophagy regulators and infectious disease predisposition in human populations; second, demonstration of a role for autophagy in infection control in vivo in animal models; third, the definition of specific antiautophagic defenses in highly evolved pathogens; and fourth, recognition of connections between the ubiquitin system and autophagy of bacteria (and interestingly mitochondria, which are incidentally organelles of bacterial evolutionary origin) via a growing list of modifier and adapter proteins including p62/SQSTM1, NDP52, Atg32, Parkin, and Nix/BNIP3L. PMID:20116986

  7. Autophagy facilitates TLR4- and TLR3-triggered migration and invasion of lung cancer cells through the promotion of TRAF6 ubiquitination.

    Science.gov (United States)

    Zhan, Zhenzhen; Xie, Xuefeng; Cao, Hao; Zhou, Xiaohui; Zhang, Xu Dong; Fan, Huimin; Liu, Zhongmin

    2014-02-01

    Autophagy contributes to the pathogenesis of cancer, whereas toll-like receptors (TLRs) also play an important role in cancer development and immune escape. However, little is known about the potential interaction between TLR signaling and autophagy in cancer cells. Here we show that autophagy induced by TLR4 or TLR3 activation enhances various cytokine productions through promoting TRAF6 (TNF receptor-associated factor 6, E3 ubiquitin protein ligase) ubiquitination and thus facilitates migration and invasion of lung cancer cells. Stimulation of TLR4 and TLR3 with lipopolysaccharide (LPS) and polyinosinic-polycytidylic acid [poly(I:C)] respectively triggered autophagy in lung cancer cells. This was mediated by the adaptor protein, toll-like receptor adaptor molecule 1 (TICAM1/TRIF), and was required for TLR4- and TLR3-induced increases in the production of IL6, CCL2/MCP-1 [chemokine (C-C motif) ligand 2], CCL20/MIP-3α [chemokine (C-C motif) ligand 20], VEGFA (vascular endothelial growth factor A), and MMP2 [matrix metallopeptidase 2 (gelatinase A, 72 kDa gelatinase, 72 kDa type IV collagenase)]. These cytokines appeared to be necessary for enhanced migration and invasion of lung cancer cells upon TLR activation. Remarkably, inhibition of autophagy by chemical or genetic approaches blocked TLR4- or TLR3-induced Lys63 (K63)-linked ubiquitination of TRAF6 that was essential for activation of MAPK and NFKB (nuclear factor of kappa light polypeptide gene enhancer in B-cells) pathways, both of which were involved in the increased production of the cytokines. Collectively, these results identify induction of autophagy by TLR4 and TLR3 as an important mechanism that drives lung cancer progression, and indicate that inhibition of autophagy may be a useful strategy in the treatment of lung cancer.

  8. Autophagy in Macrophages: Impacting Inflammation and Bacterial Infection

    Directory of Open Access Journals (Sweden)

    Ali Vural

    2014-01-01

    Full Text Available Macrophages are on the front line of host defense. They possess an array of germline-encoded pattern recognition receptors/sensors (PRRs that recognize pathogen-associated molecular patterns (PAMPs and which activate downstream effectors/pathways to help mediate innate immune responses and host defense. Innate immune responses include the rapid induction of transcriptional networks that trigger the production of cytokines, chemokines, and cytotoxic molecules; the mobilization of cells including neutrophils and other leukocytes; the engulfment of pathogens by phagocytosis and their delivery to lysosome for degradation; and the induction of autophagy. Autophagy is a catabolic process that normally maintains cellular homeostasis in a lysosome-dependent manner, but it also functions as a cytoprotective response that intersects with a variety of general stress-response pathways. This review focuses on the intimately linked molecular mechanisms that help govern the autophagic pathway and macrophage innate immune responses.

  9. Autophagy, inflammation and innate immunity in inflammatory myopathies.

    Directory of Open Access Journals (Sweden)

    Cristina Cappelletti

    Full Text Available Autophagy has a large range of physiological functions and its dysregulation contributes to several human disorders, including autoinflammatory/autoimmune diseases such as inflammatory myopathies (IIMs. In order to better understand the pathogenetic mechanisms of these muscular disorders, we sought to define the role of autophagic processes and their relation with the innate immune system in the three main subtypes of IIM, specifically sporadic inclusion body myositis (sIBM, polymyositis (PM, dermatomyositis (DM and juvenile dermatomyositis (JDM. We found that although the mRNA transcript levels of the autophagy-related genes BECN1, ATG5 and FBXO32 were similar in IIM and controls, autophagy activation in all IIM subgroups was suggested by immunoblotting results and confirmed by immunofluorescence. TLR4 and TLR3, two potent inducers of autophagy, were highly increased in IIM, with TLR4 transcripts significantly more expressed in PM and DM than in JDM, sIBM and controls, and TLR3 transcripts highly up-regulated in all IIM subgroups compared to controls. Co-localization between autophagic marker, LC3, and TLR4 and TLR3 was observed not only in sIBM but also in PM, DM and JDM muscle tissues. Furthermore, a highly association with the autophagic processes was observed in all IIM subgroups also for some TLR4 ligands, endogenous and bacterial HSP60, other than the high-mobility group box 1 (HMGB1. These findings indicate that autophagic processes are active not only in sIBM but also in PM, DM and JDM, probably in response to an exogenous or endogenous 'danger signal'. However, autophagic activation and regulation, and also interaction with the innate immune system, differ in each type of IIM. Better understanding of these differences may lead to new therapies for the different IIM types.

  10. Ubiquitin

    DEFF Research Database (Denmark)

    Vinther-Jensen, T.; Simonsen, A. H.; Budtz-Jorgensen, E.;

    2015-01-01

    BACKGROUND: Finding early and dynamic biomarkers in Huntington's disease is a key to understanding the early pathology of Huntington's disease and potentially to tracking disease progression. This would benefit the future evaluation of potential neuroprotective and disease-modifying therapies......, as well as aid in identifying an optimal time point for initiating a potential therapeutic intervention. METHODS: This explorative proteomics study evaluated cerebrospinal fluid from 94 Huntington's disease gene-expansion carriers (39 premanifest and 55 manifest) and 27 Huntington's disease gene...... and controls. One of them identified as ubiquitin was shown to be dependent on the Unified Huntington Disease Rating Scale Total Functional Capacity, a pseudo-measure of disease severity (P = 0.001), and the Symbol Digit Modalities Test (0.04) in manifest and CAG-age product score (P = 0.019) in all gene...

  11. Magnetic ferroferric oxide nanoparticles induce vascular endothelial cell dysfunction and inflammation by disturbing autophagy.

    Science.gov (United States)

    Zhang, Lu; Wang, XueQin; Miao, YiMing; Chen, ZhiQiang; Qiang, PengFei; Cui, LiuQing; Jing, Hongjuan; Guo, YuQi

    2016-03-01

    Despite the considerable use of magnetic ferroferric oxide nanoparticles (Fe3O4NPs) worldwide, their safety is still an important topic of debate. In the present study, we detected the toxicity and biological behavior of bare-Fe3O4NPs (B-Fe3O4NPs) on human umbilical vascular endothelial cells (HUVECs). Our results showed that B-Fe3O4NPs did not induce cell death within 24h even at concentrations up to 400 μg/ml. The level of nitric oxide (NO) and the activity of endothelial NO synthase (eNOS) were decreased after exposure to B-Fe3O4NPs, whereas the levels of proinflammatory cytokines were elevated. Importantly, B-Fe3O4NPs increased the accumulation of autophagosomes and LC3-II in HUVECs through both autophagy induction and the blockade of autophagy flux. The levels of Beclin 1 and VPS34, but not phosphorylated mTOR, were increased in the B-Fe3O4NP-treated HUVECs. Suppression of autophagy induction or stimulation of autophagy flux, at least partially, attenuated the B-Fe3O4NP-induced HUVEC dysfunction. Additionally, enhanced autophagic activity might be linked to the B-Fe3O4NP-induced production of proinflammatory cytokines. Taken together, these results demonstrated that B-Fe3O4NPs disturb the process of autophagy in HUVECs, and eventually lead to endothelial dysfunction and inflammation.

  12. The autophagy gene Atg16l1 differentially regulates Treg and TH2 cells to control intestinal inflammation.

    Science.gov (United States)

    Kabat, Agnieszka M; Harrison, Oliver J; Riffelmacher, Thomas; Moghaddam, Amin E; Pearson, Claire F; Laing, Adam; Abeler-Dörner, Lucie; Forman, Simon P; Grencis, Richard K; Sattentau, Quentin; Simon, Anna Katharina; Pott, Johanna; Maloy, Kevin J

    2016-01-01

    A polymorphism in the autophagy gene Atg16l1 is associated with susceptibility to inflammatory bowel disease (IBD); however, it remains unclear how autophagy contributes to intestinal immune homeostasis. Here, we demonstrate that autophagy is essential for maintenance of balanced CD4(+) T cell responses in the intestine. Selective deletion of Atg16l1 in T cells in mice resulted in spontaneous intestinal inflammation that was characterized by aberrant type 2 responses to dietary and microbiota antigens, and by a loss of Foxp3(+) Treg cells. Specific ablation of Atg16l1 in Foxp3(+) Treg cells in mice demonstrated that autophagy directly promotes their survival and metabolic adaptation in the intestine. Moreover, we also identify an unexpected role for autophagy in directly limiting mucosal TH2 cell expansion. These findings provide new insights into the reciprocal control of distinct intestinal TH cell responses by autophagy, with important implications for understanding and treatment of chronic inflammatory disorders. PMID:26910010

  13. Mycobacterium tuberculosis eis regulates autophagy, inflammation, and cell death through redox-dependent signaling.

    Directory of Open Access Journals (Sweden)

    Dong-Min Shin

    Full Text Available The "enhanced intracellular survival" (eis gene of Mycobacterium tuberculosis (Mtb is involved in the intracellular survival of M. smegmatis. However, its exact effects on host cell function remain elusive. We herein report that Mtb Eis plays essential roles in modulating macrophage autophagy, inflammatory responses, and cell death via a reactive oxygen species (ROS-dependent pathway. Macrophages infected with an Mtb eis-deletion mutant H37Rv (Mtb-Δeis displayed markedly increased accumulation of massive autophagic vacuoles and formation of autophagosomes in vitro and in vivo. Infection of macrophages with Mtb-Δeis increased the production of tumor necrosis factor-α and interleukin-6 over the levels produced by infection with wild-type or complemented strains. Elevated ROS generation in macrophages infected with Mtb-Δeis (for which NADPH oxidase and mitochondria were largely responsible rendered the cells highly sensitive to autophagy activation and cytokine production. Despite considerable activation of autophagy and proinflammatory responses, macrophages infected with Mtb-Δeis underwent caspase-independent cell death. This cell death was significantly inhibited by blockade of autophagy and c-Jun N-terminal kinase-ROS signaling, suggesting that excessive autophagy and oxidative stress are detrimental to cell survival. Finally, artificial over-expression of Eis or pretreatment with recombinant Eis abrogated production of both ROS and proinflammatory cytokines, which depends on the N-acetyltransferase domain of the Eis protein. Collectively, these data indicate that Mtb Eis suppresses host innate immune defenses by modulating autophagy, inflammation, and cell death in a redox-dependent manner.

  14. Inhibition of autophagy ameliorates atherogenic inflammation by augmenting apigenin-induced macrophage apoptosis.

    Science.gov (United States)

    Wang, Qun; Zeng, Ping; Liu, Yuanliang; Wen, Ge; Fu, Xiuqiong; Sun, Xuegang

    2015-07-01

    Increasing evidences showed that the survival of macrophages promotes atherogenesis. Macrophage apoptosis in the early phase of atherosclerotic process negatively regulates the progression of atherosclerotic lesions. We demonstrated that a natural anti-oxidant apigenin could ameliorate atherogenesis in ApoE(-/-) mice. It reduced the number of foam cells and decreased the serum levels of tumor necrosis factor α, interleukin 1β (IL-1β) and IL-6. Our results showed that oxidized low-density lipoprotein (oxLDL) led to the secretion of pro-inflammatory cytokines. Apigenin-induced apoptosis and downregulated the secretion of TNF-α, IL-6 and IL-1β. It is further supported by the use of zVAD, a pan-caspase inhibitor, demonstrating that apigenin lowered cytokine profile through induction of macrophage apoptosis. Moreover, apigenin-induced Atg5/Atg7-dependent autophagy in macrophages pretreated with oxLDL. Results illustrated that autophagy inhibition increased apigenin-induced apoptosis through activation of Bax. The present findings suggest that oxLDL maintained the survival of macrophages and activated the secretion of pro-inflammatory cytokines to initiate atherosclerosis. Apigenin-induced apoptosis of lipid-laden macrophages and resolved inflammation to ameliorate atherosclerosis. In conclusion, combination of apigenin with autophagy inhibition may be a promising strategy to induce foam cell apoptosis and subdue atherogenic cytokines.

  15. Mitochondrial DNA that escapes from autophagy causes inflammation and heart failure.

    Science.gov (United States)

    Oka, Takafumi; Hikoso, Shungo; Yamaguchi, Osamu; Taneike, Manabu; Takeda, Toshihiro; Tamai, Takahito; Oyabu, Jota; Murakawa, Tomokazu; Nakayama, Hiroyuki; Nishida, Kazuhiko; Akira, Shizuo; Yamamoto, Akitsugu; Komuro, Issei; Otsu, Kinya

    2012-05-10

    Heart failure is a leading cause of morbidity and mortality in industrialized countries. Although infection with microorganisms is not involved in the development of heart failure in most cases, inflammation has been implicated in the pathogenesis of heart failure. However, the mechanisms responsible for initiating and integrating inflammatory responses within the heart remain poorly defined. Mitochondria are evolutionary endosymbionts derived from bacteria and contain DNA similar to bacterial DNA. Mitochondria damaged by external haemodynamic stress are degraded by the autophagy/lysosome system in cardiomyocytes. Here we show that mitochondrial DNA that escapes from autophagy cell-autonomously leads to Toll-like receptor (TLR) 9-mediated inflammatory responses in cardiomyocytes and is capable of inducing myocarditis and dilated cardiomyopathy. Cardiac-specific deletion of lysosomal deoxyribonuclease (DNase) II showed no cardiac phenotypes under baseline conditions, but increased mortality and caused severe myocarditis and dilated cardiomyopathy 10 days after treatment with pressure overload. Early in the pathogenesis, DNase II-deficient hearts showed infiltration of inflammatory cells and increased messenger RNA expression of inflammatory cytokines, with accumulation of mitochondrial DNA deposits in autolysosomes in the myocardium. Administration of inhibitory oligodeoxynucleotides against TLR9, which is known to be activated by bacterial DNA, or ablation of Tlr9 attenuated the development of cardiomyopathy in DNase II-deficient mice. Furthermore, Tlr9 ablation improved pressure overload-induced cardiac dysfunction and inflammation even in mice with wild-type Dnase2a alleles. These data provide new perspectives on the mechanism of genesis of chronic inflammation in failing hearts.

  16. Autophagy Genes Enhance Murine Gammaherpesvirus 68 Reactivation from Latency by Preventing Virus-Induced Systemic Inflammation.

    Science.gov (United States)

    Park, Sunmin; Buck, Michael D; Desai, Chandni; Zhang, Xin; Loginicheva, Ekaterina; Martinez, Jennifer; Freeman, Michael L; Saitoh, Tatsuya; Akira, Shizuo; Guan, Jun-Lin; He, You-Wen; Blackman, Marcia A; Handley, Scott A; Levine, Beth; Green, Douglas R; Reese, Tiffany A; Artyomov, Maxim N; Virgin, Herbert W

    2016-01-13

    Host genes that regulate systemic inflammation upon chronic viral infection are incompletely understood. Murine gammaherpesvirus 68 (MHV68) infection is characterized by latency in macrophages, and reactivation is inhibited by interferon-γ (IFN-γ). Using a lysozyme-M-cre (LysMcre) expression system, we show that deletion of autophagy-related (Atg) genes Fip200, beclin 1, Atg14, Atg16l1, Atg7, Atg3, and Atg5, in the myeloid compartment, inhibited MHV68 reactivation in macrophages. Atg5 deficiency did not alter reactivation from B cells, and effects on reactivation from macrophages were not explained by alterations in productive viral replication or the establishment of latency. Rather, chronic MHV68 infection triggered increased systemic inflammation, increased T cell production of IFN-γ, and an IFN-γ-induced transcriptional signature in macrophages from Atg gene-deficient mice. The Atg5-related reactivation defect was partially reversed by neutralization of IFN-γ. Thus Atg genes in myeloid cells dampen virus-induced systemic inflammation, creating an environment that fosters efficient MHV68 reactivation from latency. PMID:26764599

  17. Inhalation of titanium dioxide induces endoplasmic reticulum stress-mediated autophagy and inflammation in mice.

    Science.gov (United States)

    Yu, Kyeong-Nam; Sung, Jae Hyuck; Lee, Somin; Kim, Ji-Eun; Kim, Sanghwa; Cho, Won-Young; Lee, Ah Young; Park, Soo Jin; Lim, Joohyun; Park, Changhoon; Chae, Chanhee; Lee, Jin Kyu; Lee, Jinkyu; Kim, Jun-Sung; Cho, Myung-Haing

    2015-11-01

    Titanium dioxide (TiO2) nanoparticles are widely used in cosmetics, sunscreen, electronics, drug delivery systems, and diverse bio-application fields. In the workplace, the primary exposure route for TiO2 nanoparticles is inhalation through the respiratory system. Because TiO2 nanoparticles have different physiological properties, in terms of size and bioactivity, their toxic effects in the respiratory system must be determined. In this study, to determine the toxic effect of inhaled TiO2 nanoparticles in the lung and the underlying mechanism, we used a whole-body chamber inhalation system to expose A/J mice to TiO2 nanoparticles for 28 days. During the experiments, the inhaled TiO2 nanoparticles were characterized using a cascade impactor and transmission electron microscopy. After inhalation of the TiO2 nanoparticles, hyperplasia and inflammation were observed in a TiO2 dose-dependent manner. To determine the biological mechanism of the toxic response in the lung, we examined endoplasmic reticulum (ER) and mitochondria in lung. The ER and mitochondria were disrupted and dysfunctional in the TiO2-exposed lung leading to abnormal autophagy. In summary, we assessed the potential risk of TiO2 nanoparticles in the respiratory system, which contributed to our understanding of the mechanism underlining TiO2 nanoparticle toxicity in the lung.

  18. Defective CFTR induces aggresome formation and lung inflammation in cystic fibrosis through ROS-mediated autophagy inhibition.

    Science.gov (United States)

    Luciani, Alessandro; Villella, Valeria Rachela; Esposito, Speranza; Brunetti-Pierri, Nicola; Medina, Diego; Settembre, Carmine; Gavina, Manuela; Pulze, Laura; Giardino, Ida; Pettoello-Mantovani, Massimo; D'Apolito, Maria; Guido, Stefano; Masliah, Eliezer; Spencer, Brian; Quaratino, Sonia; Raia, Valeria; Ballabio, Andrea; Maiuri, Luigi

    2010-09-01

    Accumulation of unwanted/misfolded proteins in aggregates has been observed in airways of patients with cystic fibrosis (CF), a life-threatening genetic disorder caused by mutations in the gene encoding the cystic fibrosis transmembrane conductance regulator (CFTR). Here we show how the defective CFTR results in defective autophagy and decreases the clearance of aggresomes. Defective CFTR-induced upregulation of reactive oxygen species (ROS) and tissue transglutaminase (TG2) drive the crosslinking of beclin 1, leading to sequestration of phosphatidylinositol-3-kinase (PI(3)K) complex III and accumulation of p62, which regulates aggresome formation. Both CFTR knockdown and the overexpression of green fluorescent protein (GFP)-tagged-CFTR(F508del) induce beclin 1 downregulation and defective autophagy in non-CF airway epithelia through the ROS-TG2 pathway. Restoration of beclin 1 and autophagy by either beclin 1 overexpression, cystamine or antioxidants rescues the localization of the beclin 1 interactome to the endoplasmic reticulum and reverts the CF airway phenotype in vitro, in vivo in Scnn1b-transgenic and Cftr(F508del) homozygous mice, and in human CF nasal biopsies. Restoring beclin 1 or knocking down p62 rescued the trafficking of CFTR(F508del) to the cell surface. These data link the CFTR defect to autophagy deficiency, leading to the accumulation of protein aggregates and to lung inflammation.

  19. Ubiquitin Ligase TRIM62 Regulates CARD9-Mediated Anti-fungal Immunity and Intestinal Inflammation.

    Science.gov (United States)

    Cao, Zhifang; Conway, Kara L; Heath, Robert J; Rush, Jason S; Leshchiner, Elizaveta S; Ramirez-Ortiz, Zaida G; Nedelsky, Natalia B; Huang, Hailiang; Ng, Aylwin; Gardet, Agnès; Cheng, Shih-Chin; Shamji, Alykhan F; Rioux, John D; Wijmenga, Cisca; Netea, Mihai G; Means, Terry K; Daly, Mark J; Xavier, Ramnik J

    2015-10-20

    CARD9 is a central component of anti-fungal innate immune signaling via C-type lectin receptors, and several immune-related disorders are associated with CARD9 alterations. Here, we used a rare CARD9 variant that confers protection against inflammatory bowel disease as an entry point to investigating CARD9 regulation. We showed that the protective variant of CARD9, which is C-terminally truncated, acted in a dominant-negative manner for CARD9-mediated cytokine production, indicating an important role for the C terminus in CARD9 signaling. We identified TRIM62 as a CARD9 binding partner and showed that TRIM62 facilitated K27-linked poly-ubiquitination of CARD9. We identified K125 as the ubiquitinated residue on CARD9 and demonstrated that this ubiquitination was essential for CARD9 activity. Furthermore, we showed that similar to Card9-deficient mice, Trim62-deficient mice had increased susceptibility to fungal infection. In this study, we utilized a rare protective allele to uncover a TRIM62-mediated mechanism for regulation of CARD9 activation. PMID:26488816

  20. Coordination of autophagy with other cellular activities

    Institute of Scientific and Technical Information of China (English)

    Yan WANG; Zheng-hong QIN

    2013-01-01

    The cell biological phenomenon of autophagy has attracted increasing attention in recent years,partly as a consequence of the discovery of key components of its cellular machinery.Autophagy plays a crucial role in a myriad of cellular functions.Autophagy has its own regulatory mechanisms,but this process is not isolated.Autophagy is coordinated with other cellular activities to maintain cell homeostasis.Autophagy is critical for a range of human physiological processes.The multifunctional roles of autophagy are explained by its ability to interact with several key components of various cell pathways.In this review,we focus on the coordination between autophagy and other physiological processes,including the ubiquitin-proteasome system (UPS),energy homeostasis,aging,programmed cell death,the immune responses,microbial invasion and inflammation.The insights gained from investigating autophagic networks should increase our understanding of their roles in human diseases and their potential as targets for therapeutic intervention.

  1. Clearance of autophagy-associated dying retinal pigment epithelial cells - a possible source for inflammation in age-related macular degeneration.

    Science.gov (United States)

    Szatmári-Tóth, M; Kristóf, E; Veréb, Z; Akhtar, S; Facskó, A; Fésüs, L; Kauppinen, A; Kaarniranta, K; Petrovski, G

    2016-01-01

    Retinal pigment epithelial (RPE) cells can undergo different forms of cell death, including autophagy-associated cell death during age-related macular degeneration (AMD). Failure of macrophages or dendritic cells (DCs) to engulf the different dying cells in the retina may result in the accumulation of debris and progression of AMD. ARPE-19 and primary human RPE cells undergo autophagy-associated cell death upon serum depletion and oxidative stress induced by hydrogen peroxide (H2O2). Autophagy was revealed by elevated light-chain-3 II (LC3-II) expression and electron microscopy, while autophagic flux was confirmed by blocking the autophago-lysosomal fusion using chloroquine (CQ) in these cells. The autophagy-associated dying RPE cells were engulfed by human macrophages, DCs and living RPE cells in an increasing and time-dependent manner. Inhibition of autophagy by 3-methyladenine (3-MA) decreased the engulfment of the autophagy-associated dying cells by macrophages, whereas sorting out the GFP-LC3-positive/autophagic cell population or treatment by the glucocorticoid triamcinolone (TC) enhanced it. Increased amounts of IL-6 and IL-8 were released when autophagy-associated dying RPEs were engulfed by macrophages. Our data suggest that cells undergoing autophagy-associated cell death engage in clearance mechanisms guided by professional and non-professional phagocytes, which is accompanied by inflammation as part of an in vitro modeling of AMD pathogenesis. PMID:27607582

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

    Directory of Open Access Journals (Sweden)

    Won-Hee Song

    2014-01-01

    Full Text Available 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.

  3. Autophagy in Inflammatory Diseases

    Directory of Open Access Journals (Sweden)

    Alexander J. S. Choi

    2011-01-01

    Full Text Available Autophagy provides a mechanism for the turnover of cellular organelles and proteins through a lysosome-dependent degradation pathway. During starvation, autophagy exerts a homeostatic function that promotes cell survival by recycling metabolic precursors. Additionally, autophagy can interact with other vital processes such as programmed cell death, inflammation, and adaptive immune mechanisms, and thereby potentially influence disease pathogenesis. Macrophages deficient in autophagic proteins display enhanced caspase-1-dependent proinflammatory cytokine production and the activation of the inflammasome. Autophagy provides a functional role in infectious diseases and sepsis by promoting intracellular bacterial clearance. Mutations in autophagy-related genes, leading to loss of autophagic function, have been implicated in the pathogenesis of Crohn's disease. Furthermore, autophagy-dependent mechanisms have been proposed in the pathogenesis of several pulmonary diseases that involve inflammation, including cystic fibrosis and pulmonary hypertension. Strategies aimed at modulating autophagy may lead to therapeutic interventions for diseases associated with inflammation.

  4. Ubiquitin at the crossroad of cell death and survival

    Institute of Scientific and Technical Information of China (English)

    Yu-Shan Chen; Xiao-Bo Qiu

    2013-01-01

    Ubiquitination is crucial for cellular processes, such as protein degradation, apoptosis, autophagy, and cell cycle progression. Dysregulation of the ubiquitination network accounts for the development of numerous diseases, including cancer. Thus, targeting ubiquitination is a promising strategy in cancer therapy. Both apoptosis and autophagy are involved in tumorigenesis and response to cancer therapy. Although both are categorized as types of celldeath, autophagy is general y considered to have protective functions, including protecting cells from apoptosis under certain cellular stress conditions. This review highlights recent advances in understanding the regulation of apoptosis and autophagy by ubiquitination.

  5. Autophagy in Tuberculosis

    Science.gov (United States)

    Deretic, Vojo

    2014-01-01

    Autophagy as an immune mechanism controls inflammation and acts as a cell-autonomous defense against intracellular microbes including Mycobacterium tuberculosis. An equally significant role of autophagy is its anti-inflammatory and tissue-sparing function. This combination of antimicrobial and anti-inflammatory actions prevents active disease in animal models. In human populations, genetic links between autophagy, inflammatory bowel disease, and susceptibility to tuberculosis provide further support to these combined roles of autophagy. The autophagic control of M. tuberculosis and prevention of progressive disease provide novel insights into physiological and immune control of tuberculosis. It also offers host-based therapeutic opportunities because autophagy can be pharmacologically modulated. PMID:25167980

  6. Tart cherry supplementation improves working memory, hippocampal inflammation and autophagy in aged rats

    Science.gov (United States)

    High consumption of fruits and vegetables has been associated with reduced risk of debilitating diseases and improved cognition in aged populations. These beneficial effects have been attributed to the antioxidant/anti-inflammation properties of phytochemicals found in fruits and vegetables. Tart ch...

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

    Science.gov (United States)

    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...

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

    Directory of Open Access Journals (Sweden)

    Antje eVoigt

    2013-03-01

    Full Text Available 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.

  9. IL-1 receptor blockade restores autophagy and reduces inflammation in chronic granulomatous disease in mice and in humans

    NARCIS (Netherlands)

    Luca, A. De; Smeekens, S.P.; Casagrande, A.; Iannitti, R.; Conway, K.L.; Gresnigt, M.S.; Begun, J.; Plantinga, T.S.; Joosten, L.A.B.; Meer, J.W.M. van der; Chamilos, G.; Netea, M.G.; Xavier, R.J.; Dinarello, C.A.; Romani, L.; Veerdonk, F.L. van de

    2014-01-01

    Patients with chronic granulomatous disease (CGD) have a mutated NADPH complex resulting in defective production of reactive oxygen species; these patients can develop severe colitis and are highly susceptible to invasive fungal infection. In NADPH oxidase-deficient mice, autophagy is defective but

  10. Phosphorylation of the autophagy receptor optineurin restricts Salmonella growth

    DEFF Research Database (Denmark)

    Wild, Philipp; Farhan, Hesso; McEwan, David G;

    2011-01-01

    Selective autophagy can be mediated via receptor molecules that link specific cargoes to the autophagosomal membranes decorated by ubiquitin-like microtubule-associated protein light chain 3 (LC3) modifiers. Although several autophagy receptors have been identified, little is known about mechanisms...... controlling their functions in vivo. In this work, we found that phosphorylation of an autophagy receptor, optineurin, promoted selective autophagy of ubiquitin-coated cytosolic Salmonella enterica. The protein kinase TANK binding kinase 1 (TBK1) phosphorylated optineurin on serine-177, enhancing LC3 binding...... affinity and autophagic clearance of cytosolic Salmonella. Conversely, ubiquitin- or LC3-binding optineurin mutants and silencing of optineurin or TBK1 impaired Salmonella autophagy, resulting in increased intracellular bacterial proliferation. We propose that phosphorylation of autophagy receptors might...

  11. Involvement of Ubiquitin-Editing Protein A20 in Modulating Inflammation in Rat Cochlea Associated with Silver Nanoparticle-Induced CD68 Upregulation and TLR4 Activation

    Science.gov (United States)

    Feng, Hao; Pyykkö, Ilmari; Zou, Jing

    2016-05-01

    Silver nanoparticles (AgNPs) were shown to temporarily impair the biological barriers in the skin of the external ear canal, mucosa of the middle ear, and inner ear, causing partially reversible hearing loss after delivery into the middle ear. The current study aimed to elucidate the molecular mechanism, emphasizing the TLR signaling pathways in association with the potential recruitment of macrophages in the cochlea and the modulation of inflammation by ubiquitin-editing protein A20. Molecules potentially involved in these signaling pathways were thoroughly analysed using immunohistochemistry in the rat cochlea exposed to AgNPs at various concentrations through intratympanic injection. The results showed that 0.4 % AgNPs but not 0.02 % AgNPs upregulated the expressions of CD68, TLR4, MCP1, A20, and RNF11 in the strial basal cells, spiral ligament fibrocytes, and non-sensory supporting cells of Corti's organ. 0.4 % AgNPs had no effect on CD44, TLR2, MCP2, Rac1, myosin light chain, VCAM1, Erk1/2, JNK, p38, IL-1β, TNF-α, TNFR1, TNFR2, IL-10, or TGF-β. This study suggested that AgNPs might confer macrophage-like functions on the strial basal cells and spiral ligament fibrocytes and enhance the immune activities of non-sensory supporting cells of Corti's organ through the upregulation of CD68, which might be involved in TLR4 activation. A20 and RNF11 played roles in maintaining cochlear homeostasis via negative regulation of the expressions of inflammatory cytokines.

  12. Involvement of Ubiquitin-Editing Protein A20 in Modulating Inflammation in Rat Cochlea Associated with Silver Nanoparticle-Induced CD68 Upregulation and TLR4 Activation.

    Science.gov (United States)

    Feng, Hao; Pyykkö, Ilmari; Zou, Jing

    2016-12-01

    Silver nanoparticles (AgNPs) were shown to temporarily impair the biological barriers in the skin of the external ear canal, mucosa of the middle ear, and inner ear, causing partially reversible hearing loss after delivery into the middle ear. The current study aimed to elucidate the molecular mechanism, emphasizing the TLR signaling pathways in association with the potential recruitment of macrophages in the cochlea and the modulation of inflammation by ubiquitin-editing protein A20. Molecules potentially involved in these signaling pathways were thoroughly analysed using immunohistochemistry in the rat cochlea exposed to AgNPs at various concentrations through intratympanic injection. The results showed that 0.4 % AgNPs but not 0.02 % AgNPs upregulated the expressions of CD68, TLR4, MCP1, A20, and RNF11 in the strial basal cells, spiral ligament fibrocytes, and non-sensory supporting cells of Corti's organ. 0.4 % AgNPs had no effect on CD44, TLR2, MCP2, Rac1, myosin light chain, VCAM1, Erk1/2, JNK, p38, IL-1β, TNF-α, TNFR1, TNFR2, IL-10, or TGF-β. This study suggested that AgNPs might confer macrophage-like functions on the strial basal cells and spiral ligament fibrocytes and enhance the immune activities of non-sensory supporting cells of Corti's organ through the upregulation of CD68, which might be involved in TLR4 activation. A20 and RNF11 played roles in maintaining cochlear homeostasis via negative regulation of the expressions of inflammatory cytokines. PMID:27142878

  13. Autophagy and cytokines.

    Science.gov (United States)

    Harris, James

    2011-11-01

    Autophagy is a highly conserved homoeostatic mechanism for the lysosomal degradation of cytosolic constituents, including long-lived macromolecules, organelles and intracellular pathogens. Autophagosomes are formed in response to a number of environmental stimuli, including amino acid deprivation, but also by both host- and pathogen-derived molecules, including toll-like receptor ligands and cytokines. In particular, IFN-γ, TNF-α, IL-1, IL-2, IL-6 and TGF-β have been shown to induce autophagy, while IL-4, IL-10 and IL-13 are inhibitory. Moreover, autophagy can itself regulate the production and secretion of cytokines, including IL-1, IL-18, TNF-α, and Type I IFN. This review discusses the potentially pivotal roles of autophagy in the regulation of inflammation and the coordination of innate and adaptive immune responses.

  14. Autophagy in mammalian cells

    Institute of Scientific and Technical Information of China (English)

    Kadija; Abounit; Tiziano; M; Scarabelli; Roy; B; McCauley

    2012-01-01

    Autophagy is a regulated process for the degradation of cellular components that has been well conserved in eukaryotic cells. The discovery of autophagy-regulating proteins in yeast has been important in understanding this process. Although many parallels exist between fungi and mammals in the regulation and execution of autophagy, there are some important differences. The preautophagosomal structure found in yeast has not been identified in mammals, and it seems that there may be multiple origins for autophagosomes, including endoplasmic reticulum, plasma membrane and mitochondrial outer membrane. The maturation of the phagophore is largely dependent on 5’-AMP activated protein kinase and other factors that lead to the dephosphorylation of mammalian target of rapamycin. Once the process is initiated, the mammalian phagophore elongates and matures into an autophagosome by processes that are similar to those in yeast. Cargo selection is dependent on the ubiquitin conjugation of protein aggregates and organelles and recognition of these conjugates by autophagosomal receptors. Lysosomal degradation of cargo produces metabolites that can be recycled during stress. Autophagy is an impor-tant cellular safeguard during starvation in all eukaryotes; however, it may have more complicated, tissue specific roles in mammals. With certain exceptions, autophagy seems to be cytoprotective, and defects in the process have been associated with human disease.

  15. Nutritional Status and Cardiac Autophagy

    Directory of Open Access Journals (Sweden)

    Jihyun Ahn

    2013-02-01

    Full Text Available Autophagy is necessary for the degradation of long-lasting proteins and nonfunctional organelles, and is activated to promote cellular survival. However, overactivation of autophagy may deplete essential molecules and organelles responsible for cellular survival. Lifelong calorie restriction by 40% has been shown to increase the cardiac expression of autophagic markers, which suggests that it may have a cardioprotective effect by decreasing oxidative damage brought on by aging and cardiovascular diseases. Although cardiac autophagy is critical to regulating protein quality and maintaining cellular function and survival, increased or excessive autophagy may have deleterious effects on the heart under some circumstances, including pressure overload-induced heart failure. The importance of autophagy has been shown in nutrient supply and preservation of energy in times of limitation, such as ischemia. Some studies have suggested that a transition from obesity to metabolic syndrome may involve progressive changes in myocardial inflammation, mitochondrial dysfunction, fibrosis, apoptosis, and myocardial autophagy.

  16. Effects of the Total Saponins from Rosa laevigata Michx Fruit against Acetaminophen-Induced Liver Damage in Mice via Induction of Autophagy and Suppression of Inflammation and Apoptosis

    Directory of Open Access Journals (Sweden)

    Deshi Dong

    2014-05-01

    Full Text Available The effect of the total saponins from Rosa laevigata Michx fruit (RLTS against acetaminophen (APAP-induced liver damage in mice was evaluated in the present paper. The results showed that RLTS markedly improved the levels of liver SOD, CAT, GSH, GSH-Px, MDA, NO and iNOS, and the activities of serum ALT and AST caused by APAP. Further research confirmed that RLTS prevented fragmentation of DNA and mitochondrial ultrastructural alterations based on TdT-mediated dUTP nick end labeling (TUNEL and transmission electron microscopy (TEM assays. In addition, RLTS decreased the gene or protein expressions of cytochrome P450 (CYP2E1, pro-inflammatory mediators (IL-1β, IL-4, IL-6, TNF-α, iNOS, Bax, HMGB-1 and COX-2, pro-inflammatory transcription factors (NF-κB and AP-1, pro-apoptotic proteins (cytochrome C, p53, caspase-3, caspase-9, p-JNK, p-p38 and p-ERK, and increased the protein expressions of Bcl-2 and Bcl-xL. Moreover, the gene expression of IL-10, and the proteins including LC3, Beclin-1 and Atg5 induced by APAP were even more augmented by the extract. These results demonstrate that RLTS has hepatoprotective effects through antioxidative action, induction of autophagy, and suppression of inflammation and apoptosis, and could be developed as a potential candidate to treat APAP-induced liver damage in the future.

  17. Sch9 regulates intracellular protein ubiquitination by controlling stress responses

    Directory of Open Access Journals (Sweden)

    Beibei Qie

    2015-08-01

    Full Text Available Protein ubiquitination and the subsequent degradation are important means by which aberrant proteins are removed from cells, a key requirement for long-term survival. In this study, we found that the overall level of ubiquitinated proteins dramatically decreased as yeast cell grew from log to stationary phase. Deletion of SCH9, a gene encoding a key protein kinase for longevity control, decreased the level of ubiquitinated proteins in log phase and this effect could be reversed by restoring Sch9 function. We demonstrate here that the decrease of ubiquitinated proteins in sch9Δ cells in log phase is not caused by changes in ubiquitin expression, proteasome activity, or autophagy, but by enhanced expression of stress response factors and a decreased level of oxidative stress. Our results revealed for the first time how Sch9 regulates the level of ubiquitinated proteins and provides new insight into how Sch9 controls longevity.

  18. Autophagy in dementias.

    Science.gov (United States)

    Kragh, Christine Lund; Ubhi, Kiren; Wyss-Coray, Tony; Wyss-Corey, Tony; Masliah, Eliezer

    2012-01-01

    Dementias are a varied group of disorders typically associated with memory loss, impaired judgment and/or language and by symptoms affecting other cognitive and social abilities to a degree that interferes with daily functioning. Alzheimer's disease (AD) is the most common cause of a progressive dementia, followed by dementia with Lewy bodies (DLB), frontotemporal dementia (FTD), (VaD) and HIV-associated neurocognitive disorders (HAND). The pathogenesis of this group of disorders has been linked to the abnormal accumulation of proteins in the brains of affected individuals, which in turn has been related to deficits in protein clearance. Autophagy is a key cellular protein clearance pathway with proteolytic cleavage and degradation via the ubiquitin-proteasome pathway representing another important clearance mechanism. Alterations in the levels of autophagy and the proteins associated with the autophagocytic pathway have been reported in various types of dementias. This review will examine recent literature across these disorders and highlight a common theme of altered autophagy across the spectrum of the dementias. PMID:22150925

  19. The protein ATG16L1 suppresses inflammatory cytokines induced by the intracellular sensors Nod1 and Nod2 in an autophagy-independent manner.

    Science.gov (United States)

    Sorbara, Matthew T; Ellison, Lisa K; Ramjeet, Mahendrasingh; Travassos, Leonardo H; Jones, Nicola L; Girardin, Stephen E; Philpott, Dana J

    2013-11-14

    The peptidoglycan sensor Nod2 and the autophagy protein ATG16L1 have been linked to Crohn's disease (CD). Although Nod2 and the related sensor, Nod1, direct ATG16L1 to initiate anti-bacterial autophagy, whether ATG16L1 affects Nod-driven inflammation has not been examined. Here, we uncover an unanticipated autophagy-independent role for ATG16L1 in negatively regulating Nod-driven inflammatory responses. Knockdown of ATG16L1 expression, but not that of ATG5 or ATG9a, specifically enhanced Nod-driven cytokine production. In addition, autophagy-incompetent truncated forms of ATG16L1 regulated Nod-driven cytokine responses. Mechanistically, we demonstrated that ATG16L1 interfered with poly-ubiquitination of the Rip2 adaptor and recruitment of Rip2 into large signaling complexes. The CD-associated allele of ATG16L1 was impaired in its ability to regulate Nod-driven inflammatory responses. Overall, these results suggest that ATG16L1 is critical for Nod-dependent regulation of cytokine responses and that disruption of this Nod1- or Nod2-ATG16L1 signaling axis could contribute to the chronic inflammation associated with CD.

  20. Dysregulation of the Autophagy-Endolysosomal System in Amyotrophic Lateral Sclerosis and Related Motor Neuron Diseases

    Directory of Open Access Journals (Sweden)

    Asako Otomo

    2012-01-01

    Full Text Available Amyotrophic lateral sclerosis (ALS is a heterogeneous group of incurable motor neuron diseases (MNDs characterized by a selective loss of upper and lower motor neurons in the brain and spinal cord. Most cases of ALS are sporadic, while approximately 5–10% cases are familial. More than 16 causative genes for ALS/MNDs have been identified and their underlying pathogenesis, including oxidative stress, endoplasmic reticulum stress, excitotoxicity, mitochondrial dysfunction, neural inflammation, protein misfolding and accumulation, dysfunctional intracellular trafficking, abnormal RNA processing, and noncell-autonomous damage, has begun to emerge. It is currently believed that a complex interplay of multiple toxicity pathways is implicated in disease onset and progression. Among such mechanisms, ones that are associated with disturbances of protein homeostasis, the ubiquitin-proteasome system and autophagy, have recently been highlighted. Although it remains to be determined whether disease-associated protein aggregates have a toxic or protective role in the pathogenesis, the formation of them results from the imbalance between generation and degradation of misfolded proteins within neuronal cells. In this paper, we focus on the autophagy-lysosomal and endocytic degradation systems and implication of their dysfunction to the pathogenesis of ALS/MNDs. The autophagy-endolysosomal pathway could be a major target for the development of therapeutic agents for ALS/MNDs.

  1. Autophagy selectivity through receptor clustering

    Science.gov (United States)

    Rutenberg, Andrew; Brown, Aidan

    Substrate selectivity in autophagy requires an all-or-none cellular response. We focus on peroxisomes, for which autophagy receptor proteins NBR1 and p62 are well characterized. Using computational models, we explore the hypothesis that physical clustering of autophagy receptor proteins on the peroxisome surface provides an appropriate all-or-none response. We find that larger peroxisomes nucleate NBR1 clusters first, and lose them due to competitive coarsening last, resulting in significant size-selectivity. We then consider a secondary hypothesis that p62 inhibits NBR1 cluster formation. We find that p62 inhibition enhances size-selectivity enough that, even if there is no change of the pexophagy rate, the volume of remaining peroxisomes can significantly decrease. We find that enhanced ubiquitin levels suppress size-selectivity, and that this effect is more pronounced for individual peroxisomes. Sufficient ubiquitin allows receptor clusters to form on even the smallest peroxisomes. We conclude that NBR1 cluster formation provides a viable physical mechanism for all-or-none substrate selectivity in pexophagy. We predict that cluster formation is associated with significant size-selectivity. Now at Simon Fraser University.

  2. Roles of STAT3/SOCS3 Pathway in Regulating the Visual Function and Ubiquitin-Proteasome-dependent Degradation of Rhodopsin during Retinal Inflammation*

    OpenAIRE

    Ozawa, Yoko; Nakao, Keiko; Kurihara, Toshihide; Shimazaki, Takuya; Shimmura, Shigeto; Ishida, Susumu; Yoshimura, Akihiko; Tsubota, Kazuo; Okano, Hideyuki

    2008-01-01

    Inflammatory cytokines cause tissue dysfunction. We previously reported that retinal inflammation down-regulates rhodopsin expression and impairs visual function by an unknown mechanism. Here, we demonstrate that rhodopsin levels were preserved by suppressor of cytokine signaling 3 (SOCS3), a negative feedback regulator of STAT3 activation. SOCS3 was expressed mainly in photoreceptor cells in the retina. In the SOCS3-deficient retinas, rhodopsin protein levels dropped sooner, and the reductio...

  3. Advantages of Extracellular Ubiquitin in Modulation of Immune Responses.

    Science.gov (United States)

    Sujashvili, Rusudan

    2016-01-01

    T and B lymphocytes play a central role in protecting the human body from infectious pathogens but occasionally they can escape immune tolerance, become activated, and induce autoimmune diseases. All deregulated cellular processes are associated with improper functioning of the ubiquitin-proteasome system (UPS) in eukaryotic cells. The role of ubiquitin in regulation of immune responses and in autoimmune diseases is only beginning to emerge. Ubiquitin is found in intra- and extracellular fluids and is involved in regulation of numerous cellular processes. Extracellular ubiquitin ascribed a role in lymphocyte differentiation. It regulates differentiation and maturation of hematopoietic cell lines. Ubiquitination is involved in initiation, propagation, and termination of immune responses. Disrupted ubiquitination can lead to autoimmunity. Recent observations showed that it can suppress immune response and prevent inflammation. Exogenous ubiquitin may provide good potential as a new tool for targeted therapy for immune mediated disorders of various etiologies. PMID:27642236

  4. Inducing autophagy

    DEFF Research Database (Denmark)

    Harder, Lea M; Bunkenborg, Jakob; Andersen, Jens S.

    2014-01-01

    Autophagy is a lysosomal-mediated catabolic process, which through degradation of different cytoplasmic components aids in maintaining cellular homeostasis and survival during exposure to extra- or intracellular stresses. Ammonia is a potential toxic and stress-inducing byproduct of glutamine...... catabolism, which has recently been found to induce autophagy in an MTOR independent way and support cancer cell survival. In this study, quantitative phosphoproteomics was applied to investigate the initial signaling events linking ammonia to the induction of autophagy. The MTOR inhibitor rapamycin was used...... as a reference treatment to emphasize the differences between an MTOR-dependent and -independent autophagy-induction. By this means 5901 phosphosites were identified of which 626 were treatment-specific regulated and 175 were coregulated. Investigation of the ammonia-specific regulated sites supported that MTOR...

  5. [Progress in ubiquitin, ubiquitin chain and protein ubiquitination].

    Science.gov (United States)

    Lan, Qiuyan; Gao, Yuan; Li, Yanchang; Hong, Xuechuan; Xu, Ping

    2016-01-01

    Protein ubiquitination is one of the most important and widely exist protein post-translational modifications in eukaryotic cells, which takes the ubiquitin and ubiquitin chains as signal molecules to covalently modify other protein substrates. It plays an important roles in the control of almost all of the life processes, including gene transcription and translation, signal transduction and cell-cycle progression, besides classical 26S protesome degradation pathway. Varied modification sites in the same substrates as well as different types of ubiquitin linkages in the same modification sites contain different structural information, which conduct different signal or even determine the fate of the protein substrates in the cell. Any abnormalities in ubiquitin chain formation or its modification process may cause severe problem in maintaining the balance of intracellular environment and finally result in serious health problem of human being. In this review, we discussed the discovery, genetic characteristics and the crystal structure of the ubiquitin. We also emphasized the recent progresses of the assembly processes, structure and their biological function of ubiquitin chains. The relationship between the disregulation and related human diseases has also been discussed. These progress will shed light on the complexity of proteome, which may also provide tools in the new drug research and development processes. PMID:27363196

  6. Burkholderia cenocepacia J2315 escapes to the cytosol and actively subverts autophagy in human macrophages.

    Science.gov (United States)

    Al-Khodor, Souhaila; Marshall-Batty, Kimberly; Nair, Vinod; Ding, Li; Greenberg, David E; Fraser, Iain D C

    2014-03-01

    Selective autophagy functions to specifically degrade cellular cargo tagged by ubiquitination, including bacteria. Strains of the Burkholderia cepacia complex (Bcc) are opportunistic pathogens that cause life-threatening infections in patients with cystic fibrosis (CF) and chronic granulomatous disease (CGD). While there is evidence that defective macrophage autophagy in a mouse model of CF can influence B. cenocepacia susceptibility, there have been no comprehensive studies on how this bacterium is sensed and targeted by the host autophagy response in human macrophages. Here, we describe the intracellular life cycle of B. cenocepacia J2315 and its interaction with the autophagy pathway in human cells. Electron and confocal microscopy analyses demonstrate that the invading bacteria interact transiently with the endocytic pathway before escaping to the cytosol. This escape triggers theselective autophagy pathway, and the recruitment of ubiquitin, the ubiquitin-binding adaptors p62 and NDP52 and the autophagosome membrane-associated protein LC3B, to the bacterial vicinity. However, despite recruitment of these key autophagy pathway effectors, B. cenocepacia blocks autophagosome completion and replicates in the host cytosol. We find that a pre-infection increase in cellular autophagy flux can significantly inhibit B. cenocepacia replication and that lower autophagy flux in macrophages from immunocompromised CGD patients could contribute to increased B. cenocepacia susceptibility, identifying autophagy manipulation as a potential therapeutic approach to reduce bacterial burden in B. cenocepacia infections. PMID:24119232

  7. 细胞自噬和炎症反应的相互调控与牙周炎%Interaction of autophagy and inflammation in periodontitis

    Institute of Scientific and Technical Information of China (English)

    任静宜; 刘歆婵; 丁烨; 于洪强; 周延民; 于维先

    2016-01-01

    destroy periodontal tissues through the interaction of Toll-like receptor(TLR) with various components, such as lipopolysaccharide, peptidoglycan, and bacterial DNA. As a consequence, inflammatory cells are recruited and inflammatory cytokines are released. In local periodontal tissues, TLR or nucleotide-binding oligomerization domain-like receptor activates innate immune responses, induces autophagy-related pathways, and recognizes pathogen- and damage-associated molecular patterns. Autophagy can also influence inflammatory responses by negatively regulating TLR signals. This review focuses on recent progress in the mutual regulation of autophagy and inflammation. This review also describes the potential relations between autophagy and periodontitis to elucidate disease pathogenesis and to develop new therapies.

  8. Autophagy in lung disease pathogenesis and therapeutics

    OpenAIRE

    Ryter, Stefan W.; Augustine M K Choi

    2015-01-01

    Autophagy, a cellular pathway for the degradation of damaged organelles and proteins, has gained increasing importance in human pulmonary diseases, both as a modulator of pathogenesis and as a potential therapeutic target. In this pathway, cytosolic cargos are sequestered into autophagosomes, which are delivered to the lysosomes where they are enzymatically degraded and then recycled as metabolic precursors. Autophagy exerts an important effector function in the regulation of inflammation, an...

  9. Autophagy and its neuroprotection in neurodegenerative diseases

    Institute of Scientific and Technical Information of China (English)

    Ping Gu; Avaneesh Jakkoju; Mingwei Wang; Weidong Le

    2011-01-01

    It has been suggested that protein misfolding and aggregation contribute significantly to the development of neurodegenerative diseases. Misfolded and aggregated proteins are cleared by ubiquitin proteasomal system (UPS) and by both Micro and Macro autophagy lysosomal pathway (ALP). Autophagosomal dysfunction has been implicated in an increasing number of diseases including neurodegenerative diseases. Autophagy is a cellular self-eating process that plays an important role in neuroprotection as well as neuronal injury and death. While a decrease in autophagic activity interferes with protein degradation and possibly organelle turnover, increased autophagy has been shown to facilitate the clearance of aggregation-prone proteins and promote neuronal survival in a number of disease models. On the other hand, too much autophagic activity can be detrimental, suggesting the regulation of autophagy is critical in dictating cell fate. In this review paper, we will discuss various aspects of ALP biology and its dual functions in neuronal cell death and survival. We will also evaluate the role of autophagy in neurodegenerative diseases including Alzheimer's disease, Parkinson's disease, Huntington's disease, amyotrophic lateral sclerosis. Finally, we will explore the therapeutic potential of autophagy modifiers in several neurodegenerative diseases.

  10. Macrophage Autophagy in Atherosclerosis

    Directory of Open Access Journals (Sweden)

    Maria Chiara Maiuri

    2013-01-01

    Full Text Available Macrophages play crucial roles in atherosclerotic immune responses. Recent investigation into macrophage autophagy (AP in atherosclerosis has demonstrated a novel pathway through which these cells contribute to vascular inflammation. AP is a cellular catabolic process involving the delivery of cytoplasmic contents to the lysosomal machinery for ultimate degradation and recycling. Basal levels of macrophage AP play an essential role in atheroprotection during early atherosclerosis. However, AP becomes dysfunctional in the more advanced stages of the pathology and its deficiency promotes vascular inflammation, oxidative stress, and plaque necrosis. In this paper, we will discuss the role of macrophages and AP in atherosclerosis and the emerging evidence demonstrating the contribution of macrophage AP to vascular pathology. Finally, we will discuss how AP could be targeted for therapeutic utility.

  11. Sent to destroy: the ubiquitin proteasome system regulates cell signaling and protein quality control in cardiovascular development and disease.

    Science.gov (United States)

    Willis, Monte S; Townley-Tilson, W H Davin; Kang, Eunice Y; Homeister, Jonathon W; Patterson, Cam

    2010-02-19

    The ubiquitin proteasome system (UPS) plays a crucial role in biological processes integral to the development of the cardiovascular system and cardiovascular diseases. The UPS prototypically recognizes specific protein substrates and places polyubiquitin chains on them for subsequent destruction by the proteasome. This system is in place to degrade not only misfolded and damaged proteins, but is essential also in regulating a host of cell signaling pathways involved in proliferation, adaptation to stress, regulation of cell size, and cell death. During the development of the cardiovascular system, the UPS regulates cell signaling by modifying transcription factors, receptors, and structural proteins. Later, in the event of cardiovascular diseases as diverse as atherosclerosis, cardiac hypertrophy, and ischemia/reperfusion injury, ubiquitin ligases and the proteasome are implicated in protecting and exacerbating clinical outcomes. However, when misfolded and damaged proteins are ubiquitinated by the UPS, their destruction by the proteasome is not always possible because of their aggregated confirmations. Recent studies have discovered how these ubiquitinated misfolded proteins can be destroyed by alternative "specific" mechanisms. The cytosolic receptors p62, NBR, and histone deacetylase 6 recognize aggregated ubiquitinated proteins and target them for autophagy in the process of "selective autophagy." Even the ubiquitination of multiple proteins within whole organelles that drive the more general macro-autophagy may be due, in part, to similar ubiquitin-driven mechanisms. In summary, the crosstalk between the UPS and autophagy highlight the pivotal and diverse roles the UPS plays in maintaining protein quality control and regulating cardiovascular development and disease. PMID:20167943

  12. Monitoring Ubiquitin-Coated Bacteria via Confocal Microscopy.

    Science.gov (United States)

    Lork, Marie; Delvaeye, Mieke; Gonçalves, Amanda; Van Hamme, Evelien; Beyaert, Rudi

    2016-01-01

    Salmonella is a gram-negative facultative intracellular pathogen that is capable of infecting a variety of hosts. Inside host cells, most Salmonella bacteria reside and replicate within Salmonella-containing vacuoles. They use virulence proteins to manipulate the host cell machinery for their own benefit and hijack the host cytoskeleton to travel toward the perinuclear area. However, a fraction of bacteria escapes into the cytosol where they get decorated with a dense layer of polyubiquitin, which labels the bacteria for clearance by autophagy. More specifically, autophagy receptor proteins recognize the ubiquitinated bacteria and deliver them to autophagosomes, which subsequently fuse to lysosomes. Here, we describe methods used to infect HeLa cells with Salmonella bacteria and to detect their ubiquitination via immunofluorescence and laser scanning confocal microscopy. PMID:27613040

  13. Exposure to O-16 particle irradiation causes age-like decrements in rats through increased oxidative stress, inflammation and loss of autophagy

    Science.gov (United States)

    Exposing young rats to particles of high energy and charge (HZE particles) enhances indices of oxidative stress and inflammation, disrupts the functioning of neuronal communication, and alters cognitive behaviors. Even though exposure to these highly charged particles occurs at low fluence rates, p...

  14. Anti-inflammation activity of sinomenine-induced autophagy in EA.hy926 cells%盐酸青藤碱诱导EA.hy926细胞自噬及其在抗炎中的作用

    Institute of Scientific and Technical Information of China (English)

    杨亭; 倪振洪; 龚薇; 何凤田

    2013-01-01

    Objective To investigate the underlying mechanism of sinomenine,a traditional Chinese herb extract monomer,in inducing autophagy and its anti-inflammation activity in EA.hy926 cells.Methods Western blotting was taken to detect the expression of autophagic markers LC3 Ⅱ,ERK2,phosphorylation ERK2 and inflammatory cytokine HMGB1 in the EA.hy926 cells after the treatment of sinomenine at 50 or 100 μg/mL for 12 h.Acridine orange staining was used to observe the formation of acidic vesicular organelles in the cells by fluorescence microscopy.Results Western blotting revealed that the expression of LC3 Ⅱ(0.67 ± 0.05) and phosphorylation ERK2 (1.08 ± 0.05) were remarkably increased after the treatment of sinomenine at 100 μg/mL (P < 0.05).Autophagic vesicles were induced by sinomenine,while both LC3 Ⅱ and autophagic vesicles were partially decreased by ERK-specific inhibitor U0126 (P < 0.05).LPS-induced HMGB1 expression could be reduced by sinomenine,which was partially reversed by autophagy inhibitor CQ (P < 0.05).Conclusion It may be one of the mechanisms of anti-inflammation activity of sinomenine that can reduce HMGB1 by promoting autophagy through the ERK pathway in EA.hy926 cells.%目的 探讨中药单体提取物盐酸青藤碱诱导人内皮细胞EA.hy926自噬的机制及其在抗炎中发挥的作用.方法 Western blot检测分别以终浓度为50、100 μg/mL的盐酸青藤碱处理12 h后的EA.hy926细胞中自噬相关蛋白LC3Ⅱ、ERK2、磷酸化ERK2及炎症细胞因子HMGB1表达变化情况;荧光显微镜观察吖啶橙染色的经盐酸青藤碱诱导后EA.hy926细胞酸性小体变化情况.结果 经终浓度为50、100 μg/mL的盐酸青藤碱处理EA.hy926细胞后,与对照组比较,100 μg/mL的盐酸青藤碱可使自噬相关蛋白LC3Ⅱ表达(0.67±0.05)及ERK2的磷酸化水平(1.08 ±0.05)上调(P<0.05),ERK抑制剂U0126可使LC3Ⅱ表达下调(P<0.05)及EA.hy926细胞中酸性小体减少;

  15. In the family with ubiquitin

    OpenAIRE

    Alexandru, Gabriela; Pariente, Nonia; Xirodimas, Dimitris

    2011-01-01

    The Cold Spring Harbor meeting on ‘The Ubiquitin family', held in May 2011, brought together scientists from a wide range of fields, all under the common umbrella of ubiquitin and ubiquitin-like protein structure, function and regulation.

  16. Impaired macrophage autophagy increases the immune response in obese mice by promoting proinflammatory macrophage polarization.

    Science.gov (United States)

    Liu, Kun; Zhao, Enpeng; Ilyas, Ghulam; Lalazar, Gadi; Lin, Yu; Haseeb, Muhammad; Tanaka, Kathryn E; Czaja, Mark J

    2015-01-01

    Recent evidence that excessive lipid accumulation can decrease cellular levels of autophagy and that autophagy regulates immune responsiveness suggested that impaired macrophage autophagy may promote the increased innate immune activation that underlies obesity. Primary bone marrow-derived macrophages (BMDM) and peritoneal macrophages from high-fat diet (HFD)-fed mice had decreased levels of autophagic flux indicating a generalized impairment of macrophage autophagy in obese mice. To assess the effects of decreased macrophage autophagy on inflammation, mice with a Lyz2-Cre-mediated knockout of Atg5 in macrophages were fed a HFD and treated with low-dose lipopolysaccharide (LPS). Knockout mice developed systemic and hepatic inflammation with HFD feeding and LPS. This effect was liver specific as knockout mice did not have increased adipose tissue inflammation. The mechanism by which the loss of autophagy promoted inflammation was through the regulation of macrophage polarization. BMDM and Kupffer cells from knockout mice exhibited abnormalities in polarization with both increased proinflammatory M1 and decreased anti-inflammatory M2 polarization as determined by measures of genes and proteins. The heightened hepatic inflammatory response in HFD-fed, LPS-treated knockout mice led to liver injury without affecting steatosis. These findings demonstrate that autophagy has a critical regulatory function in macrophage polarization that downregulates inflammation. Defects in macrophage autophagy may underlie inflammatory disease states such as the decrease in macrophage autophagy with obesity that leads to hepatic inflammation and the progression to liver injury.

  17. The Impact of Autophagy on Cell Death Modalities

    Directory of Open Access Journals (Sweden)

    Stefan W. Ryter

    2014-01-01

    Full Text Available Autophagy represents a homeostatic cellular mechanism for the turnover of organelles and proteins, through a lysosome-dependent degradation pathway. During starvation, autophagy facilitates cell survival through the recycling of metabolic precursors. Additionally, autophagy can modulate other vital processes such as programmed cell death (e.g., apoptosis, inflammation, and adaptive immune mechanisms and thereby influence disease pathogenesis. Selective pathways can target distinct cargoes (e.g., mitochondria and proteins for autophagic degradation. At present, the causal relationship between autophagy and various forms of regulated or nonregulated cell death remains unclear. Autophagy can occur in association with necrosis-like cell death triggered by caspase inhibition. Autophagy and apoptosis have been shown to be coincident or antagonistic, depending on experimental context, and share cross-talk between signal transduction elements. Autophagy may modulate the outcome of other regulated forms of cell death such as necroptosis. Recent advances suggest that autophagy can dampen inflammatory responses, including inflammasome-dependent caspase-1 activation and maturation of proinflammatory cytokines. Autophagy may also act as regulator of caspase-1 dependent cell death (pyroptosis. Strategies aimed at modulating autophagy may lead to therapeutic interventions for diseases in which apoptosis or other forms of regulated cell death may play a cardinal role.

  18. Autophagy and intestinal homeostasis.

    Science.gov (United States)

    Patel, Khushbu K; Stappenbeck, Thaddeus S

    2013-01-01

    Nutrient absorption is the basic function that drives mammalian intestinal biology. To facilitate nutrient uptake, the host's epithelial barrier is composed of a single layer of cells. This constraint is problematic, as a design of this type can be easily disrupted. The solution during the course of evolution was to add numerous host defense mechanisms that can help prevent local and systemic infection. These mechanisms include specialized epithelial cells that produce a physiochemical barrier overlying the cellular barrier, robust and organized adaptive and innate immune cells, and the ability to mount an inflammatory response that is commensurate with a specific threat level. The autophagy pathway is a critical cellular process that strongly influences all these functions. Therefore, a fundamental understanding of the components of this pathway and their influence on inflammation, immunity, and barrier function will facilitate our understanding of homeostasis in the gastrointestinal tract. PMID:23216414

  19. Mechanisms of silver nanoparticle-induced toxicity and important role of autophagy.

    Science.gov (United States)

    Mao, Bin-Hsu; Tsai, Jui-Chen; Chen, Chun-Wan; Yan, Shian-Jang; Wang, Ying-Jan

    2016-10-01

    Safety concerns have been raised over the extensive applications of silver nanoparticles (AgNPs) because nano dimensions make them highly bioactive, being potentially harmful to the exposed humans. Surface physico-chemistry (shape, surface charge, chemical composition, etc.) that mainly dictates nano-bio interactions is relevant for influencing their biocompatibility and toxicity. Although the hazardousness of AgNPs has been demonstrated in vitro and in vivo, mechanistic understanding of the toxicity particularly at the molecular and organismal levels, in addition to oxidative stress and silver ion dissolution, has remained unclear. A growing body of research has elucidated that autophagy, being activated in response to exposure to various nanomaterials, may serve as a cellular defense mechanism against nanotoxicity. Recently, autophagy activation was shown to correlate with AgNPs exposure; however, the subsequent autophagosome-lysosome fusion was defective. As autophagy plays a crucial role in selective removal of stress-mediated protein aggregates and injured organelles, AgNPs-induced autophagic flux defect may consequently lead to aggravated cytotoxic responses. Furthermore, we suggest that p62 accumulation resulting from defective autophagy may also potentially account for AgNPs cytotoxicity. Intriguingly, AgNPs have been shown to interfere with ubiquitin modifications, either via upregulating levels of enzymes participating in ubiquitination, or through impairing the biological reactivity of ubiquitin (due to formation of AgNPs-ubiquitin corona). Ubiquitination both confers selectivity to autophagy as well as modulates stabilization, activation, and trafficking of proteins involved in autophagic clearance pathways. In this regard, we offer a new perspective that interference of AgNPs with ubiquitination may account for AgNPs-induced defective autophagy and cytotoxic effects. PMID:27240148

  20. It's all about talking: two-way communication between proteasomal and lysosomal degradation pathways via ubiquitin.

    Science.gov (United States)

    Liebl, Martina P; Hoppe, Thorsten

    2016-08-01

    Selective degradation of proteins requires a fine-tuned coordination of the two major proteolytic pathways, the ubiquitin-proteasome system (UPS) and autophagy. Substrate selection and proteolytic activity are defined by a plethora of regulatory cofactors influencing each other. Both proteolytic pathways are initiated by ubiquitylation to mark substrate proteins for degradation, although the size and/or topology of the modification are different. In this context E3 ubiquitin ligases, ensuring the covalent attachment of activated ubiquitin to the substrate, are of special importance. The regulation of E3 ligase activity, competition between different E3 ligases for binding E2 conjugation enzymes and substrates, as well as their interplay with deubiquitylating enzymes (DUBs) represent key events in the cross talk between the UPS and autophagy. The coordination between both degradation routes is further influenced by heat shock factors and ubiquitin-binding proteins (UBPs) such as p97, p62, or optineurin. Mutations in enzymes and ubiquitin-binding proteins or a general decline of both proteolytic systems during aging result in accumulation of damaged and aggregated proteins. Thus further mechanistic understanding of how UPS and autophagy communicate might allow therapeutic intervention especially against age-related diseases. PMID:27225656

  1. The Selective Autophagy Receptor p62 Forms a Flexible Filamentous Helical Scaffold

    Directory of Open Access Journals (Sweden)

    Rodolfo Ciuffa

    2015-05-01

    Full Text Available The scaffold protein p62/SQSTM1 is involved in protein turnover and signaling and is commonly found in dense protein bodies in eukaryotic cells. In autophagy, p62 acts as a selective autophagy receptor that recognizes and shuttles ubiquitinated proteins to the autophagosome for degradation. The structural organization of p62 in cellular bodies and the interplay of these assemblies with ubiquitin and the autophagic marker LC3 remain to be elucidated. Here, we present a cryo-EM structural analysis of p62. Together with structures of assemblies from the PB1 domain, we show that p62 is organized in flexible polymers with the PB1 domain constituting a helical scaffold. Filamentous p62 is capable of binding LC3 and addition of long ubiquitin chains induces disassembly and shortening of filaments. These studies explain how p62 assemblies provide a large molecular scaffold for the nascent autophagosome and reveal how they can bind ubiquitinated cargo.

  2. Autophagy is essential to support skeletal muscle plasticity in response to endurance exercise

    OpenAIRE

    Bernardi, Henri; Py, Guillaume; Candau, Robin B.

    2014-01-01

    Physical exercise is a stress that can substantially modulate cellular signaling mechanisms to promote morphological and metabolic adaptations. Skeletal muscle protein and organelle turnover is dependent on two major cellular pathways: Forkhead box class O proteins (FOXO) transcription factors that regulate two main proteolytic systems, the ubiquitin-proteasome, and the autophagy-lysosome systems, including mitochondrial autophagy, and the MTORC1 signaling associated with protein translation ...

  3. Autophagy, Metabolism, and Cancer.

    Science.gov (United States)

    White, Eileen; Mehnert, Janice M; Chan, Chang S

    2015-11-15

    Macroautophagy (autophagy hereafter) captures intracellular proteins and organelles and degrades them in lysosomes. The degradation breakdown products are released from lysosomes and recycled into metabolic and biosynthetic pathways. Basal autophagy provides protein and organelle quality control by eliminating damaged cellular components. Starvation-induced autophagy recycles intracellular components into metabolic pathways to sustain mitochondrial metabolic function and energy homeostasis. Recycling by autophagy is essential for yeast and mammals to survive starvation through intracellular nutrient scavenging. Autophagy suppresses degenerative diseases and has a context-dependent role in cancer. In some models, cancer initiation is suppressed by autophagy. By preventing the toxic accumulation of damaged protein and organelles, particularly mitochondria, autophagy limits oxidative stress, chronic tissue damage, and oncogenic signaling, which suppresses cancer initiation. This suggests a role for autophagy stimulation in cancer prevention, although the role of autophagy in the suppression of human cancer is unclear. In contrast, some cancers induce autophagy and are dependent on autophagy for survival. Much in the way that autophagy promotes survival in starvation, cancers can use autophagy-mediated recycling to maintain mitochondrial function and energy homeostasis to meet the elevated metabolic demand of growth and proliferation. Thus, autophagy inhibition may be beneficial for cancer therapy. Moreover, tumors are more autophagy-dependent than normal tissues, suggesting that there is a therapeutic window. Despite these insights, many important unanswered questions remain about the exact mechanisms of autophagy-mediated cancer suppression and promotion, how relevant these observations are to humans, and whether the autophagy pathway can be modulated therapeutically in cancer. See all articles in this CCR Focus section, "Cell Death and Cancer Therapy."

  4. Emerging role of selective autophagy in human diseases.

    Directory of Open Access Journals (Sweden)

    Kenji eMizumura

    2014-11-01

    Full Text Available AbstractAutophagy was originally described as a highly conserved system for the degradation of cytosol through a lysosome-dependent pathway. In response to starvation, autophagy degrades organelles and proteins to provide metabolites and energy for its pro-survival effects. Autophagy is recognized as playing a role in the pathogenesis of disease either directly or indirectly, through the regulation of vital processes such as programmed cell death, inflammation, and adaptive immune mechanisms. Recent studies have demonstrated that autophagy is not only a simple metabolite recycling system, but also has the ability to degrade specific cellular targets, such as mitochondria, cilia, and invading bacteria. In addition, selective autophagy has also been implicated in vesicle trafficking pathways, with potential roles in secretion and other intracellular transport processes. Selective autophagy has drawn the attention of researchers because of its potential importance in clinical diseases. Therapeutic strategies to target selective autophagy rather than general autophagy may maximize clinical benefit by enhancing selectivity. In this review, we outline the principle components of selective autophagy processes and their emerging importance in human disease, with an emphasis on pulmonary diseases.

  5. Autophagy is required for IL-2-mediated fibroblast growth

    Energy Technology Data Exchange (ETDEWEB)

    Kang, Rui [Department of Surgery, University of Pittsburgh, Pittsburgh, Pennsylvania 15219 (United States); Tang, Daolin, E-mail: tangd2@upmc.edu [Department of Surgery, University of Pittsburgh, Pittsburgh, Pennsylvania 15219 (United States); Lotze, Michael T., E-mail: lotzemt@upcm.edu [Department of Surgery, University of Pittsburgh, Pittsburgh, Pennsylvania 15219 (United States); Zeh III, Herbert J., E-mail: zehh@upmc.edu [Department of Surgery, University of Pittsburgh, Pittsburgh, Pennsylvania 15219 (United States)

    2013-02-15

    Autophagy is an evolutionarily conserved pathway responsible for delivery of cytoplasmic material into the lysosomal degradation pathway to enable vesicular exocytosis. Interleukin (IL)-2 is produced by T-cells and its activity is important for immunoregulation. Fibroblasts are an immune competent cell type, playing a critical role in wound healing, chronic inflammation, and tumor development. Although autophagy plays an important role in each of these processes, whether it regulates IL-2 activity in fibroblasts is unknown. Here, we show that autophagy is required for IL-2-induced cell growth in fibroblasts. IL-2 significantly induced autophagy in mouse embryonic fibroblasts (MEFs) and primary lung fibroblasts. Autophagy inhibitors (e.g., 3-methylamphetamine and bafilomycin A1) or knockdown of ATG5 and beclin 1 blocked clinical grade IL-2-induced autophagy. Moreover, IL-2 induced HMGB1 cytoplasmic translocation in MEFs and promoted interaction between HMGB1 and beclin1, which is required for autophagy induction. Pharmacological and genetic inhibition of autophagy inhibited IL-2-induced cell proliferation and enhanced IL-2-induced apoptosis. These findings suggest that autophagy is an important pro-survival regulator for IL-2-induced cell growth in fibroblasts.

  6. Hypoxia, MTOR and autophagy

    OpenAIRE

    Blagosklonny, Mikhail V.

    2013-01-01

    Although hypoxia can cause cell cycle arrest, it may simultaneously suppress a conversion from this arrest to senescence. Furthermore, hypoxia can suppress senescence caused by diverse stimuli, maintaining reversible quiescence instead. Hypoxia activates autophagy and inhibits MTOR, thus also activating autophagy. What is the relationship between autophagy and cellular senescence? Also, can inhibition of MTOR and stimulation of autophagy explain the gerosuppressive effects of hypoxia?

  7. Oxidative stress, redox signaling pathways, and autophagy in cachectic muscles of male patients with advanced COPD and lung cancer.

    Science.gov (United States)

    Puig-Vilanova, Ester; Rodriguez, Diego A; Lloreta, Josep; Ausin, Pilar; Pascual-Guardia, Sergio; Broquetas, Joan; Roca, Josep; Gea, Joaquim; Barreiro, Esther

    2015-02-01

    Muscle dysfunction and wasting are predictors of mortality in advanced COPD and malignancies. Redox imbalance and enhanced protein catabolism are underlying mechanisms in COPD. We hypothesized that the expression profile of several biological markers share similarities in patients with cachexia associated with either COPD or lung cancer (LC). In vastus lateralis of cachectic patients with either LC (n=10) or advanced COPD (n=16) and healthy controls (n=10), markers of redox balance, inflammation, proteolysis, autophagy, signaling pathways, mitochondrial function, muscle structure, and sarcomere damage were measured using laboratory and light and electron microscopy techniques. Systemic redox balance and inflammation were also determined. All subjects were clinically evaluated. Compared to controls, in both cachectic groups of patients, a similar expression profile of different biological markers was observed in their muscles: increased levels of muscle protein oxidation and ubiquitination (p<0.05, both), which positively correlated (r=0.888), redox-sensitive signaling pathways (NF-κB and FoxO) were activated (p<0.05, all), fast-twitch fiber sizes were atrophied, muscle structural abnormalities and sarcomere disruptions were significantly greater (p<0.05, both). Structural and functional protein levels were lower in muscles of both cachectic patient groups than in controls (p<0.05, all). However, levels of autophagy markers including ultrastructural autophagosome counts were increased only in muscles of cachectic COPD patients (p<0.05). Systemic oxidative stress and inflammation levels were also increased in both patient groups compared to controls (p<0.005, both). Oxidative stress and redox-sensitive signaling pathways are likely to contribute to the etiology of muscle wasting and sarcomere disruption in patients with respiratory cachexia: LC and COPD. PMID:25464271

  8. Autophagy deficiency in macrophages enhances NLRP3 inflammasome activity and chronic lung disease following silica exposure.

    Science.gov (United States)

    Jessop, Forrest; Hamilton, Raymond F; Rhoderick, Joseph F; Shaw, Pamela K; Holian, Andrij

    2016-10-15

    Autophagy is an important metabolic mechanism that can promote cellular survival following injury. The specific contribution of autophagy to silica-induced inflammation and disease is not known. The objective of these studies was to determine the effects of silica exposure on the autophagic pathway in macrophages, as well as the general contribution of autophagy in macrophages to inflammation and disease. Silica exposure enhanced autophagic activity in vitro in Bone Marrow derived Macrophages and in vivo in Alveolar Macrophages isolated from silica-exposed mice. Impairment of autophagy in myeloid cells in vivo using Atg5(fl/fl)LysM-Cre(+) mice resulted in enhanced cytotoxicity and inflammation after silica exposure compared to littermate controls, including elevated IL-18 and the alarmin HMGB1 in the whole lavage fluid. Autophagy deficiency caused some spontaneous inflammation and disease. Greater silica-induced acute inflammation in Atg5(fl/fl)LysM-Cre(+) mice correlated with increased fibrosis and chronic lung disease. These studies demonstrate a critical role for autophagy in suppressing silica-induced cytotoxicity and inflammation in disease development. Furthermore, this data highlights the importance of basal autophagy in macrophages and other myeloid cells in maintaining lung homeostasis.

  9. Reciprocal regulation of cilia and autophagy via the MTOR and proteasome pathways.

    Science.gov (United States)

    Wang, Shixuan; Livingston, Man J; Su, Yunchao; Dong, Zheng

    2015-04-01

    Primary cilium is an organelle that plays significant roles in a number of cellular functions ranging from cell mechanosensation, proliferation, and differentiation to apoptosis. Autophagy is an evolutionarily conserved cellular function in biology and indispensable for cellular homeostasis. Both cilia and autophagy have been linked to different types of genetic and acquired human diseases. Their interaction has been suggested very recently, but the underlying mechanisms are still not fully understood. We examined autophagy in cells with suppressed cilia and measured cilium length in autophagy-activated or -suppressed cells. It was found that autophagy was repressed in cells with short cilia. Further investigation showed that MTOR activation was enhanced in cilia-suppressed cells and the MTOR inhibitor rapamycin could largely reverse autophagy suppression. In human kidney proximal tubular cells (HK2), autophagy induction was associated with cilium elongation. Conversely, autophagy inhibition by 3-methyladenine (3-MA) and chloroquine (CQ) as well as bafilomycin A1 (Baf) led to short cilia. Cilia were also shorter in cultured atg5-knockout (KO) cells and in atg7-KO kidney proximal tubular cells in mice. MG132, an inhibitor of the proteasome, could significantly restore cilium length in atg5-KO cells, being concomitant with the proteasome activity. Together, the results suggest that cilia and autophagy regulate reciprocally through the MTOR signaling pathway and ubiquitin-proteasome system.

  10. Ubiquitination of plant immune receptors.

    Science.gov (United States)

    Zhou, Jinggeng; He, Ping; Shan, Libo

    2014-01-01

    Ubiquitin is a highly conserved regulatory protein consisting of 76 amino acids and ubiquitously expressed in all eukaryotic cells. The reversible ubiquitin conjugation to a wide variety of target proteins, a process known as ubiquitination or ubiquitylation, serves as one of the most important and prevalent posttranslational modifications to regulate the myriad actions of protein cellular functions, including protein degradation, vesicle trafficking, and subcellular localization. Protein ubiquitination is an ATP-dependent stepwise covalent attachment of one or more ubiquitin molecules to target proteins mediated by a hierarchical enzymatic cascade consisting of an E1 ubiquitin-activating enzyme, E2 ubiquitin-conjugating enzyme, and E3 ubiquitin ligase. The plant plasma membrane resident receptor-like kinase Flagellin Sensing 2 (FLS2) recognizes bacterial flagellin and initiates innate immune signaling to defend against pathogen attacks. We have recently shown that two plant U-box E3 ubiquitin ligases PUB12 and PUB13 directly ubiquitinate FLS2 and promote flagellin-induced FLS2 degradation, which in turn attenuates FLS2 signaling to prevent excessive or prolonged activation of immune responses. Here, we use FLS2 as an example to describe a protocol for detection of protein ubiquitination in plant cells in vivo and in test tubes in vitro. In addition, we elaborate the approach to identify different types of ubiquitin linkages by using various lysine mutants of ubiquitin. The various in vivo and in vitro ubiquitination assays will provide researchers with the tools to address how ubiquitination regulates diverse cellular functions of target proteins. PMID:25117287

  11. Interplay between Leucine-Rich Repeat Kinase 2 (LRRK2) and p62/SQSTM-1 in Selective Autophagy

    Science.gov (United States)

    Park, Sangwook; Han, Seulki; Choi, Insup; Kim, Beomsue; Park, Seung Pyo; Joe, Eun-Hye; Suh, Young Ho

    2016-01-01

    The deposit of polyubiquitinated aggregates has been implicated in the pathophysiology of Parkinson’s disease (PD), and growing evidence indicates that selective autophagy plays a critical role in the clearance of ubiquitin-positive protein aggregates by autophagosomes. The selective autophagic receptor p62/SQSTM-1, which associates directly with both ubiquitin and LC3, transports ubiquitin conjugates to autophagosomes for degradation. Leucine-rich repeat kinase 2 (LRRK2), a PD-associated protein kinase, is tightly controlled by autophagy-lysosome degradation as well as by the ubiquitin-proteasome pathway. However, little is known about the degradation of ubiquitinated LRRK2 via selective autophagy. In the present study, we found that p62/SQSTM-1 physically interacts with LRRK2 as a selective autophagic receptor. The overexpression of p62 leads to the robust degradation of LRRK2 through the autophagy-lysosome pathway. In addition, LRRK2 indirectly regulates Ser351 and Ser403 phosphorylation of p62. Of particular interest, the interaction between phosphorylated p62 and Keap1 is reduced by LRRK2 overexpression. Therefore, we propose that the interplay between LRRK2 and p62 may contribute to the pathophysiological function and homeostasis of LRRK2 protein. PMID:27631370

  12. Selective Autophagy in Drosophila

    Directory of Open Access Journals (Sweden)

    Ioannis P. Nezis

    2012-01-01

    Full Text Available Autophagy is an evolutionarily conserved process of cellular self-eating and is a major pathway for degradation of cytoplasmic material by the lysosomal machinery. Autophagy functions as a cellular response in nutrient starvation, but it is also associated with the removal of protein aggregates and damaged organelles and therefore plays an important role in the quality control of proteins and organelles. Although it was initially believed that autophagy occurs randomly in the cell, during the last years, there is growing evidence that sequestration and degradation of cytoplasmic material by autophagy can be selective. Given the important role of autophagy and selective autophagy in several disease-related processes such as neurodegeneration, infections, and tumorigenesis, it is important to understand the molecular mechanisms of selective autophagy, especially at the organismal level. Drosophila is an excellent genetically modifiable model organism exhibiting high conservation in the autophagic machinery. However, the regulation and mechanisms of selective autophagy in Drosophila have been largely unexplored. In this paper, I will present an overview of the current knowledge about selective autophagy in Drosophila.

  13. Blocking autophagy prevents bortezomib-induced NF-κB activation by reducing I-κBα degradation in lymphoma cells.

    Directory of Open Access Journals (Sweden)

    Li Jia

    Full Text Available 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-κBα, to the autophagosome. Degradation of I-κBα results in increased NF-κB nuclear translocation and transcription activity. Since bortezomib treatment promoted I-κBα phosphorylation, ubiquitination and degradation, this suggests that the route of I-κBα degradation was not via the ubiquitin-proteasome degradation system. The autophagy inhibitor chloroquine (CQ significantly inhibited bortezomib-induced I-κBα degradation, increased complex formation with NF-κB and reduced NF-κB 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-κBα. Inhibition of both autophagy and the proteasome has great potential to kill apoptosis-resistant lymphoma cells.

  14. Regulation of autophagy in oxygen-dependent cellular stress.

    Science.gov (United States)

    Ryter, Stefan W; Choi, Augustine M K

    2013-01-01

    Oxidative stress caused by supraphysiological production of reactive oxygen species (ROS), can cause cellular injury associated with protein and lipid oxidation, DNA damage, and mitochondrial dysfunction. The cellular responses triggered by oxidative stress include the altered regulation of signaling pathways that culminate in the regulation of cell survival or cell death pathways. Recent studies suggest that autophagy, a cellular homeostatic process that governs the turnover of damaged organelles and proteins, may represent a general cellular and tissue response to oxidative stress. The autophagic pathway involves the encapsulation of substrates in double-membraned vesicles, which are subsequently delivered to the lysosome for enzymatic degradation and recycling of metabolic precursors. Autophagy may play multifunctional roles in cellular adaptation to stress, by maintaining mitochondrial integrity, and removing damaged proteins. Additionally, autophagy may play important roles in the regulation of inflammation and immune function. Modulation of the autophagic pathway has been reported in cell culture models of oxidative stress, including altered states of oxygen tension (i.e., hypoxia, hyperoxia), and exposure to oxidants. Furthermore, proteins that regulate autophagy may be subject to redox regulation. The heme oxygenase- 1 (HO)-1 enzyme system may have a role in the regulation of autophagy. Recent studies suggest that carbon monoxide (CO), a reaction product of HO activity which can alter mitochondrial function, may induce autophagy in cultured epithelial cells. In conclusion, current research suggests a central role for autophagy as a mammalian oxidative stress response and its interrelationship to other stress defense systems. PMID:23092322

  15. Enhanced autophagy as a potential mechanism for the improved physiological function by simvastatin in muscular dystrophy.

    Science.gov (United States)

    Whitehead, Nicholas P

    2016-04-01

    Autophagy has recently emerged as an important cellular process for the maintenance of skeletal muscle health and function. Excessive autophagy can trigger muscle catabolism, leading to atrophy. In contrast, reduced autophagic flux is a characteristic of several muscle diseases, including Duchenne muscular dystrophy, the most common and severe inherited muscle disorder. Recent evidence demonstrates that enhanced reactive oxygen species (ROS) production by CYBB/NOX2 impairs autophagy in muscles from the dmd/mdx mouse, a genetic model of Duchenne muscular dystrophy. Statins decrease CYBB/NOX2 expression and activity and stimulate autophagy in skeletal muscle. Therefore, we treated dmd/mdx mice with simvastatin and showed decreased CYBB/NOX2-mediated oxidative stress and enhanced autophagy induction. This was accompanied by reduced muscle damage, inflammation and fibrosis, and increased muscle force production. Our data suggest that increased autophagy may be a potential mechanism by which simvastatin improves skeletal muscle health and function in muscular dystrophy. PMID:26890413

  16. Autophagy and cancer

    Institute of Scientific and Technical Information of China (English)

    Si-Zhao; Lu; Duygu; Dee; Harrison-Findik

    2013-01-01

    Autophagy is a homeostatic and evolutionarily conserved mechanism of self-digestion by which the cells degrade and recycle long-lived proteins and excess or damaged organelles.Autophagy is activated in response to both physiological and pathological stimuli including growth factor depletion,energy deficiency or the upregulation of Bcl-2 protein expression.A novel role of autophagy in various cancers has been proposed.Interestingly,evidence that supports both a positive and negative role of autophagy in the pathogenesis of cancer has been reported.As a tumor suppression mechanism,autophagy maintains genome stability,induces senescence and possibly autophagic cell death.On the other hand,autophagy participates in tumor growth and maintenance by supplying metabolic substrate,limiting oxidative stress,and maintaining cancer stem cell population.It has been proposed that the differential roles of autophagy in cancer are disease type and stage specific.In addition,substrate selectivity might be involved in carrying out the specific effect of autophagy in cancer,and represents one of the potential directions for future studies.

  17. Drosophila Parkin requires PINK1 for mitochondrial translocation and ubiquitinates Mitofusin

    OpenAIRE

    Ziviani, E.; Tao, R.N.; Whitworth, A. J.

    2010-01-01

    Loss of the E3 ubiquitin ligase Parkin causes early onset Parkinson's disease, a neurodegenerative disorder of unknown etiology. Parkin has been linked to multiple cellular processes including protein degradation, mitochondrial homeostasis, and autophagy; however, its precise role in pathogenesis is unclear. Recent evidence suggests that Parkin is recruited to damaged mitochondria, possibly affecting mitochondrial fission and/or fusion, to mediate their autophagic turnover. The precise mechan...

  18. Chemical Inhibition of Autophagy

    DEFF Research Database (Denmark)

    Baek, Eric; Lin Kim, Che; Gyeom Kim, Mi;

    2016-01-01

    Chinese hamster ovary (CHO) cells activate and undergo apoptosis and autophagy for various environmental stresses. Unlike apoptosis, studies on increasing the production of therapeutic proteins in CHO cells by targeting the autophagy pathway are limited. In order to identify the effects of chemical...... autophagy inhibitors on the specific productivity (qp), nine chemical inhibitors that had been reported to target three different phases of autophagy (metformin, dorsomorphin, resveratrol, and SP600125 against initiation and nucleation; 3-MA, wortmannin, and LY294002 against elongation, and chloroquine...... significantly increased the qp of DG44-Fc and DUKX-Fc. In contrast, for DG44-Ab, only 3-MA significantly increased the qp. The autophagy-inhibiting activity of the nine chemical inhibitors on the rCHO cell lines was evaluated through Western blot analysis and flow cytometry. Unexpectedly, some chemical...

  19. Autophagy in Hepatic Fibrosis

    Directory of Open Access Journals (Sweden)

    Yang Song

    2014-01-01

    Full Text Available Hepatic fibrosis is a leading cause of morbidity and mortality worldwide. Hepatic fibrosis is usually associated with chronic liver diseases caused by infection, drugs, metabolic disorders, or autoimmune imbalances. Effective clinical therapies are still lacking. Autophagy is a cellular process that degrades damaged organelles or protein aggregation, which participates in many pathological processes including liver diseases. Autophagy participates in hepatic fibrosis by activating hepatic stellate cells and may participate as well through influencing other fibrogenic cells. Besides that, autophagy can induce some liver diseases to develop while it may play a protective role in hepatocellular abnormal aggregates related liver diseases and reduces fibrosis. With a better understanding of the potential effects of autophagy on hepatic fibrosis, targeting autophagy might be a novel therapeutic strategy for hepatic fibrosis in the near future.

  20. SLC9A3R1 stimulates autophagy via BECN1 stabilization in breast cancer cells.

    Science.gov (United States)

    Liu, Hong; Ma, Yan; He, Hong-Wei; Wang, Jia-Ping; Jiang, Jian-Dong; Shao, Rong-Guang

    2015-01-01

    Autophagy, a self-catabolic process, has been found to be involved in abrogating the proliferation and metastasis of breast cancer. SLC9A3R1 (solute carrier family 9, subfamily A [NHE3, cation proton antiporter 3], member 3 regulator 1), a multifunctional scaffold protein, is involved in suppressing breast cancer cells proliferation and the SLC9A3R1-related signaling pathway regulates the activation of autophagy processes. However, the precise regulatory mechanism and signaling pathway of SLC9A3R1 in the regulation of autophagy processes in breast cancer cells remains unknown. Here, we report that the stability of BECN1, the major component of the autophagic core lipid kinase complex, is augmented in SLC9A3R1-overexpressing breast cancer MDA-MB-231 cells, subsequently stimulating autophagy by attenuating the interaction between BECN1 and BCL2. Initially, we found that SLC9A3R1 partially stimulated autophagy through the PTEN-PI3K-AKT1 signaling cascade in MDA-MB-231 cells. SLC9A3R1 then attenuated the interaction between BECN1 and BCL2 to stimulate the autophagic core lipid kinase complex. Further findings revealed that SLC9A3R1 bound to BECN1 and subsequently blocked ubiquitin-dependent BECN1 degradation. And the deletion of the C-terminal domain of SLC9A3R1 resulted in significantly reduced binding to BECN1. Moreover, the lack of C-terminal of SLC9A3R1 neither reduced the ubiquitination of BECN1 nor induced autophagy in breast cancer cells. The decrease in BECN1 degradation induced by SLC9A3R1 resulted in the activity of autophagy stimulation in breast cancer cells. These findings indicate that the SLC9A3R1-BECN1 signaling pathway participates in the activation of autophagy processes in breast cancer cells.

  1. TNFAIP3 promotes survival of CD4 T cells by restricting MTOR and promoting autophagy.

    Science.gov (United States)

    Matsuzawa, Yu; Oshima, Shigeru; Takahara, Masahiro; Maeyashiki, Chiaki; Nemoto, Yasuhiro; Kobayashi, Masanori; Nibe, Yoichi; Nozaki, Kengo; Nagaishi, Takashi; Okamoto, Ryuichi; Tsuchiya, Kiichiro; Nakamura, Tetsuya; Ma, Averil; Watanabe, Mamoru

    2015-01-01

    Autophagy plays important roles in metabolism, differentiation, and survival in T cells. TNFAIP3/A20 is a ubiquitin-editing enzyme that is thought to be a negative regulator of autophagy in cell lines. However, the role of TNFAIP3 in autophagy remains unclear. To determine whether TNFAIP3 regulates autophagy in CD4 T cells, we first analyzed Tnfaip3-deficient naïve CD4 T cells in vitro. We demonstrated that Tnfaip3-deficient CD4 T cells exhibited reduced MAP1LC3/LC3 (microtubule-associated protein 1 light chain 3) puncta formation, increased mitochondrial content, and exaggerated reactive oxygen species (ROS) production. These results indicate that TNFAIP3 promotes autophagy after T cell receptor (TCR) stimulation in CD4 T cells. We then investigated the mechanism by which TNFAIP3 promotes autophagy signaling. We found that TNFAIP3 bound to the MTOR (mechanistic target of rapamycin) complex and that Tnfaip3-deficient cells displayed enhanced ubiquitination of the MTOR complex and MTOR activity. To confirm the effects of enhanced MTOR activity in Tnfaip3-deficient cells, we analyzed cell survival following treatment with Torin1, an MTOR inhibitor. Tnfaip3-deficient CD4 T cells exhibited fewer cell numbers than the control cells in vitro and in vivo. In addition, the impaired survival of Tnfaip3-deficient cells was ameliorated with Torin1 treatment in vitro and in vivo. The effect of Torin1 was abolished by Atg5 deficiency. Thus, enhanced MTOR activity regulates the survival of Tnfaip3-deficient CD4 T cells. Taken together, our findings illustrate that TNFAIP3 restricts MTOR signaling and promotes autophagy, providing new insight into the manner in which MTOR and autophagy regulate survival in CD4 T cells.

  2. Ubiquitin domain proteins in disease

    DEFF Research Database (Denmark)

    Klausen, Louise Kjær; Schulze, Andrea; Seeger, Michael;

    2007-01-01

    The human genome encodes several ubiquitin-like (UBL) domain proteins (UDPs). Members of this protein family are involved in a variety of cellular functions and many are connected to the ubiquitin proteasome system, an essential pathway for protein degradation in eukaryotic cells. Despite...... and cancer. Publication history: Republished from Current BioData's Targeted Proteins database (TPdb; http://www.targetedproteinsdb.com)....

  3. Inhibition of autophagy induced by proteasome inhibition increases cell death in human SHG-44 glioma cells

    Institute of Scientific and Technical Information of China (English)

    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

    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 investigate whether inhibition of the proteasome can induce autophagy and the effects of autophagy on the fate of human SHG-44 glioma cells.Methods:The proteasome inhibitor MG-132 was used to induce autophagy in SHG-44 glioma cells,and the effect of autophagy on the survival of SHG-44 glioma cells was investigated using an autophagy inhibitor 3-MA.Cell viability was measured by MTT assay.Apoptosis and cell cycle were detected by flow cytometry.The expression of autophagy related proteins was determined by Western blot.Results:MG-132 inhibited cell proliferation,induced cell death and cell cycle arrest at G~JM phase,and activated autophagy in SHG-44 glioma cells.The expression of autophagy-related Beclin-1 and LC3-1 was significantly up-regulated and part of LC3-1 was converted into LC3-11.However,when SHG-44 glioma cells were co-treated with MG-132 and 3-MA,the cells became less viable,but cell death and cell numbers at G2/M phase increased.Moreover,the accumulation of acidic vesicular organelles was decreased,the expression of Beclin-1 and LC3 was significantly down-regulated and the conversion of LC3-11 from LC3-1 was also inhibited.Conclusion:Inhibition of the proteasome can induce autophagy in human SHG-44 glioma cells,and inhibition of autophagy increases cell death.This discovery may shed new light on the effect of autophagy on modulating the fate of SHG-44 glioma cells.

  4. Macrophage migration inhibitory factor induces autophagy via reactive oxygen species generation.

    Directory of Open Access Journals (Sweden)

    Yung-Chun Chuang

    Full Text Available Autophagy is an evolutionarily conserved catabolic process that maintains cellular homeostasis under stress conditions such as starvation and pathogen infection. Macrophage migration inhibitory factor (MIF is a multifunctional cytokine that plays important roles in inflammation and tumorigenesis. Cytokines such as IL-1β and TNF-α that are induced by MIF have been shown to be involved in the induction of autophagy. However, the actual role of MIF in autophagy remains unclear. Here, we have demonstrated that incubation of human hepatoma cell line HuH-7 cells with recombinant MIF (rMIF induced reactive oxygen species (ROS production and autophagy formation, including LC3-II expression, LC3 punctae formation, autophagic flux, and mitochondria membrane potential loss. The autophagy induced by rMIF was inhibited in the presence of MIF inhibitor, ISO-1 as well as ROS scavenger N-acetyl-L-cysteine (NAC. In addition, serum starvation-induced MIF release and autophagy of HuH-7 cells were partly blocked in the presence of NAC. Moreover, diminished MIF expression by shRNA transfection or inhibition of MIF by ISO-1 decreased serum starvation-induced autophagy of HuH-7 cells. Taken together, these data suggest that cell autophagy was induced by MIF under stress conditions such as inflammation and starvation through ROS generation.

  5. Phospho-ubiquitin: upending the PINK-Parkin-ubiquitin cascade.

    Science.gov (United States)

    Matsuda, Noriyuki

    2016-04-01

    Mitochondria with decreased membrane potential are characterized by defects in protein import into the matrix and impairments in high-efficiency synthesis of ATP. These low-quality mitochondria are marked with ubiquitin for selective degradation. Key factors in this mechanism are PTEN-induced putative kinase 1 (PINK1, a mitochondrial kinase) and Parkin (a ubiquitin ligase), disruption of which has been implicated in predisposition to Parkinson's disease. Previously, the clearance of damaged mitochondria had been thought to be the end result of a simple cascading reaction of PINK1-Parkin-ubiquitin. However, in the past year, several research groups including ours unexpectedly revealed that Parkin regulation is mediated by PINK1-dependent phosphorylation of ubiquitin. These results overturned the simple hierarchy that posited PINK1 and ubiquitin as the upstream and downstream factors of Parkin, respectively. Although ubiquitylation is well-known as a post-translational modification, it has recently become clear that ubiquitin itself can be modified, and that this modification unexpectedly converts ubiquitin to a factor that functions in retrograde signalling.

  6. Suppression of autophagy dysregulates the antioxidant response and causes premature senescence of melanocytes.

    Science.gov (United States)

    Zhang, Cheng-Feng; Gruber, Florian; Ni, Chunya; Mildner, Michael; Koenig, Ulrich; Karner, Susanne; Barresi, Caterina; Rossiter, Heidemarie; Narzt, Marie-Sophie; Nagelreiter, Ionela M; Larue, Lionel; Tobin, Desmond J; Eckhart, Leopold; Tschachler, Erwin

    2015-05-01

    Autophagy is the central cellular mechanism for delivering organelles and cytoplasm to lysosomes for degradation and recycling of their molecular components. To determine the contribution of autophagy to melanocyte (MC) biology, we inactivated the essential autophagy gene Atg7 specifically in MCs using the Cre-loxP system. This gene deletion efficiently suppressed a key step in autophagy, lipidation of microtubule-associated protein 1 light chain 3 beta (LC3), in MCs and induced slight hypopigmentation of the epidermis in mice. The melanin content of hair was decreased by 10-15% in mice with autophagy-deficient MC as compared with control animals. When cultured in vitro, MCs from mutant and control mice produced equal amounts of melanin per cell. However, Atg7-deficient MCs entered into premature growth arrest and accumulated reactive oxygen species (ROS) damage, ubiquitinated proteins, and the multi-functional adapter protein SQSTM1/p62. Moreover, nuclear factor erythroid 2-related factor 2 (Nrf2)-dependent expression of NAD(P)H dehydrogenase, quinone 1, and glutathione S-transferase Mu 1 was increased, indicating a contribution of autophagy to redox homeostasis in MCs. In summary, the results of our study suggest that Atg7-dependent autophagy is dispensable for melanogenesis but necessary for achieving the full proliferative capacity of MCs. PMID:25290687

  7. Autophagy activity is up-regulated in adipose tissue of obese individuals and modulates proinflammantory cytokine expression.

    NARCIS (Netherlands)

    Jansen, H.J.; Essen, van P.; Koenen, T.; Joosten, L.A.; Netea, M.G.; Tack, C.J.; Stienstra, R.

    2012-01-01

    Autophagy, an evolutionary conserved process aimed at recycling damaged organelles and protein aggregates in the cell, also modulates proinflammatory cytokine production in peripheral blood mononuclear cells. Because adipose tissue inflammation accompanied by elevated levels of proinflammatory cytok

  8. Autophagy activity is up-regulated in adipose tissue of obese individuals and modulates proinflammatory cytokine expression

    NARCIS (Netherlands)

    Jansen, H.J.; Essen, P. van; Koenen, T.; Joosten, L.A.B.; Netea, M.G.; Tack, C.J.J.; Stienstra, R.

    2012-01-01

    Autophagy, an evolutionary conserved process aimed at recycling damaged organelles and protein aggregates in the cell, also modulates proinflammatory cytokine production in peripheral blood mononuclear cells. Because adipose tissue inflammation accompanied by elevated levels of proinflammatory cytok

  9. Targeted deletion of Atg5 reveals differential roles of autophagy in keratin K5-expressing epithelia

    Energy Technology Data Exchange (ETDEWEB)

    Sukseree, Supawadee [Research Division of Biology and Pathobiology of the Skin, Department of Dermatology, Medical University of Vienna, Vienna (Austria); Department of Biochemistry, Faculty of Medicine, Srinakharinwirot University, Bangkok (Thailand); Rossiter, Heidemarie; Mildner, Michael [Research Division of Biology and Pathobiology of the Skin, Department of Dermatology, Medical University of Vienna, Vienna (Austria); Pammer, Johannes [Institute of Clinical Pathology, Medical University of Vienna, Vienna (Austria); Buchberger, Maria; Gruber, Florian [Research Division of Biology and Pathobiology of the Skin, Department of Dermatology, Medical University of Vienna, Vienna (Austria); Watanapokasin, Ramida [Department of Biochemistry, Faculty of Medicine, Srinakharinwirot University, Bangkok (Thailand); Tschachler, Erwin [Research Division of Biology and Pathobiology of the Skin, Department of Dermatology, Medical University of Vienna, Vienna (Austria); Eckhart, Leopold, E-mail: leopold.eckhart@meduniwien.ac.at [Research Division of Biology and Pathobiology of the Skin, Department of Dermatology, Medical University of Vienna, Vienna (Austria)

    2013-01-11

    Highlights: Black-Right-Pointing-Pointer We generated mice lacking Atg5 and autophagy in keratin K5-positive epithelia. Black-Right-Pointing-Pointer Suppression of autophagy in thymic epithelium was not associated with signs of autoimmunity. Black-Right-Pointing-Pointer Autophagy was required for normal terminal differentiation of preputial gland cells. Black-Right-Pointing-Pointer Autophagy-deficient cells of the preputial glands degraded nuclear DNA prematurely. -- Abstract: Autophagy contributes to the homeostasis of many tissues, yet its role in epithelia is incompletely understood. A recent report proposed that Atg5-dependent autophagy in thymic epithelial cells is essential for their function in the negative selection of self-reactive T-cells and, thus, for the suppression of tissue inflammation. Here we crossed mice carrying floxed alleles of the Atg5 gene with mice expressing the Cre recombinase under the control of the keratin K5 promoter to suppress autophagy in all K5-positive epithelia. The efficiency of autophagy abrogation was confirmed by immunoanalyses of LC3, which was converted to the autophagy-associated LC3-II form in normal but not Atg5-deficient cells, and of p62, which accumulated in Atg5-deficient cells. Mice carrying the epithelium-specific deletion of Atg5 showed normal weight gain, absence of tissue inflammation, and a normal morphology of the thymic epithelium. By contrast, autophagy-deficient epithelial cells of the preputial gland showed aberrant eosinophilic staining in histology and premature degradation of nuclear DNA during terminal differentiation. Taken together, the results of this study suggest that autophagy is dispensable for the suppression of autoimmunity by thymic epithelial cells but essential for normal differentiation of the preputial gland in mice.

  10. Targeted deletion of Atg5 reveals differential roles of autophagy in keratin K5-expressing epithelia

    International Nuclear Information System (INIS)

    Highlights: ► We generated mice lacking Atg5 and autophagy in keratin K5-positive epithelia. ► Suppression of autophagy in thymic epithelium was not associated with signs of autoimmunity. ► Autophagy was required for normal terminal differentiation of preputial gland cells. ► Autophagy-deficient cells of the preputial glands degraded nuclear DNA prematurely. -- Abstract: Autophagy contributes to the homeostasis of many tissues, yet its role in epithelia is incompletely understood. A recent report proposed that Atg5-dependent autophagy in thymic epithelial cells is essential for their function in the negative selection of self-reactive T-cells and, thus, for the suppression of tissue inflammation. Here we crossed mice carrying floxed alleles of the Atg5 gene with mice expressing the Cre recombinase under the control of the keratin K5 promoter to suppress autophagy in all K5-positive epithelia. The efficiency of autophagy abrogation was confirmed by immunoanalyses of LC3, which was converted to the autophagy-associated LC3-II form in normal but not Atg5-deficient cells, and of p62, which accumulated in Atg5-deficient cells. Mice carrying the epithelium-specific deletion of Atg5 showed normal weight gain, absence of tissue inflammation, and a normal morphology of the thymic epithelium. By contrast, autophagy-deficient epithelial cells of the preputial gland showed aberrant eosinophilic staining in histology and premature degradation of nuclear DNA during terminal differentiation. Taken together, the results of this study suggest that autophagy is dispensable for the suppression of autoimmunity by thymic epithelial cells but essential for normal differentiation of the preputial gland in mice.

  11. The role of autophagy in microbial infection and immunity

    Directory of Open Access Journals (Sweden)

    Desai M

    2015-01-01

    Full Text Available Mayura Desai,1 Rong Fang,2 Jiaren Sun11Department of Microbiology and Immunology, 2Department of Pathology, University of Texas Medical Branch at Galveston, Galveston, TX, USAAbstract: The autophagy pathway represents an evolutionarily conserved cell recycling process that is activated in response to nutrient deprivation and other stress signals. Over the years, it has been linked to an array of cellular functions. Equally, a wide range of cell-intrinsic, as well as extracellular, factors have been implicated in the induction of the autophagy pathway. Microbial infections represent one such factor that can not only activate autophagy through specific mechanisms but also manipulate the response to the invading microbe's advantage. Moreover, in many cases, particularly among viruses, the pathway has been shown to be intricately involved in the replication cycle of the pathogen. Conversely, autophagy also plays a role in combating the infection process, both through direct destruction of the pathogen and as one of the key mediating factors in the host defense mechanisms of innate and adaptive immunity. Further, the pathway also plays a role in controlling the pathogenesis of infectious diseases by regulating inflammation. In this review, we discuss various interactions between pathogens and the cellular autophagic response and summarize the immunological functions of the autophagy pathway.Keywords: autophagy, xenophagy, antiviral, antibacterial

  12. Autophagy as a Possible Underlying Mechanism of Nanomaterial Toxicity

    Directory of Open Access Journals (Sweden)

    Vanessa Cohignac

    2014-07-01

    Full Text Available The rapid development of nanotechnologies is raising safety concerns because of the potential effects of engineered nanomaterials on human health, particularly at the respiratory level. Since the last decades, many in vivo studies have been interested in the pulmonary effects of different classes of nanomaterials. It has been shown that some of them can induce toxic effects, essentially depending on their physico-chemical characteristics, but other studies did not identify such effects. Inflammation and oxidative stress are currently the two main mechanisms described to explain the observed toxicity. However, the exact underlying mechanism(s still remain(s unknown and autophagy could represent an interesting candidate. Autophagy is a physiological process in which cytoplasmic components are digested via a lysosomal pathway. It has been shown that autophagy is involved in the pathogenesis and the progression of human diseases, and is able to modulate the oxidative stress and pro-inflammatory responses. A growing amount of literature suggests that a link between nanomaterial toxicity and autophagy impairment could exist. In this review, we will first summarize what is known about the respiratory effects of nanomaterials and we will then discuss the possible involvement of autophagy in this toxicity. This review should help understand why autophagy impairment could be taken as a promising candidate to fully understand nanomaterials toxicity.

  13. Autophagy and lysosomal dysfunction as emerging mechanisms of nanomaterial toxicity

    Directory of Open Access Journals (Sweden)

    Stern Stephan T

    2012-06-01

    Full Text Available Abstract The study of the potential risks associated with the manufacture, use, and disposal of nanoscale materials, and their mechanisms of toxicity, is important for the continued advancement of nanotechnology. Currently, the most widely accepted paradigms of nanomaterial toxicity are oxidative stress and inflammation, but the underlying mechanisms are poorly defined. This review will highlight the significance of autophagy and lysosomal dysfunction as emerging mechanisms of nanomaterial toxicity. Most endocytic routes of nanomaterial cell uptake converge upon the lysosome, making the lysosomal compartment the most common intracellular site of nanoparticle sequestration and degradation. In addition to the endo-lysosomal pathway, recent evidence suggests that some nanomaterials can also induce autophagy. Among the many physiological functions, the lysosome, by way of the autophagy (macroautophagy pathway, degrades intracellular pathogens, and damaged organelles and proteins. Thus, autophagy induction by nanoparticles may be an attempt to degrade what is perceived by the cell as foreign or aberrant. While the autophagy and endo-lysosomal pathways have the potential to influence the disposition of nanomaterials, there is also a growing body of literature suggesting that biopersistent nanomaterials can, in turn, negatively impact these pathways. Indeed, there is ample evidence that biopersistent nanomaterials can cause autophagy and lysosomal dysfunctions resulting in toxicological consequences.

  14. Lithium and autophagy.

    Science.gov (United States)

    Motoi, Yumiko; Shimada, Kohei; Ishiguro, Koichi; Hattori, Nobutaka

    2014-06-18

    Lithium, a drug used to treat bipolar disorders, has a variety of neuroprotective mechanisms, including autophagy regulation, in various neuropsychiatric conditions. In neurodegenerative diseases, lithium enhances degradation of aggregate-prone proteins, including mutated huntingtin, phosphorylated tau, and α-synuclein, and causes damaged mitochondria to degrade, while in a mouse model of cerebral ischemia and Alzheimer's disease autophagy downregulation by lithium is observed. The signaling pathway of lithium as an autophagy enhancer might be associated with the mammalian target of rapamycin (mTOR)-independent pathway, which is involved in myo-inositol-1,4,5-trisphosphate (IP3) in Huntington's disease and Parkinson's disease. However, the mTOR-dependent pathway might be involved in inhibiting glycogen synthase kinase-3β (GSK3β) in other diseases. Lithium's autophagy-enhancing property may contribute to the therapeutic benefit of patients with neuropsychiatric disorders. PMID:24738557

  15. The ubiquitin-proteasome system

    Indian Academy of Sciences (India)

    Dipankar Nandi; Pankaj Tahiliani; Anujith Kumar; Dilip Chandu

    2006-03-01

    The 2004 Nobel Prize in chemistry for the discovery of protein ubiquitination has led to the recognition of cellular proteolysis as a central area of research in biology. Eukaryotic proteins targeted for degradation by this pathway are first ‘tagged’ by multimers of a protein known as ubiquitin and are later proteolyzed by a giant enzyme known as the proteasome. This article recounts the key observations that led to the discovery of ubiquitin-proteasome system (UPS). In addition, different aspects of proteasome biology are highlighted. Finally, some key roles of the UPS in different areas of biology and the use of inhibitors of this pathway as possible drug targets are discussed.

  16. Autophagy research: Lessons from metabolism

    NARCIS (Netherlands)

    A.J. Meijer

    2009-01-01

    Autophagy research continues to expand exponentially. Clearly autophagy and metabolism are intimately connected; however, the rapid expansion of research into this topic inevitably brings the risk that important basic knowledge of metabolism will be overlooked when considering experimental data. Unf

  17. [Advances in the study of the relationship between autophagy and sepsis-induced lung injury].

    Science.gov (United States)

    Wang, Xingtong; Li, Hengyu; Xia, Zhaofan

    2014-08-01

    Sepsis is one of the most common pathogenetic causes of acute lung injury (ALI), and at present there is still a lack of effective targeted techniques and methods for its prevention and treatment. Autophagy is a homeostatic mecha- nism common to all eukaryotic cells, including adaption to environment, defense against invasion of pathogens, and maintenance of cellular homeostasis. Autophagy is also involved in a variety of lung-related diseases. In septic lung injury, autophagy not only serves to dissipate dysfunctional organelles, but also inhibits the release of inflammatory cytokines. This review aims at eliciting the role of autophagy in sepsis-induced ALI and further exploring the potential targets of autophagy in inhibiting inflammation, in an effort to provide a new perspective for clinical treatment of sepsis-induced ALI.

  18. New Insights Into Roles of Ubiquitin Modification in Regulating Plastids and Other Endosymbiotic Organelles.

    Science.gov (United States)

    Broad, W; Ling, Q; Jarvis, P

    2016-01-01

    Recent findings have revealed important and diverse roles for the ubiquitin modification of proteins in the regulation of endosymbiotic organelles, which include the primary plastids of plants as well as complex plastids: the secondary endosymbiotic organelles of cryptophytes, alveolates, stramenopiles, and haptophytes. Ubiquitin modifications have a variety of potential consequences, both to the modified protein itself and to cellular regulation. The ubiquitin-proteasome system (UPS) can target individual proteins for selective degradation by the cytosolic 26S proteasome. Ubiquitin modifications can also signal the removal of whole endosymbiotic organelles, for example, via autophagy as has been well characterized in mitochondria. As plastids must import over 90% of their proteins from the cytosol, the observation that the UPS selectively targets the plastid protein import machinery is particularly significant. In this way, the UPS may influence the development and interconversions of different plastid types, as well as plastid responses to stress, by reconfiguring the organellar proteome. In complex plastids, the Symbiont-derived ERAD-Like Machinery (SELMA) has coopted the protein transport capabilities of the ER-Associated Degradation (ERAD) system, whereby misfolded proteins are retrotranslocated from ER for proteasomal degradation, uncoupling them from proteolysis: SELMA components have been retargeted to the second outermost plastid membrane to mediate protein import. In spite of this wealth of new information, there still remain a large number of unanswered questions and a need to define the roles of ubiquitin modification further in the regulation of plastids. PMID:27241217

  19. Receptor Proteins in Selective Autophagy

    Directory of Open Access Journals (Sweden)

    Christian Behrends

    2012-01-01

    Full Text Available Autophagy has long been thought to be an essential but unselective bulk degradation pathway. However, increasing evidence suggests selective autophagosomal turnover of a broad range of substrates. Bifunctional autophagy receptors play a key role in selective autophagy by tethering cargo to the site of autophagosomal engulfment. While the identity of molecular components involved in selective autophagy has been revealed at least to some extent, we are only beginning to understand how selectivity is achieved in this process. Here, we summarize the mechanistic and structural basis of receptor-mediated selective autophagy.

  20. Selective autophagy of non-ubiquitylated targets in plants: looking for cognate receptor/adaptor proteins

    Directory of Open Access Journals (Sweden)

    Vasko eVeljanovski

    2014-06-01

    Full Text Available Cellular homeostasis is essential for the physiology of eukaryotic cells. Eukaryotic cells, including plant cells, utilize two main pathways to adjust the level of cytoplasmic components, namely the proteasomal and the lysosomal/vacuolar pathways. Macroautophagy is a lysosomal/vacuolar pathway which, until recently, was thought to be non-specific and a bulk degradation process. However, selective autophagy which can be activated in the cell under various physiological conditions, involves the specific degradation of defined macromolecules or organelles by a conserved molecular mechanism. For this process to be efficient, the mechanisms underlying the recognition and selection of the cargo to be engulfed by the double-membrane autophagosome are critical, and not yet well understood. Ubiquitin (poly-ubiquitin conjugation to the target appears to be a conserved ligand mechanism in many types of selective autophagy, and defined receptors/adaptors recognizing and regulating the autophagosomal capture of the ubiquitylated target have been characterized. However, non-proteinaceous and non-ubiquitylated cargoes are also selectively degraded by this pathway. This ubiquitin-independent selective autophagic pathway also involves receptor and/or adaptor proteins linking the cargo to the autophagic machinery. Some of these receptor/adaptor proteins including accessory autophagy-related (Atg and non-Atg proteins have been described in yeast and animal cells but not yet in plants. In this review we discuss the ubiquitin-independent cargo selection mechanisms in selective autophagy degradation of organelles and macromolecules and speculate on potential plant receptor/adaptor proteins.

  1. 细胞自噬与肿瘤耐药的研究进展%Recent progress of autophagy and drug-resistance

    Institute of Scientific and Technical Information of China (English)

    叶丽霖; 李玉峰

    2013-01-01

    Summary Autophagy is a self degradation process that are responsible for the removal of long lived proteins and damaged organelles, which plays a key role in the elimination of such intracellular waste, reconstruction of structure,growth and development of cells and maintainance of cellular homeostasis. During tumour development, autophagy has paradoxically been reported to have roles in promoting both cell survival and growth. In tumour cells with defects in apoptosis, autophagy allows prolonged survival. Recent evidence suggests that autophagy provides a protective function to limit tumour necrosis and inflammation,and to mitigate genome damage in tumour cells in response to metabolic stress. Many studies show that many cytoxic drugs can induce autophagy in its induction of apoptosis. Activation of autophagy can lead to drug resistance in tumour cells. In this review, we will summarize the relationship between autophagy and drug resistance.

  2. Mechanism of action of the tuberculosis and Crohn disease risk factor IRGM in autophagy

    Science.gov (United States)

    Chauhan, Santosh; Mandell, Michael A.; Deretic, Vojo

    2016-01-01

    ABSTRACT Polymorphisms in the IRGM gene, associated with Crohn disease (CD) and tuberculosis, are among the earliest identified examples documenting the role of autophagy in human disease. Functional studies have shown that IRGM protects against these diseases by modulating autophagy, yet the exact molecular mechanism of IRGM's activity has remained unknown. We have recently elucidated IRGM's mechanism of action. IRGM functions as a platform for assembling, stabilizing, and activating the core autophagic machinery, while at the same time physically coupling it to conventional innate immunity receptors. Exposure to microbial products or bacterial invasion increases IRGM expression, which leads to stabilization of AMPK. Specific protein-protein interactions and post-translational modifications such as ubiquitination of IRGM, lead to a co-assembly with IRGM of the key autophagy regulators ULK1 and BECN1 in their activated forms. IRGM physically interacts with 2 other CD risk factors, ATG16L1 and NOD2, placing these 3 principal players in CD within the same molecular complex. This explains how polymorphisms altering expression or function of any of the 3 factors individually can affect the same process—autophagy. Furthermore, IRGM's interaction with NOD2, and additional pattern recognition receptors such as NOD1, RIG-I, and select TLRs, transduces microbial signals to the core autophagy apparatus. This work solves the long-standing enigma of how IRGM controls autophagy. PMID:26313894

  3. The Deubiquitinating Enzyme UBPY Is Required for Lysosomal Biogenesis and Productive Autophagy in Drosophila.

    Directory of Open Access Journals (Sweden)

    Anne-Claire Jacomin

    Full Text Available Autophagy is a catabolic process that delivers cytoplasmic components to the lysosomes. Protein modification by ubiquitination is involved in this pathway: it regulates the stability of autophagy regulators such as BECLIN-1 and it also functions as a tag targeting specific substrates to autophagosomes. In order to identify deubiquitinating enzymes (DUBs involved in autophagy, we have performed a genetic screen in the Drosophila larval fat body. This screen identified Uch-L3, Usp45, Usp12 and Ubpy. In this paper, we show that Ubpy loss of function results in the accumulation of autophagosomes due to a blockade of the autophagy flux. Furthermore, analysis by electron and confocal microscopy of Ubpy-depleted fat body cells revealed altered lysosomal morphology, indicating that Ubpy inactivation affects lysosomal maintenance and/or biogenesis. Lastly, we have shown that shRNA mediated inactivation of UBPY in HeLa cells affects autophagy in a different way: in UBPY-depleted HeLa cells autophagy is deregulated.

  4. Autophagy regulates keratin 8 homeostasis in mammary epithelial cells and in breast tumors

    Science.gov (United States)

    Kongara, Sameera; Kravchuk, Olga; Teplova, Irina; Lozy, Fred; Schulte, Jennifer; Moore, Dirk; Barnard, Nicola; Neumann, Carola A.; White, Eileen; Karantza, Vassiliki

    2010-01-01

    Autophagy is activated in response to cellular stressors and mediates lysosomal degradation and recycling of cytoplasmic material and organelles as a temporary cell survival mechanism. Defective autophagy is implicated in human pathology, as disruption of protein and organelle homeostasis enables disease-promoting mechanisms such as toxic protein aggregation, oxidative stress, genomic damage and inflammation. We previously showed that autophagy-defective immortalized mouse mammary epithelial cells (iMMECs) are susceptible to metabolic stress, DNA damage and genomic instability. We now report that autophagy deficiency was associated with ER and oxidative stress, and deregulation of p62-mediated keratin homeostasis in mammary cells and allograft tumors and in mammary tissues from genetically engineered mice. In human breast tumors, high phospho(Ser73)-K8 levels inversely correlated with Beclin 1 expression. Thus, autophagy preserves cellular fitness by limiting ER and oxidative stress, a function potentially important in autophagy-mediated suppression of mammary tumorigenesis. Furthermore, autophagy regulates keratin homeostasis in the mammary gland via a p62-dependent mechanism. High phospho(Ser73)-K8 expression may be a marker of autophagy functional status in breast tumors and, as such, could have therapeutic implications for breast cancer patients. PMID:20530580

  5. DNA damage and autophagy

    Energy Technology Data Exchange (ETDEWEB)

    Rodriguez-Rocha, Humberto; Garcia-Garcia, Aracely [Redox Biology Center and School of Veterinary Medicine and Biomedical Sciences, University of Nebraska-Lincoln, Lincoln, NE 68583 (United States); Panayiotidis, Mihalis I. [School of Community Health Sciences, University of Nevada, Reno, NV 89557 (United States); Franco, Rodrigo, E-mail: rfrancocruz2@unl.edu [Redox Biology Center and School of Veterinary Medicine and Biomedical Sciences, University of Nebraska-Lincoln, Lincoln, NE 68583 (United States)

    2011-06-03

    Both exogenous and endogenous agents are a threat to DNA integrity. Exogenous environmental agents such as ultraviolet (UV) and ionizing radiation, genotoxic chemicals and endogenous byproducts of metabolism including reactive oxygen species can cause alterations in DNA structure (DNA damage). Unrepaired DNA damage has been linked to a variety of human disorders including cancer and neurodegenerative disease. Thus, efficient mechanisms to detect DNA lesions, signal their presence and promote their repair have been evolved in cells. If DNA is effectively repaired, DNA damage response is inactivated and normal cell functioning resumes. In contrast, when DNA lesions cannot be removed, chronic DNA damage triggers specific cell responses such as cell death and senescence. Recently, DNA damage has been shown to induce autophagy, a cellular catabolic process that maintains a balance between synthesis, degradation, and recycling of cellular components. But the exact mechanisms by which DNA damage triggers autophagy are unclear. More importantly, the role of autophagy in the DNA damage response and cellular fate is unknown. In this review we analyze evidence that supports a role for autophagy as an integral part of the DNA damage response.

  6. Dopamine Oxidation and Autophagy

    Directory of Open Access Journals (Sweden)

    Patricia Muñoz

    2012-01-01

    Full Text Available The molecular mechanisms involved in the neurodegenerative process of Parkinson's disease remain unclear. Currently, there is a general agreement that mitochondrial dysfunction, α-synuclein aggregation, oxidative stress, neuroinflammation, and impaired protein degradation are involved in the neurodegeneration of dopaminergic neurons containing neuromelanin in Parkinson's disease. Aminochrome has been proposed to play an essential role in the degeneration of dopaminergic neurons containing neuromelanin by inducing mitochondrial dysfunction, oxidative stress, the formation of neurotoxic α-synuclein protofibrils, and impaired protein degradation. Here, we discuss the relationship between the oxidation of dopamine to aminochrome, the precursor of neuromelanin, autophagy dysfunction in dopaminergic neurons containing neuromelanin, and the role of dopamine oxidation to aminochrome in autophagy dysfunction in dopaminergic neurons. Aminochrome induces the following: (i the formation of α-synuclein protofibrils that inactivate chaperone-mediated autophagy; (ii the formation of adducts with α- and β-tubulin, which induce the aggregation of the microtubules required for the fusion of autophagy vacuoles and lysosomes.

  7. Autophagy studies in Bombyx mori

    Directory of Open Access Journals (Sweden)

    L Tian

    2015-03-01

    Full Text Available Autophagy, which is well conserved from yeast to mammals, plays essential roles in development and diseases. Using the domesticated silkworm, Bombyx mori, as a model insect, several reports on autophagy have been made recently. Autophagic features are observed in the midgut and fat body during the larval-pupal transition as well as the silk gland and ovarian nurse cells during the pupal stage. There are 14 autophagy related (Atg genes, including at least two transcript variants of Atg1, predicated in Bombyx. Expression of most Atg genes is consistent with the autophagy process in the fat body during the larval-pupal transition, and reduction of Atg1 expression by RNAi blocks this process. The molting hormone, 20-hydroxyecdysone (20E, and starvation induce autophagy in the fat body by upregulating Atg gene expression and blocking the PI3K-TORC1 pathway. Meanwhile, autophagy precedes apoptosis in the midgut and other larval tissues during the larval-pupal transition, while the detailed mechanism is not illustrated yet. We assume that there are at least four future directions about autophagy studies in Bombyx during the next years: (1 physiological functions of autophagy; (2 identification of new components involved in the autophagy process; (3 detailed molecular mechanism of autophagosome formation; (4 functional relationship between autophagy and apoptosis.

  8. Ubiquitin and proteasomes in transcription.

    Science.gov (United States)

    Geng, Fuqiang; Wenzel, Sabine; Tansey, William P

    2012-01-01

    Regulation of gene transcription is vitally important for the maintenance of normal cellular homeostasis. Failure to correctly regulate gene expression, or to deal with problems that arise during the transcription process, can lead to cellular catastrophe and disease. One of the ways cells cope with the challenges of transcription is by making extensive use of the proteolytic and nonproteolytic activities of the ubiquitin-proteasome system (UPS). Here, we review recent evidence showing deep mechanistic connections between the transcription and ubiquitin-proteasome systems. Our goal is to leave the reader with a sense that just about every step in transcription-from transcription initiation through to export of mRNA from the nucleus-is influenced by the UPS and that all major arms of the system--from the first step in ubiquitin (Ub) conjugation through to the proteasome-are recruited into transcriptional processes to provide regulation, directionality, and deconstructive power. PMID:22404630

  9. In Vitro Ubiquitination: Self-Ubiquitination, Chain Formation, and Substrate Ubiquitination Assays.

    Science.gov (United States)

    Maspero, Elena; Polo, Simona

    2016-01-01

    Ubiquitination of proteins in vitro has evolved as an indispensable tool for the functional analysis of this posttranslational modification. In vitro ubiquitination is particularly helpful to study conjugation mechanisms. The efficiency of the ubiquitination reaction depends in part on the quality of the enzymes utilized. Here we introduce the assay developed in our lab to study HECT E3 ligases. It involves bacterially expressed E1, His-tagged Ube2D3 (also called UbcH5c, the best E2 for Nedd4), untagged Nedd4, and untagged ubiquitin (Ub). As tags may impair specific activity of the enzymes or even interfere with the enzymatic reaction, they should be avoided, removed, or kept to a minimal size whenever possible, unless proven to be without consequence. The protocol described here is suitable for other E3 ligases capable of forming Ub chains as pseudo-product of the enzyme reaction. It is also adapted to include substrates. In this case, substrates should be tagged and purified after the reaction is completed to allow the detection of the ubiquitinated products. PMID:27613033

  10. Rapamycin Improves Palmitate-Induced ER Stress/NF κ B Pathways Associated with Stimulating Autophagy in Adipocytes

    Directory of Open Access Journals (Sweden)

    Jiajing Yin

    2015-01-01

    Full Text Available Obesity-induced endoplasmic reticulum (ER stress and inflammation lead to adipocytes dysfunction. Autophagy helps to adapt to cellular stress and involves in regulating innate inflammatory response. In present study, we examined the activity of rapamycin, a mTOR kinase inhibitor, against endoplasmic reticulum stress and inflammation in adipocytes. An in vitro model was used in which 3T3-L1 adipocytes were preloaded with palmitate (PA to generate artificial hypertrophy mature adipocytes. Elevated autophagy flux and increased number of autophagosomes were observed in response to PA and rapamycin treatment. Rapamycin attenuated PA-induced PERK and IRE1-associated UPR pathways, evidenced by decreased protein levels of eIF2α phosphorylation, ATF4, CHOP, and JNK phosphorylation. Inhibiting autophagy with chloroquine (CQ exacerbated these ER stress markers, indicating the role of autophagy in ameliorating ER stress. In addition, cotreatment of CQ abolished the anti-ER stress effects of rapamycin, which confirms the effect of rapamycin on ERs is autophagy-dependent. Furthermore, rapamycin decreased PA-induced nuclear translocation of NFκB P65 subunit, thereby NFκB-dependent inflammatory cytokines MCP-1 and IL-6 expression and secretion. In conclusion, rapamycin attenuated PA-induced ER stress/NFκB pathways to counterbalance adipocytes stress and inflammation. The beneficial of rapamycin in this context partly depends on autophagy. Stimulating autophagy may become a way to attenuate adipocytes dysfunction.

  11. Rapamycin improves palmitate-induced ER stress/NF κ B pathways associated with stimulating autophagy in adipocytes.

    Science.gov (United States)

    Yin, Jiajing; Gu, Liping; Wang, Yufan; Fan, Nengguang; Ma, Yuhang; Peng, Yongde

    2015-01-01

    Obesity-induced endoplasmic reticulum (ER) stress and inflammation lead to adipocytes dysfunction. Autophagy helps to adapt to cellular stress and involves in regulating innate inflammatory response. In present study, we examined the activity of rapamycin, a mTOR kinase inhibitor, against endoplasmic reticulum stress and inflammation in adipocytes. An in vitro model was used in which 3T3-L1 adipocytes were preloaded with palmitate (PA) to generate artificial hypertrophy mature adipocytes. Elevated autophagy flux and increased number of autophagosomes were observed in response to PA and rapamycin treatment. Rapamycin attenuated PA-induced PERK and IRE1-associated UPR pathways, evidenced by decreased protein levels of eIF2α phosphorylation, ATF4, CHOP, and JNK phosphorylation. Inhibiting autophagy with chloroquine (CQ) exacerbated these ER stress markers, indicating the role of autophagy in ameliorating ER stress. In addition, cotreatment of CQ abolished the anti-ER stress effects of rapamycin, which confirms the effect of rapamycin on ERs is autophagy-dependent. Furthermore, rapamycin decreased PA-induced nuclear translocation of NFκB P65 subunit, thereby NFκB-dependent inflammatory cytokines MCP-1 and IL-6 expression and secretion. In conclusion, rapamycin attenuated PA-induced ER stress/NFκB pathways to counterbalance adipocytes stress and inflammation. The beneficial of rapamycin in this context partly depends on autophagy. Stimulating autophagy may become a way to attenuate adipocytes dysfunction.

  12. Non-degradative Ubiquitination of Protein Kinases.

    OpenAIRE

    K Aurelia Ball; Johnson, Jeffrey R.; Lewinski, Mary K; John Guatelli; Erik Verschueren; Krogan, Nevan J.; Matthew P Jacobson

    2016-01-01

    Growing evidence supports other regulatory roles for protein ubiquitination in addition to serving as a tag for proteasomal degradation. In contrast to other common post-translational modifications, such as phosphorylation, little is known about how non-degradative ubiquitination modulates protein structure, dynamics, and function. Due to the wealth of knowledge concerning protein kinase structure and regulation, we examined kinase ubiquitination using ubiquitin remnant immunoaffinity enrichm...

  13. Autophagy During Cardiac Stress: Joys and Frustrations of Autophagy

    Science.gov (United States)

    Gottlieb, Roberta A.; Mentzer, Robert M.

    2013-01-01

    The study of autophagy has been transformed by the cloning of most genes in the pathway and the introduction of GFP-LC3 as a reporter to allow visual assessment of autophagy. The field of cardiac biology is not alone in attempting to understand the implications of autophagy. The purpose of this review is to address some of the controversies and conundrums associated with the evolving studies of autophagy in the heart. Autophagy is a cellular process involving a complex orchestration of regulatory gene products as well as machinery for assembly, selective targeting, and degradation of autophagosomes and their contents. Our understanding of the role of autophagy in human disease is rapidly evolving as investigators examine the process in different tissues and different pathophysiological contexts. In the field of heart disease, autophagy has been examined in the settings of ischemia and reperfusion, preconditioning, cardiac hypertrophy, and heart failure. This review addresses the role of autophagy in cardioprotection, the balance of catabolism and anabolism, the concept of mitochondrial quality control, and the implications of impaired autophagic flux or frustrated autophagy. PMID:20148666

  14. Posttranslational modification of autophagy-related proteins in macroautophagy.

    Science.gov (United States)

    Xie, Yangchun; Kang, Rui; Sun, Xiaofang; Zhong, Meizuo; Huang, Jin; Klionsky, Daniel J; Tang, Daolin

    2015-01-01

    Macroautophagy is an intracellular catabolic process involved in the formation of multiple membrane structures ranging from phagophores to autophagosomes and autolysosomes. Dysfunction of macroautophagy is implicated in both physiological and pathological conditions. To date, 38 autophagy-related (ATG) genes have been identified as controlling these complicated membrane dynamics during macroautophagy in yeast; approximately half of these genes are clearly conserved up to human, and there are additional genes whose products function in autophagy in higher eukaryotes that are not found in yeast. The function of the ATG proteins, in particular their ability to interact with a number of macroautophagic regulators, is modulated by posttranslational modifications (PTMs) such as phosphorylation, glycosylation, ubiquitination, acetylation, lipidation, and proteolysis. In this review, we summarize our current knowledge of the role of ATG protein PTMs and their functional relevance in macroautophagy. Unraveling how these PTMs regulate ATG protein function during macroautophagy will not only reveal fundamental mechanistic insights into the regulatory process, but also provide new therapeutic targets for the treatment of autophagy-associated diseases.

  15. Role of autophagy in the pathogenesis of amyotrophic lateral sclerosis.

    Science.gov (United States)

    Lee, Jae Keun; Shin, Jin Hee; Lee, Ji Eun; Choi, Eui-Ju

    2015-11-01

    Amyotrophic lateral sclerosis (ALS) is a late-onset neurodegenerative disease characterized by the selective degeneration of upper and lower motor neurons associated with the abnormal aggregation of ubiquitinated proteins. The molecular mechanisms underlying the pathogenesis of ALS remain unclear, however. Autophagy is a major pathway for the elimination of protein aggregates and damaged organelles and therefore contributes to cellular homeostasis. This catabolic process begins with the formation of the double membrane-bound autophagosome that engulfs portions of the cytoplasm and subsequently fuses with a lysosome to form an autolysosome, in which lysosomal enzymes digest autophagic substrates. Defects at various stages of autophagy have been associated with pathological mutations of several ALS-linked genes including SOD1, p62, TDP-43, and optineurin, suggesting that such defects may play a causative role in the pathogenesis of this condition. In this review, we summarize the dysregulation of autophagy associated with ALS as well as potential therapeutic strategies based on modulation of the autophagic process. PMID:26264610

  16. Inhibition of glycogen synthase kinase 3β promotes autophagy to protect mice from acute liver failure mediated by peroxisome proliferator-activated receptor α

    OpenAIRE

    Ren, F.; Zhang, L; Zhang, X; Shi, H; T. Wen; Bai, L.; S. Zheng; Y. Chen; Chen, D.; Li, L.; Duan, Z

    2016-01-01

    Our previous studies have demonstrated that inhibition of glycogen synthase kinase 3β (GSK3β) activity protects mice from acute liver failure (ALF), whereas its protective and regulatory mechanism remains elusive. Autophagy is a recently recognized rudimentary cellular response to inflammation and injury. The aim of the present study was to test the hypothesis that inhibition of GSK3β mediates autophagy to inhibit liver inflammation and protect against ALF. In ALF mice model induced by d-gala...

  17. Autophagy in plants and phytopathogens.

    Science.gov (United States)

    Yoshimoto, Kohki; Takano, Yoshitaka; Sakai, Yasuyoshi

    2010-04-01

    Plants and plant-associated microorganisms including phytopathogens have to adapt to drastic changes in environmental conditions. Because of their immobility, plants must cope with various types of environmental stresses such as starvation, oxidative stress, drought stress, and invasion by phytopathogens during their differentiation, development, and aging processes. Here we briefly describe the early studies of plant autophagy, summarize recent studies on the molecular functions of ATG genes, and speculate on the role of autophagy in plants and phytopathogens. Autophagy regulates senescence and pathogen-induced cell death in plants, and autophagy and pexophagy play critical roles in differentiation and the invasion of host cells by phytopathogenic fungi. PMID:20079356

  18. Autophagy to Survive

    Directory of Open Access Journals (Sweden)

    Muzeyyen Izmirli

    2014-06-01

    Full Text Available Autophagy is the catabolic mechanism that involves cell degradation of unnecessary or dysfunctional cellular components through the actions of lysosomes. It helps to keep the cells alive in such cases like oxidative stress, lack of nutrients and growth factors providing recycling of intracellular molecules. However, it works as a part of metabolism regulation, morphogenesis, cell differentiation, senescence, cell death and immune system. As a result of impairment of this mechanism, pathological situations arise including cancer, neurodegenerative and infectious diseases. Consequently, researches about autophagy mechanism are important for the development of novel diagnosis, follow-up and treatment modalities in health problems. [Archives Medical Review Journal 2014; 23(3.000: 411-419

  19. Autophagy in cardiovascular biology

    OpenAIRE

    Lavandero, Sergio; Chiong, Mario; Rothermel, Beverly A.; Hill, Joseph A.

    2015-01-01

    Cardiovascular disease is the leading cause of death worldwide. As such, there is great interest in identifying novel mechanisms that govern the cardiovascular response to disease-related stress. First described in failing hearts, autophagy within the cardiovascular system has been widely characterized in cardiomyocytes, cardiac fibroblasts, endothelial cells, vascular smooth muscle cells, and macrophages. In all cases, a window of optimal autophagic activity appears to be critical to the mai...

  20. AUTOPHAGY IN LUNG CANCER

    OpenAIRE

    Jaboin, Jerry J.; Hwang, Misun; Lu, Bo

    2009-01-01

    Lung cancer is the leading cause of cancer-related deaths worldwide. The relatively poor cure rate in lung cancer patients has been associated with a resistance to chemotherapy and radiation that is at least in part related to defects in cellular apoptotic machinery. Exploitation of another form of cell death, autophagy, has the capacity to improve the therapeutic gain of current therapies. In an effort to develop novel treatment strategies to enhance the therapeutic ratio for lung cancer, we...

  1. Bcl-2-dependent upregulation of autophagy by sequestosome 1/p62 in vitro

    Institute of Scientific and Technical Information of China (English)

    Liang ZHOU; Hong-feng WANG; Hai-gang REN; Dong CHEN; Feng GAO; Qing-song HU; Chen FU

    2013-01-01

    To investigate whether sequestosome 1/p62 (p62),a key cargo adaptor protein involved in both the ubiquitin-proteasome system and the autophagy-lysosome system,could directly regulate autophagy in vitro.Methods:HEK 293 cells or HeLa cells were transfected with p62-expressing plasmids or siRNA targeting p62.The cells or the cell lysates were subsequently subjected to immunofluorescence assay,immunoprecipitation assay,or immunoblot analysis.In vitro pulldown assay was used to study the interaction of p62 with Bcl-2.Results:Overexpression of p62 significantly increased the basal level of autophagy in both HEK 293 cells and HeLa cells,whereas knockdown of p62 significantly decreased the basal level of autophagy.In vitro pulldown assay showed that p62 directly interacted with Bcl-2.It was observed in HeLa cells that p62 co-localized with Bcl-2.Furthermore,knockdown of p62 in HEK 293 cells significantly increased the amount of Beclin 1 that co-immunoprecipitated with Bcl-2.Conclusion:p62 induces autophagy by disrupting the association between Bcl-2 and Beclin 1.

  2. Relationship between autophagy and aging%细胞自噬与衰老

    Institute of Scientific and Technical Information of China (English)

    席兴字

    2011-01-01

    Aging is an intrinsic property of organism, which is characterized by the accumulation of damaged protein aggregates in cells and tissues. Clearance of protein aggregates mainly relies on ubiquitin-proteasome system and lysosome-dependent autophagy pathway.In recent years, some autophagy genes have been reported to play an important role in aging in models organisms, and these findings motivate interests in developing anti-aging drugs. This paper summarized the mechanisms of autophagy, and discussed the connections between aging and autophagy.%衰老是生物体内在的性质,细胞中损伤蛋白质积累是生物体衰老的一个重要特征.蛋门质聚集体的清除主要依赖于泛素-蛋白酶体途径和依赖溶酶体的细胞自噬(autophagy)途径.近年来,研究人员在许多模式生物中发现一些细胞自噬基因在衰老中起重要作用,这些研究激发人们利用细胞自噬开发抗衰老药物的兴趣.本文就细胞自噬的机理、衰老与自噬之间的联系等进行综述.

  3. Regulation of Autophagy by Kinases

    Energy Technology Data Exchange (ETDEWEB)

    Sridharan, Savitha; Jain, Kirti; Basu, Alakananda, E-mail: alakananda.basu@unthsc.edu [Department of Molecular Biology and Immunology, Institute for Cancer Research, University of North Texas Health Science Center, Fort Worth, TX 76107 (United States)

    2011-06-09

    Autophagy is a process of self-degradation that maintains cellular viability during periods of metabolic stress. Although autophagy is considered a survival mechanism when faced with cellular stress, extensive autophagy can also lead to cell death. Aberrations in autophagy are associated with several diseases, including cancer. Therapeutic exploitation of this process requires a clear understanding of its regulation. Although the core molecular components involved in the execution of autophagy are well studied there is limited information on how cellular signaling pathways, particularly kinases, regulate this complex process. Protein kinases are integral to the autophagy process. Atg1, the first autophagy-related protein identified, is a serine/threonine kinase and it is regulated by another serine/threonine kinase mTOR. Emerging studies suggest the participation of many different kinases in regulating various components/steps of this catabolic process. This review focuses on the regulation of autophagy by several kinases with particular emphasis on serine/threonine protein kinases such as mTOR, AMP-activated protein kinase, Akt, mitogen-activated protein kinase (ERK, p38 and JNK) and protein kinase C that are often deregulated in cancer and are important therapeutic targets.

  4. Autophagy: Regulation by Energy Sensing

    NARCIS (Netherlands)

    A.J. Meijer; P. Codogno

    2011-01-01

    Autophagy is inhibited by the mTOR signaling pathway, which is stimulated by increased amino acid levels. When cellular energy production is compromised, AMP-activated protein kinase is activated, mTOR is inhibited and autophagy is stimulated. Two recent studies have shed light on the molecular mech

  5. A genetic screen for modifiers of Drosophila caspase Dcp-1 reveals caspase involvement in autophagy and novel caspase-related genes

    Directory of Open Access Journals (Sweden)

    Ahnn Joohong

    2010-01-01

    Full Text Available Abstract Background Caspases are cysteine proteases with essential functions in the apoptotic pathway; their proteolytic activity toward various substrates is associated with the morphological changes of cells. Recent reports have described non-apoptotic functions of caspases, including autophagy. In this report, we searched for novel modifiers of the phenotype of Dcp-1 gain-of-function (GF animals by screening promoter element- inserted Drosophila melanogaster lines (EP lines. Results We screened ~15,000 EP lines and identified 72 Dcp-1-interacting genes that were classified into 10 groups based on their functions and pathways: 4 apoptosis signaling genes, 10 autophagy genes, 5 insulin/IGF and TOR signaling pathway genes, 6 MAP kinase and JNK signaling pathway genes, 4 ecdysone signaling genes, 6 ubiquitination genes, 11 various developmental signaling genes, 12 transcription factors, 3 translation factors, and 11 other unclassified genes including 5 functionally undefined genes. Among them, insulin/IGF and TOR signaling pathway, MAP kinase and JNK signaling pathway, and ecdysone signaling are known to be involved in autophagy. Together with the identification of autophagy genes, the results of our screen suggest that autophagy counteracts Dcp-1-induced apoptosis. Consistent with this idea, we show that expression of eGFP-Atg5 rescued the eye phenotype caused by Dcp-1 GF. Paradoxically, we found that over-expression of full-length Dcp-1 induced autophagy, as Atg8b-GFP, an indicator of autophagy, was increased in the eye imaginal discs and in the S2 cell line. Taken together, these data suggest that autophagy suppresses Dcp-1-mediated apoptotic cell death, whereas Dcp-1 positively regulates autophagy, possibly through feedback regulation. Conclusions We identified a number of Dcp-1 modifiers that genetically interact with Dcp-1-induced cell death. Our results showing that Dcp-1 and autophagy-related genes influence each other will aid future

  6. Retroperitoneal inflammation

    Science.gov (United States)

    ... page: //medlineplus.gov/ency/article/001255.htm Retroperitoneal inflammation To use the sharing features on this page, please enable JavaScript. Retroperitoneal inflammation is swelling that occurs in the retroperitoneal space. ...

  7. Role and Regulation of Autophagy in Heat Stress Responses of Tomato Plants

    Directory of Open Access Journals (Sweden)

    Jie eZhou

    2014-04-01

    Full Text Available As sessile organisms, plants are constantly exposed to a wide spectrum of stress conditions such as high temperature, which causes protein misfolding. Misfolded proteins are highly toxic and must be efficiently removed to reduce cellular proteotoxic stress if restoration of native conformations is unsuccessful. Although selective autophagy is known to function in protein quality control by targeting degradation of misfolded and potentially toxic proteins, its role and regulation in heat stress responses have not been analyzed in crop plants. In the present study, we found that heat stress induced expression of autophagy-related (ATG genes and accumulation of autophagosomes in tomato plants. Virus-induced gene silencing of tomato ATG5 and ATG7 genes resulted in increased sensitivity of tomato plants to heat stress based on both increased development of heat stress symptoms and compromised photosynthetic parameters of heat-stressed leaf tissues. Silencing of tomato homologs for the selective autophagy receptor NBR1, which targets ubiquitinated protein aggregates, also compromised tomato heat tolerance. To better understand the regulation of heat-induced autophagy, we found that silencing of tomato ATG5, ATG7 or NBR1 compromised heat-induced expression of not only the targeted genes but also other autophagy-related genes. Furthermore, we identified two tomato genes encoding proteins highly homologous to Arabidopsis WRKY33 transcription factor, which has been previously shown to interact physically with an autophagy protein. Silencing of tomato WRKY33 genes compromised tomato heat tolerance and reduced heat-induced ATG gene expression and autophagosome accumulation. Based on these results, we propose that heat-induced autophagy in tomato is subject to cooperative regulation by both WRKY33 and ATG proteins and plays a critical role in tomato heat tolerance, mostly likely through selective removal of heat-induced protein aggregates.

  8. Role and regulation of autophagy in heat stress responses of tomato plants.

    Science.gov (United States)

    Zhou, Jie; Wang, Jian; Yu, Jing-Quan; Chen, Zhixiang

    2014-01-01

    As sessile organisms, plants are constantly exposed to a wide spectrum of stress conditions such as high temperature, which causes protein misfolding. Misfolded proteins are highly toxic and must be efficiently removed to reduce cellular proteotoxic stress if restoration of native conformations is unsuccessful. Although selective autophagy is known to function in protein quality control by targeting degradation of misfolded and potentially toxic proteins, its role and regulation in heat stress responses have not been analyzed in crop plants. In the present study, we found that heat stress induced expression of autophagy-related (ATG) genes and accumulation of autophagosomes in tomato plants. Virus-induced gene silencing (VIGS) of tomato ATG5 and ATG7 genes resulted in increased sensitivity of tomato plants to heat stress based on both increased development of heat stress symptoms and compromised photosynthetic parameters of heat-stressed leaf tissues. Silencing of tomato homologs for the selective autophagy receptor NBR1, which targets ubiquitinated protein aggregates, also compromised tomato heat tolerance. To better understand the regulation of heat-induced autophagy, we found that silencing of tomato ATG5, ATG7, or NBR1 compromised heat-induced expression of not only the targeted genes but also other autophagy-related genes. Furthermore, we identified two tomato genes encoding proteins highly homologous to Arabidopsis WRKY33 transcription factor, which has been previously shown to interact physically with an autophagy protein. Silencing of tomato WRKY33 genes compromised tomato heat tolerance and reduced heat-induced ATG gene expression and autophagosome accumulation. Based on these results, we propose that heat-induced autophagy in tomato is subject to cooperative regulation by both WRKY33 and ATG proteins and plays a critical role in tomato heat tolerance, mostly likely through selective removal of heat-induced protein aggregates.

  9. Neuronal autophagy in cerebral ischemia

    Institute of Scientific and Technical Information of China (English)

    Feng Xu; Jin-Hua Gu; Zheng-Hong Qin

    2012-01-01

    Autophagy has evolved as a conserved process for the bulk degradation and recycling of cytosolic components,such as long-lived proteins and organelles.In neurons,autophagy is important for homeostasis and protein quality control and is maintained at relatively low levels under normal conditions,while it is upregulated in response to pathophysiological conditions,such as cerebral ischemic injury.However,the role of autophagy is more complex.It depends on age or brain maturity,region,severity of insult,and the stage of ischemia.Whether autophagy plays a beneficial or a detrimental role in cerebral ischemia depends on various pathological conditions.In this review,we elucidate the role of neuronal autophagy in cerebral ischemia.

  10. Non-degradative Ubiquitination of Protein Kinases.

    Directory of Open Access Journals (Sweden)

    K Aurelia Ball

    2016-06-01

    Full Text Available Growing evidence supports other regulatory roles for protein ubiquitination in addition to serving as a tag for proteasomal degradation. In contrast to other common post-translational modifications, such as phosphorylation, little is known about how non-degradative ubiquitination modulates protein structure, dynamics, and function. Due to the wealth of knowledge concerning protein kinase structure and regulation, we examined kinase ubiquitination using ubiquitin remnant immunoaffinity enrichment and quantitative mass spectrometry to identify ubiquitinated kinases and the sites of ubiquitination in Jurkat and HEK293 cells. We find that, unlike phosphorylation, ubiquitination most commonly occurs in structured domains, and on the kinase domain, ubiquitination is concentrated in regions known to be important for regulating activity. We hypothesized that ubiquitination, like other post-translational modifications, may alter the conformational equilibrium of the modified protein. We chose one human kinase, ZAP-70, to simulate using molecular dynamics with and without a monoubiquitin modification. In Jurkat cells, ZAP-70 is ubiquitinated at several sites that are not sensitive to proteasome inhibition and thus may have other regulatory roles. Our simulations show that ubiquitination influences the conformational ensemble of ZAP-70 in a site-dependent manner. When monoubiquitinated at K377, near the C-helix, the active conformation of the ZAP-70 C-helix is disrupted. In contrast, when monoubiquitinated at K476, near the kinase hinge region, an active-like ZAP-70 C-helix conformation is stabilized. These results lead to testable hypotheses that ubiquitination directly modulates kinase activity, and that ubiquitination is likely to alter structure, dynamics, and function in other protein classes as well.

  11. Non-degradative Ubiquitination of Protein Kinases.

    Science.gov (United States)

    Ball, K Aurelia; Johnson, Jeffrey R; Lewinski, Mary K; Guatelli, John; Verschueren, Erik; Krogan, Nevan J; Jacobson, Matthew P

    2016-06-01

    Growing evidence supports other regulatory roles for protein ubiquitination in addition to serving as a tag for proteasomal degradation. In contrast to other common post-translational modifications, such as phosphorylation, little is known about how non-degradative ubiquitination modulates protein structure, dynamics, and function. Due to the wealth of knowledge concerning protein kinase structure and regulation, we examined kinase ubiquitination using ubiquitin remnant immunoaffinity enrichment and quantitative mass spectrometry to identify ubiquitinated kinases and the sites of ubiquitination in Jurkat and HEK293 cells. We find that, unlike phosphorylation, ubiquitination most commonly occurs in structured domains, and on the kinase domain, ubiquitination is concentrated in regions known to be important for regulating activity. We hypothesized that ubiquitination, like other post-translational modifications, may alter the conformational equilibrium of the modified protein. We chose one human kinase, ZAP-70, to simulate using molecular dynamics with and without a monoubiquitin modification. In Jurkat cells, ZAP-70 is ubiquitinated at several sites that are not sensitive to proteasome inhibition and thus may have other regulatory roles. Our simulations show that ubiquitination influences the conformational ensemble of ZAP-70 in a site-dependent manner. When monoubiquitinated at K377, near the C-helix, the active conformation of the ZAP-70 C-helix is disrupted. In contrast, when monoubiquitinated at K476, near the kinase hinge region, an active-like ZAP-70 C-helix conformation is stabilized. These results lead to testable hypotheses that ubiquitination directly modulates kinase activity, and that ubiquitination is likely to alter structure, dynamics, and function in other protein classes as well.

  12. Cocaine induces astrocytosis through ER stress-mediated activation of autophagy.

    Science.gov (United States)

    Periyasamy, Palsamy; Guo, Ming-Lei; Buch, Shilpa

    2016-08-01

    Cocaine is known to induce inflammation, thereby contributing in part, to the pathogenesis of neurodegeneration. A recent study from our lab has revealed a link between macroautophagy/autophagy and microglial activation. The current study was aimed at investigating whether cocaine could also mediate activation of astrocytes and, whether this process involved induction of autophagy. Our findings demonstrated that cocaine mediated the activation of astrocytes by altering the levels of autophagy markers, such as BECN1, ATG5, MAP1LC3B-II, and SQSTM1 in both human A172 astrocytoma cells and primary human astrocytes. Furthermore, cocaine treatment resulted in increased formation of endogenous MAP1LC3B puncta in human astrocytes. Additionally, astrocytes transfected with the GFP-MAP1LC3B plasmid also demonstrated cocaine-mediated upregulation of the green fluorescent MAP1LC3B puncta. Cocaine-mediated induction of autophagy involved upstream activation of ER stress proteins such as EIF2AK3, ERN1, ATF6 since blockage of autophagy using either pharmacological or gene-silencing approaches, had no effect on cocaine-mediated induction of ER stress. Using both pharmacological and gene-silencing approaches to block either ER stress or autophagy, our findings demonstrated that cocaine-induced activation of astrocytes (measured by increased levels of GFAP) involved sequential activation of ER stress and autophagy. Cocaine-mediated-increased upregulation of GFAP correlated with increased expression of proinflammatory mediators such as TNF, IL1B, and IL6. In conclusion, these findings reveal an association between ER stress-mediated autophagy and astrogliosis in cocaine-treated astrocytes. Intervention of ER stress and/or autophagy signaling would thus be promising therapeutic targets for abrogating cocaine-mediated neuroinflammation. PMID:27337297

  13. Enhancement of CIITA transcriptional function by ubiquitin.

    Science.gov (United States)

    Greer, Susanna F; Zika, Eleni; Conti, Brian; Zhu, Xin-Sheng; Ting, Jenny P-Y

    2003-11-01

    Although increasing evidence indicates that there is a direct link between ubiquitination and mono-ubiquitination and transcription in yeast, this link has not been demonstrated in higher eukaryotes. Here we show that the major histocompatibility complex (MHC) class II transactivator (CIITA), which is required for expression of genes encoding MHC class II molecules, is ubiquitinated. This ubiquitination enhanced the association of CIITA with both MHC class II transcription factors and the MHC class II promoter, resulting in an increase in transactivation function and in the expression of MHC class II mRNA. The degree of CIITA ubiquitination was controlled by histone acetylases (HATs) and deacetylases (HDACs), indicating that the crucial cellular processes mediated by these enzymes are linked to regulate transcription. Thus, ubiquitin positively regulates a mammalian coactivator by enhancing its assembly at the promoter.

  14. Autophagy regulates chlorpyrifos-induced apoptosis in SH-SY5Y cells

    Energy Technology Data Exchange (ETDEWEB)

    Park, Jae Hyeon [Department of Pharmacology, College of Medicine, Hanyang University (Korea, Republic of); Hanyang Biomedical Research Institute, Seoul (Korea, Republic of); Graduate School of Biomedical Science and Engineering, Hanyang University, Seoul (Korea, Republic of); Lee, Jeong Eun [Department of Pharmacology, College of Medicine, Hanyang University (Korea, Republic of); Hanyang Biomedical Research Institute, Seoul (Korea, Republic of); Shin, In Chul [Department of Pharmacology, College of Medicine, Hanyang University (Korea, Republic of); Koh, Hyun Chul, E-mail: hckoh@hanyang.ac.kr [Department of Pharmacology, College of Medicine, Hanyang University (Korea, Republic of); Hanyang Biomedical Research Institute, Seoul (Korea, Republic of); Graduate School of Biomedical Science and Engineering, Hanyang University, Seoul (Korea, Republic of)

    2013-04-01

    Recent studies have shown that up-regulation of autophagy may be a tractable therapeutic intervention for clearing disease-causing proteins, including α-synuclein, ubiquitin, and other misfolded or aggregated proteins in pesticide-induced neurodegeneration. In a previous study, we reported that chlorpyrifos (CPF)-induced mitochondria-dependent apoptosis is mediated through reactive oxygen species in SH-SY5Y cells. In this study, we explored a novel pharmacotherapeutic approach to prevent CPF neurotoxicity involving the regulation of autophagy. We investigated the modulation of CPF-induced apoptosis according to autophagy regulation. We found that CPF induced apoptosis in SH-SY5Y cells, as demonstrated by the activation of caspase-3 and nuclear condensation. In addition, we observed that cells treated with CPF underwent autophagic cell death by monitoring the expression of LC3-II and p62. Pretreatment with the autophagy inducer rapamycin significantly enhanced the cell viability of CPF-exposed cells, and the enhancement of cell viability was partially due to alleviation of CPF-induced apoptosis via a decrease in levels of cleaved caspase-3. Specifically, rapamycin pretreatment decreased Bax and increased Bcl-2 expression in mitochondria. In addition, rapamycin significantly decreased cytochrome c release in from mitochondria into the cytosol. However, pretreatment of cells with the autophagy inhibitor, 3-methyladenine (3MA), remarkably increased CPF toxicity in these cells; this with correlated with increased expression of Bax and decreased expression of Bcl-2 in mitochondria. Our results suggest that CPF-induced cytotoxicity is modified by autophagy regulation and that rapamycin protects against CPF-induced apoptosis by enhancing autophagy. Pharmacologic induction of autophagy by rapamycin may be a useful treatment strategy in neurodegenerative disorders. - Highlights: ► Chlorpyrifos (CPF) is cytotoxic to SH-SY5Y cells ► CPF-induced cytotoxicity is mediated by

  15. Autophagy and neurodegenerative disorders

    Institute of Scientific and Technical Information of China (English)

    Evangelia Kesidou; Roza Lagoudaki; Olga Touloumi; Kyriaki-Nefeli Poulatsidou; Constantina Simeonidou

    2013-01-01

    Accumulation of aberrant proteins and inclusion bodies are hallmarks in most neurodegenerative diseases. Consequently, these aggregates within neurons lead to toxic effects, overproduction of reactive oxygen species and oxidative stress. Autophagy is a significant intracel ular mechanism that removes damaged organelles and misfolded proteins in order to maintain cel homeostasis. Excessive or insufficient autophagic activity in neurons leads to altered homeostasis and influences their survival rate, causing neurodegeneration. The review article provides an update of the role of autophagic process in representative chronic and acute neurodegenerative disorders.

  16. Ubiquitin-mediated response to microsporidia and virus infection in C. elegans.

    Directory of Open Access Journals (Sweden)

    Malina A Bakowski

    2014-06-01

    Full Text Available Microsporidia comprise a phylum of over 1400 species of obligate intracellular pathogens that can infect almost all animals, but little is known about the host response to these parasites. Here we use the whole-animal host C. elegans to show an in vivo role for ubiquitin-mediated response to the microsporidian species Nematocida parisii, as well to the Orsay virus, another natural intracellular pathogen of C. elegans. We analyze gene expression of C. elegans in response to N. parisii, and find that it is similar to response to viral infection. Notably, we find an upregulation of SCF ubiquitin ligase components, such as the cullin ortholog cul-6, which we show is important for ubiquitin targeting of N. parisii cells in the intestine. We show that ubiquitylation components, the proteasome, and the autophagy pathway are all important for defense against N. parisii infection. We also find that SCF ligase components like cul-6 promote defense against viral infection, where they have a more robust role than against N. parisii infection. This difference may be due to suppression of the host ubiquitylation system by N. parisii: when N. parisii is crippled by anti-microsporidia drugs, the host can more effectively target pathogen cells for ubiquitylation. Intriguingly, inhibition of the ubiquitin-proteasome system (UPS increases expression of infection-upregulated SCF ligase components, indicating that a trigger for transcriptional response to intracellular infection by N. parisii and virus may be perturbation of the UPS. Altogether, our results demonstrate an in vivo role for ubiquitin-mediated defense against microsporidian and viral infections in C. elegans.

  17. IL-33对大鼠 I/R 损伤心肌炎症反应和细胞自噬的影响%Effect of interleukin 33 on inflammation response and autophagy in rats with myocardial ischemia-reperfusion injury

    Institute of Scientific and Technical Information of China (English)

    马瑞松; 李元红; 江洪; 胡笑容; 李雪飞

    2015-01-01

    Objective To investigate the protective effect of interleukin 33 (IL-33) on myocardial ischemia-reperfu-sion ( I/R) injury and the mechanism.Methods Thirty-two rats were randomly divided into 4 groups:the control group (n=10), I/R group (model group, n=10), IL-33 group (n=6) and anti-ST2 group (n=6).In addition to the control group, the left anterior descending coronary artery ligation method was adopted to establish the myocardial I/R injury model in the other groups ( the sham operation group only received anesthesia, open-chest and threading, but not ligation) .Rats in the IL-33+I/R group and anti-ST2+I/R group were separately injected to the caudal vein with 10μg IL-33 and 0.2 mL anti-ST2 (1 mg/mL) 30 min before modeling.After reperfusion for 4 h, we obtained the serum or myocardial tissues to de-tect the following indicators of each group:(1) the serum lactate dehydrogenase (LDH) and creatine kinase (CK) level:using spectrophotometry, (2) Th1 inflammation factors in the myocardial tissues (TNF-α, INF-γand IL-6) and Th2 in-flammatory cytokines (IL-4, IL-5 and IL-3):using the ELISA, (3) the relative expression of autophagy protein LC3 and beclin 1 in the myocardial tissues:using Western blotting.Results (1) LDH and CK level:the model group was signifi-cantly higher than the control group, IL-33 group was significantly lower than the model group (P<0.05), and no statisti-cal difference was found between the anti-ST2 group and the model group.(2) the inflammation factor expression in the myocardial tissues:Th1 type inflammation factor expression: the model group and IL-33 group were significantly higher than the control group, IL-33 was significantly lower than the model group (all P<0.05), and no difference was found be-tween the anti-ST2 group and the model group.Th2 type inflammation factor expression:the model group was significantly lower than the control group, IL-33 group was significantly higher than the model group, and no statistical difference was

  18. Autophagy Mediates HBx-Induced Nuclear Factor-κB Activation and Release of IL-6, IL-8, and CXCL2 in Hepatocytes.

    Science.gov (United States)

    Luo, Millore X M; Wong, Sunny H; Chan, Matthew T V; Yu, Le; Yu, Sidney S B; Wu, Feng; Xiao, Zhangang; Wang, Xiaojuan; Zhang, Lin; Cheng, Alfred S L; Ng, Simon S M; Chan, Francis K L; Cho, Chi H; Yu, Jun; Sung, Joseph J Y; Wu, William K K

    2015-10-01

    Hepatitis B virus (HBV) and one of its encoded proteins, HBV X protein (HBx), have been shown to induce autophagy in hepatoma cells. Substantial evidence indicates that autophagy is a potent suppressor of inflammation. However, sporadic reports suggest that autophagy could promote pro-inflammatory cytokine expression and inflammation in some biological contexts. Here, we show that overexpression of HBx induces LC3B-positive autophagosome formation, increases autophagic flux and enhances the expression of ATG5, ATG7, and LC3B-II in normal hepatocytes. Abrogation of autophagy by small interfering RNA against ATG5 and ATG7 prevents HBx-induced formation of autophagosomes. Autophagy inhibition also abrogates HBx-induced activation of nuclear factor-κB (NF-κB) and production of interleukin-6 (IL-6), IL-8, and CXCL2. These findings suggest that autophagy is required for HBx-induced NF-κB activation and pro-inflammatory cytokine production and could shed new light on the complex role of autophagy in the modulation of inflammation.

  19. Inhibition of caspase-9 aggravates acute liver injury through suppression of cytoprotective autophagy

    Science.gov (United States)

    Guo, Rui; Lin, Bin; Pan, Jing Fei; Liong, Emily C.; Xu, Ai Min; Youdim, Moussa; Fung, Man Lung; So, Kwok Fai; Tipoe, George L.

    2016-01-01

    Acute liver disease is characterized by inflammation, oxidative stress and necrosis, which can greatly influence the long term clinical outcome and lead to liver failure or cancer. Here, we initially demonstrated the beneficial role of caspase-9-dependent autophagy in acute liver injury. Treatment with caspase-9 inhibitor z-LEHD-FMK in HepG2 cells, AML12 cells and C57BL/b6N mice exacerbated CCl4-induced acute hepatocellular damage, and also down-regulated autophagy markers expression levels, indicating that caspase-9 inhibition may aggravate acute liver damage by suppressing cytoprotective autophagy. CCl4 was used as an acute liver injury inducer which caused oxidative stress and apoptosis through up-regulation of HIF-1α, as well as triggered hepatic inflammation and necroptosis via TLR4/NF-κB pathway. Caspase-9 Thr125 site was firstly phosphorylated by ERK1/2 which subsequently activated the cytoprotective autophagy process to attenuate acute CCl4 injury. Caspase-9 inhibition further aggravated hepatic necroptosis through NF-κB expression, leading to increased pro-inflammatory mediators levels, suggesting a protective role of caspase-9-dependent autophagy in the inflammatory process as well as its possibility being a new therapeutic target for the treatment of acute liver injury. PMID:27580936

  20. Ubiquitin and Ubiquitin-Like Modifications of the p53 Family

    Directory of Open Access Journals (Sweden)

    Ian R. Watson

    2006-08-01

    Full Text Available Regulation of p53 by the ubiquitin-proteasomal pathway has been studied considerably. Studies have also demonstrated that the ubiquitin-like proteins SUMO-1 and NEDD8 modify p53. Similarly, p63 and p73 are subject to regulation by ubiquitin and ubiquitin-like modifications, and perturbations of these pathways in the regulation of the p53 family have been implicated in tumorigenesis and developmental abnormalities. Here, we provide an overview of the current understanding of the regulation of the p53 family by covalent modification by ubiquitin, SUMO-1, and NEDD8.

  1. Notch1 regulated autophagy controls survival and suppressor activity of activated murine T-regulatory cells

    Science.gov (United States)

    Marcel, Nimi; Sarin, Apurva

    2016-01-01

    Cell survival is one of several processes regulated by the Notch pathway in mammalian cells. Here we report functional outcomes of non-nuclear Notch signaling to activate autophagy, a conserved cellular response to nutrient stress, regulating survival in murine natural T-regulatory cells (Tregs), an immune subset controlling tolerance and inflammation. Induction of autophagy required ligand-dependent, Notch intracellular domain (NIC) activity, which controlled mitochondrial organization and survival of activated Tregs. Consistently, NIC immune-precipitated Beclin and Atg14, constituents of the autophagy initiation complex. Further, ectopic expression of an effector of autophagy (Atg3) or recombinant NIC tagged to a nuclear export signal (NIC-NES), restored autophagy and suppressor function in Notch1-/- Tregs. Furthermore, Notch1 deficiency in the Treg lineage resulted in immune hyperactivity, implicating Notch activity in Treg homeostasis. Notch1 integration with autophagy, revealed in these experiments, holds implications for Notch regulated cell-fate decisions governing differentiation. DOI: http://dx.doi.org/10.7554/eLife.14023.001 PMID:27267497

  2. Mitochondrial damage contributes to Pseudomonas aeruginosa activation of the inflammasome and is downregulated by autophagy.

    Science.gov (United States)

    Jabir, Majid Sakhi; Hopkins, Lee; Ritchie, Neil D; Ullah, Ihsan; Bayes, Hannah K; Li, Dong; Tourlomousis, Panagiotis; Lupton, Alison; Puleston, Daniel; Simon, Anna Katharina; Bryant, Clare; Evans, Thomas J

    2015-01-01

    The nucleotide-binding domain, leucine-rich repeat containing family caspase recruitment domain containing 4 (NLRC4) inflammasome can be activated by pathogenic bacteria via products translocated through the microbial type III secretion apparatus (T3SS). Recent work has shown that activation of the NLRP3 inflammasome is downregulated by autophagy, but the influence of autophagy on NLRC4 activation is unclear. We set out to determine how autophagy might influence this process, using the bacterium Pseudomonas aeruginosa, which activates the NLRC4 inflammasome via its T3SS. Infection resulted in T3SS-dependent mitochondrial damage with increased production of reactive oxygen intermediates and release of mitochondrial DNA. Inhibiting mitochondrial reactive oxygen release or degrading intracellular mitochondrial DNA abrogated NLRC4 inflammasome activation. Moreover, macrophages lacking mitochondria failed to activate NLRC4 following infection. Removal of damaged mitochondria by autophagy significantly attenuated NLRC4 inflammasome activation. Mitochondrial DNA bound specifically to NLRC4 immunoprecipitates and transfection of mitochondrial DNA directly activated the NLRC4 inflammasome; oxidation of the DNA enhanced this effect. Manipulation of autophagy altered the degree of inflammasome activation and inflammation in an in vivo model of P. aeruginosa infection. Our results reveal a novel mechanism contributing to NLRC4 activation by P. aeruginosa via mitochondrial damage and release of mitochondrial DNA triggered by the bacterial T3SS that is downregulated by autophagy.

  3. Ordered bulk degradation via autophagy

    DEFF Research Database (Denmark)

    Dengjel, Jörn; Kristensen, Anders Riis; Andersen, Jens S

    2008-01-01

    During amino acid starvation, cells undergo macroautophagy which is regarded as an unspecific bulk degradation process. Lately, more and more organelle-specific autophagy subtypes such as reticulophagy, mitophagy and ribophagy have been described and it could be shown, depending on the experimental...... setup, that autophagy specifically can remove certain subcellular components. We used an unbiased quantitative proteomics approach relying on stable isotope labeling by amino acids in cell culture (SILAC) to study global protein dynamics during amino acid starvation-induced autophagy. Looking...... at proteasomal and lysosomal degradation ample cross-talk between the two degradation pathways became evident. Degradation via autophagy appeared to be ordered and regulated at the protein complex/organelle level. This raises several important questions such as: can macroautophagy itself be specific and what...

  4. Pink1, the first ubiquitin kinase

    OpenAIRE

    Zheng, Xinde; Hunter, Tony

    2014-01-01

    Pink1 and Parkin, identified through studies of hereditary early onset Parkinson's disease, are involved in mitochondria quality control. Parkin E3 ubiquitin ligase activity is activated by Pink1 kinase activity, although the mechanism is still elusive. Three recent reports uncover a surprising mechanism in which Pink1 directly phosphorylates ubiquitin to boost Parkin activity.

  5. Met1-linked Ubiquitination in Immune Signalling

    DEFF Research Database (Denmark)

    Fiil, Berthe Katrine; Gyrd-Hansen, Mads

    2014-01-01

    Methionine 1-linked ubiquitin chains (Met1-Ub), or linear ubiquitin, has emerged as a central post-translational modification in innate immune signalling. Molecular machinery that assembles, senses and, more recently, disassembles Met1-Ub has been identified, and technical advances have enabled...

  6. Autophagy in DNA Damage Response

    Directory of Open Access Journals (Sweden)

    Piotr Czarny

    2015-01-01

    Full Text Available DNA damage response (DDR involves DNA repair, cell cycle regulation and apoptosis, but autophagy is also suggested to play a role in DDR. Autophagy can be activated in response to DNA-damaging agents, but the exact mechanism underlying this activation is not fully understood, although it is suggested that it involves the inhibition of mammalian target of rapamycin complex 1 (mTORC1. mTORC1 represses autophagy via phosphorylation of the ULK1/2–Atg13–FIP200 complex thus preventing maturation of pre-autophagosomal structures. When DNA damage occurs, it is recognized by some proteins or their complexes, such as poly(ADPribose polymerase 1 (PARP-1, Mre11–Rad50–Nbs1 (MRN complex or FOXO3, which activate repressors of mTORC1. SQSTM1/p62 is one of the proteins whose levels are regulated via autophagic degradation. Inhibition of autophagy by knockout of FIP200 results in upregulation of SQSTM1/p62, enhanced DNA damage and less efficient damage repair. Mitophagy, one form of autophagy involved in the selective degradation of mitochondria, may also play role in DDR. It degrades abnormal mitochondria and can either repress or activate apoptosis, but the exact mechanism remains unknown. There is a need to clarify the role of autophagy in DDR, as this process may possess several important biomedical applications, involving also cancer therapy.

  7. Inhibiting autophagy promotes endoplasmic reticulum stress and the ROS‑induced nod‑like receptor 3‑dependent proinflammatory response in HepG2 cells.

    Science.gov (United States)

    Yin, Jia-Jing; Xie, Guangying; Zhang, Ning; Li, Yanbo

    2016-10-01

    Inflammation and endoplasmic reticulum (ER) stress are key contributors to insulin resistance and metabolic disease, and interleukin (IL)‑1β is involved in insulin resistance. The present study aimed to investigated the role of autophagy in LPS‑induced ER stress and inflammation, which may provide evidence for controlling metabolic disease associated with inflammation. Lipopolysaccharide (LPS) induced the activation of ER stress and the nod‑like receptor 3‑dependent expression of IL‑1β and caspase‑1, as shown by western blotting, which contributed to HepG2 cell death. This also involved the generation of mitochondrial reactive oxygen species and the autophagy signaling response, which are derived from the ER stress pathway. The percentage of apoptotic cells was measured by flow cytometry with fluorescein isothiocyanate/propidium iodide staining. Reactive oxygen species formation was detected by flow cytometry using the peroxide sensitive fluorescent probe 2',7'‑dichlorofluorescin diacetate. Autophagy activation was measured by western blotting and confirmed using transmission electron microscopy. Furthermore, inhibiting autophagy promoted ER stress and the proinflammatory response in addition to cell death. These findings provide insights into the protective role of autophagy in LPS‑induced cell death and ER stress, and further identified the association of autophagy, ER stress and inflammation in HepG2 cells.

  8. Advanced Glycation End Products (AGE) Potently Induce Autophagy through Activation of RAF Protein Kinase and Nuclear Factor κB (NF-κB).

    Science.gov (United States)

    Verma, Neeharika; Manna, Sunil K

    2016-01-15

    Advanced glycation end products (AGE) accumulate in diabetic patients and aging people because of high amounts of three- or four-carbon sugars derived from glucose, thereby causing multiple consequences, including inflammation, apoptosis, obesity, and age-related disorders. It is important to understand the mechanism of AGE-mediated signaling leading to the activation of autophagy (self-eating) that might result in obesity. We detected AGE as one of the potent inducers of autophagy compared with doxorubicin and TNF. AGE-mediated autophagy is inhibited by suppression of PI3K and potentiated by the autophagosome maturation blocker bafilomycin. It increases autophagy in different cell types, and that correlates with the expression of its receptor, receptor for AGE. LC3B, the marker for autophagosomes, is shown to increase upon AGE stimulation. AGE-mediated autophagy is partially suppressed by inhibitor of NF-κB, PKC, or ERK alone and significantly in combination. AGE increases sterol regulatory element binding protein activity, which leads to an increase in lipogenesis. Although AGE-mediated lipogenesis is affected by autophagy inhibitors, AGE-mediated autophagy is not influenced by lipogenesis inhibitors, suggesting that the turnover of lipid droplets overcomes the autophagic clearance. For the first time, we provide data showing that AGE induces several cell signaling cascades, like NF-κB, PKC, ERK, and MAPK, that are involved in autophagy and simultaneously help with the accumulation of lipid droplets that are not cleared effectively by autophagy, therefore causing obesity.

  9. Regulation of nucleotide excision repair through ubiquitination

    Institute of Scientific and Technical Information of China (English)

    Jia Li; Audesh Bhat; Wei Xiao

    2011-01-01

    Nucleotide excision repair (NER) is the most versatile DNA-repair pathway in all organisms.While bacteria require only three proteins to complete the incision step of NER,eukaryotes employ about 30 proteins to complete the same step.Here we summarize recent studies demonstrating that ubiquitination,a post-translational modification,plays critical roles in regulating the NER activity either dependent on or independent of ubiquitin-proteolysis.Several NER components have been shown as targets of ubiquitination while others are actively involved in the ubiquitination process.We argue through this analysis that ubiquitination serves to coordinate various steps of NER and meanwhile connect NER with other related pathways to achieve the efficient global DNA-damage response.

  10. Regulation of plant immune receptors by ubiquitination

    Directory of Open Access Journals (Sweden)

    Giulia eFurlan

    2012-10-01

    Full Text Available From pathogen perception and the activation of signal transduction cascades to the deployment of defense responses, protein ubiquitination plays a key role in the modulation of plant immunity. Ubiquitination is mediated by three enzymes, of which the E3 ubiquitin ligases, the substrate determinants, have been the major focus of attention. Accumulating evidence suggests that ubiquitination modulates signaling mediated by pattern recognition receptors (PRRs and is important for the accumulation of nucleotide-binding leucine-rich repeat (NB-LRR type intracellular immune sensors. Recent studies also indicate that ubiquitination directs vesicle trafficking, a function that has been clearly established for immune signaling in animals. In this mini review, we discuss these and other recent advances and highlight important open questions.

  11. Regulation of plant immune receptors by ubiquitination.

    Science.gov (United States)

    Furlan, Giulia; Klinkenberg, Jörn; Trujillo, Marco

    2012-01-01

    From pathogen perception and the activation of signal transduction cascades to the deployment of defense responses, protein ubiquitination plays a key role in the modulation of plant immunity. Ubiquitination is mediated by three enzymes, of which the E3 ubiquitin ligases, the substrate determinants, have been the major focus of attention. Accumulating evidence suggests that ubiquitination modulates signaling mediated by pattern recognition receptors and is important for the accumulation of nucleotide-binding leucine-rich repeat type intracellular immune sensors. Recent studies also indicate that ubiquitination directs vesicle trafficking, a function that has been clearly established for immune signaling in animals. In this mini review, we discuss these and other recent advances and highlight important open questions. PMID:23109936

  12. The autophagy gene Wdr45/Wipi4 regulates learning and memory function and axonal homeostasis.

    Science.gov (United States)

    Zhao, Yan G; Sun, Le; Miao, Guangyan; Ji, Cuicui; Zhao, Hongyu; Sun, Huayu; Miao, Lin; Yoshii, Saori R; Mizushima, Noboru; Wang, Xiaoqun; Zhang, Hong

    2015-01-01

    WDR45/WIPI4, encoding a WD40 repeat-containing PtdIns(3)P binding protein, is essential for the basal autophagy pathway. Mutations in WDR45 cause the neurodegenerative disease β-propeller protein-associated neurodegeneration (BPAN), a subtype of NBIA. We generated CNS-specific Wdr45 knockout mice, which exhibit poor motor coordination, greatly impaired learning and memory, and extensive axon swelling with numerous axon spheroids. Autophagic flux is defective and SQSTM1 (sequestosome-1)/p62 and ubiquitin-positive protein aggregates accumulate in neurons and swollen axons. Nes-Wdr45(fl/Y) mice recapitulate some hallmarks of BPAN, including cognitive impairment and defective axonal homeostasis, providing a model for revealing the disease pathogenesis of BPAN and also for investigating the possible role of autophagy in axon maintenance.

  13. Intracellular release of rapamycin from poly (lactic acid) nanospheres modifies autophagy.

    Science.gov (United States)

    Nagata, Junpei; Matsui, Makoto; Tabata, Yasuhiko

    2016-09-01

    The objective of this study is to investigate the autophagy activity of cells by the intracellular release of rapamycin (Rapa) of an autophagy inducer. Rapa was incorporated into nanospheres of poly (lactic-co-glycolic acid) (PLGA) for the controlled release of Rapa. Rapa was released from the PLGA nanospheres incorporating rapamycin (Rapa-PLGA-NS) with time while the Rapa-PLGA-NS were hydrolytically degraded. When human hepatocellular carcinoma (HepG2) cells were incubated with the Rapa-PLGA-NS, the Rapa-PLGA-NS were internalized, and the intracellular concentration was maintained over four days, indicating the intracellular Rapa release. The microtubule-associated protein 1 light chain (LC3) of an autophagy marker was significantly high for the Rapa-PLGA-NS group compared with the free Rapa group even after four days incubation. In addition, intracellular harmful ubiquitinated proteins were degraded by the intracellular release of Rapa even after four days incubation in contrast to free Rapa. It is concluded that the intracellular Rapa release is effective in modulating the autophagy activity over a longer time period. PMID:27320771

  14. Interactions between Autophagy and Inhibitory Cytokines

    Science.gov (United States)

    Wu, Tian-tian; Li, Wei-Min; Yao, Yong-Ming

    2016-01-01

    Autophagy is a degradative pathway that plays an essential role in maintaining cellular homeostasis. Most early studies of autophagy focused on its involvement in age-associated degeneration and nutrient deprivation. However, the immunological functions of autophagy have become more widely studied in recent years. Autophagy has been shown to be an intrinsic cellular defense mechanism in the innate and adaptive immune responses. Cytokines belong to a broad and loose category of proteins and are crucial for innate and adaptive immunity. Inhibitory cytokines have evolved to permit tolerance to self while also contributing to the eradication of invading pathogens. Interactions between inhibitory cytokines and autophagy have recently been reported, revealing a novel mechanism by which autophagy controls the immune response. In this review, we discuss interactions between autophagy and the regulatory cytokines IL-10, transforming growth factor-β, and IL-27. We also mention possible interactions between two newly discovered cytokines, IL-35 and IL-37, and autophagy. PMID:27313501

  15. Autophagy and apoptosis: rivals or mates?

    Directory of Open Access Journals (Sweden)

    Yan Cheng

    2013-03-01

    Full Text Available Autophagy, a cellular process of "self-eating" by which intracellular components are degraded within the lysosome, is an evolutionarily conserved response to various stresses. Autophagy is associated with numerous patho-physiological conditions, and dysregulation of autophagy contributes to the pathogenesis of a variety of human diseases including cancer. Depending on context, activation of autophagy may promote either cell survival or death, two major events that determine pathological process of many illnesses. Importantly, the activity of autophagy is often associated with apoptosis, another critical cellular process determining cellular fate. A better understanding of biology of autophagy and its implication in human health and disorder, as well as the relationship between autophagy and apoptosis, has the potential of facilitating the development of autophagy-based therapeutic interventions for human diseases such as cancer.

  16. Autophagy and apoptosis: rivals or mates?

    Institute of Scientific and Technical Information of China (English)

    Yan Cheng; Jin-Ming Yang

    2013-01-01

    Autophagy,a cellular process of "self-eating" by which intracellular components are degraded within the lysosome,is an evolutionarily conserved response to various stresses.Autophagy is associated with numerous patho-physiological conditions,and dysregulation of autophagy contributes to the pathogenesis of a variety of human diseases including cancer.Depending on context,activation of autophagy may promote either cell survival or death,two major events that determine pathological process of many illnesses.Importantly,the activity of autophagy is often associated with apoptosis,another critical cellular process determining cellular fate.A better understanding of biology of autophagy and its implication in human health and disorder,as well as the relationship between autophagy and apoptosis,has the potential of facilitating the development of autophagy-based therapeutic interventions for human diseases such as cancer.

  17. Fluorescence microscopy: A tool to study autophagy

    Science.gov (United States)

    Rai, Shashank; Manjithaya, Ravi

    2015-08-01

    Autophagy is a cellular recycling process through which a cell degrades old and damaged cellular components such as organelles and proteins and the degradation products are reused to provide energy and building blocks. Dysfunctional autophagy is reported in several pathological situations. Hence, autophagy plays an important role in both cellular homeostasis and diseased conditions. Autophagy can be studied through various techniques including fluorescence based microscopy. With the advancements of newer technologies in fluorescence microscopy, several novel processes of autophagy have been discovered which makes it an essential tool for autophagy research. Moreover, ability to tag fluorescent proteins with sub cellular targets has enabled us to evaluate autophagy processes in real time under fluorescent microscope. In this article, we demonstrate different aspects of autophagy in two different model organisms i.e. yeast and mammalian cells, with the help of fluorescence microscopy.

  18. Interactions between Autophagy and Inhibitory Cytokines.

    Science.gov (United States)

    Wu, Tian-Tian; Li, Wei-Min; Yao, Yong-Ming

    2016-01-01

    Autophagy is a degradative pathway that plays an essential role in maintaining cellular homeostasis. Most early studies of autophagy focused on its involvement in age-associated degeneration and nutrient deprivation. However, the immunological functions of autophagy have become more widely studied in recent years. Autophagy has been shown to be an intrinsic cellular defense mechanism in the innate and adaptive immune responses. Cytokines belong to a broad and loose category of proteins and are crucial for innate and adaptive immunity. Inhibitory cytokines have evolved to permit tolerance to self while also contributing to the eradication of invading pathogens. Interactions between inhibitory cytokines and autophagy have recently been reported, revealing a novel mechanism by which autophagy controls the immune response. In this review, we discuss interactions between autophagy and the regulatory cytokines IL-10, transforming growth factor-β, and IL-27. We also mention possible interactions between two newly discovered cytokines, IL-35 and IL-37, and autophagy.

  19. Adaptive changes in autophagy after UPS impairment in Parkinson's disease

    Institute of Scientific and Technical Information of China (English)

    Yu-fei SHEN; Yu TANG; Xiao-jie ZHANG; Kai-xing HUANG; Wei-dong LE

    2013-01-01

    Ubiquitin-proteasome system (UPS) and autophagosome-lysosome pathway (ALP) are the most important machineries responsible for protein degradation in Parkinson's disease (PD).The aim of this study is to investigate the adaptive alterations in autophagy upon proteasome inhibition in dopaminergic neurons in vitro and in vivo.Methods:Human dopaminergic neuroblastoma SH-SY5Y cells were treated with the proteasome inhibitor lactacystin (5 pmol/L) for 5,12,or 24 h.The expression of autophagy-related proteins in the cells was detected with immunoblotting.UPS-impaired mouse model of PD was established by microinjection of lactacystin (2 pg) into the left hemisphere of C57BL/6 mice that were sacrificed 2 or 4weeks later.The midbrain tissues were dissected to assess alterations in autophagy using immunofluorescence,immunoblotting and electron microscopy assays.Results:Both in SH-SY5Y cells and in the midbrain of UPS-impaired mouse model of PD,treatment with lactacystin significantly increased the expression levels of LC3-Ⅰ/Ⅱ and Beclin 1,and reduced the levels of p-mTOR,mTOR and p62/SQSTM1.Furthermore,lactacystin treatment in UPS-impaired mouse model of PD caused significant loss of TH-positive neurons in the substantia nigra,and dramatically increased the number of autophagosomes in the left TH-positive neurons.Conclusion:Inhibition of UPS by lactacystin in dopaminergic neurons activates another protein degradation system,the ALP,which includes both the mTOR signaling pathway and Beclin 1-associated pathway.

  20. Crystal Structure of Oxidative Stress Sensor Keap1 in Complex with Selective Autophagy Substrate p62

    Science.gov (United States)

    Kurokawa, Hirofumi

    Keap1, an adaptor protein of cullin-RING ubiquitin ligase complex, represses cytoprotective transcription factor Nrf2 in an oxidative stress-dependent manner. The accumulation of selective autophagy substrate p62 also activates Nrf2 target genes, but the detailed mechanism has not been elucidated. Crystal structure of Keap1-p62 complex revealed the structural basis for the Nrf2 activation in which Keap1 is inactivated by p62. The accumulation of p62 is observed in hepatocellular carcinoma. The activation of Nrf2 target genes, including detoxifying enzymes and efflux transporters, by p62 may protect the cancer cells from anti-cancer drugs.

  1. STAT3-Mediated Autophagy Dependence Identifies Subtypes of Breast Cancer where Autophagy Inhibition can be Efficacious

    OpenAIRE

    Maycotte, Paola; Gearheart, Christy M.; Barnard, Rebecca; Aryal, Suraj; Mulcahy Levy, Jean M.; Fosmire, Susan P.; Hansen, Ryan J.; Morgan, Michael J.; Christopher C Porter; Gustafson, Daniel L.; Thorburn, Andrew

    2014-01-01

    Autophagy is a protein and organelle degradation pathway that is involved in diverse diseases including cancer. Recent evidence suggests that autophagy is a cell survival mechanism in tumor cells and that its inhibition especially in combination with other therapy could be beneficial but it remains unclear if all cancer cells behave the same way when autophagy is inhibited. We inhibited autophagy in a panel of breast cancer cell lines and found that some of them are dependent on autophagy for...

  2. Autophagy and mitophagy in cellular damage control

    Directory of Open Access Journals (Sweden)

    Jianhua Zhang

    2013-01-01

    Full Text Available Autophagy and mitophagy are important cellular processes that are responsible for breaking down cellular contents, preserving energy and safeguarding against accumulation of damaged and aggregated biomolecules. This graphic review gives a broad summary of autophagy and discusses examples where autophagy is important in controlling protein degradation. In addition we highlight how autophagy and mitophagy are involved in the cellular responses to reactive species and mitochondrial dysfunction. The key signaling pathways for mitophagy are described in the context of bioenergetic dysfunction.

  3. Autophagy: for better or for worse

    OpenAIRE

    Wirawan, Ellen; Berghe, Tom Vanden; Lippens, Saskia; Agostinis, Patrizia; Vandenabeele, Peter

    2011-01-01

    Autophagy is a lysosomal degradation pathway that degrades damaged or superfluous cell components into basic biomolecules, which are then recycled back into the cytosol. In this respect, autophagy drives a flow of biomolecules in a continuous degradation-regeneration cycle. Autophagy is generally considered a pro-survival mechanism protecting cells under stress or poor nutrient conditions. Current research clearly shows that autophagy fulfills numerous functions in vital biological processes....

  4. p53: The Janus of autophagy?

    OpenAIRE

    Levine, Beth; Abrams, John

    2008-01-01

    The autophagy pathway functions in adaptation to nutrient stress and tumour suppression. The p53 tumour suppressor, previously thought to positively regulate autophagy, may also inhibit it. This dual interplay between p53 and autophagy regulation is enigmatic, but may underlie key aspects of metabolism and cancer biology.

  5. Autophagy, Innate Immunity and Tissue Repair in Acute Kidney Injury

    Directory of Open Access Journals (Sweden)

    Pu Duann

    2016-05-01

    Full Text Available Kidney is a vital organ with high energy demands to actively maintain plasma hemodynamics, electrolytes and water homeostasis. Among the nephron segments, the renal tubular epithelium is endowed with high mitochondria density for their function in active transport. Acute kidney injury (AKI is an important clinical syndrome and a global public health issue with high mortality rate and socioeconomic burden due to lack of effective therapy. AKI results in acute cell death and necrosis of renal tubule epithelial cells accompanied with leakage of tubular fluid and inflammation. The inflammatory immune response triggered by the tubular cell death, mitochondrial damage, associative oxidative stress, and the release of many tissue damage factors have been identified as key elements driving the pathophysiology of AKI. Autophagy, the cellular mechanism that removes damaged organelles via lysosome-mediated degradation, had been proposed to be renoprotective. An in-depth understanding of the intricate interplay between autophagy and innate immune response, and their roles in AKI pathology could lead to novel therapies in AKI. This review addresses the current pathophysiology of AKI in aspects of mitochondrial dysfunction, innate immunity, and molecular mechanisms of autophagy. Recent advances in renal tissue regeneration and potential therapeutic interventions are also discussed.

  6. Precision autophagy: Will the next wave of selective autophagy markers and specific autophagy inhibitors feed clinical pipelines?

    Science.gov (United States)

    Lebovitz, Chandra B; DeVorkin, Lindsay; Bosc, Damien; Rothe, Katharina; Singh, Jagbir; Bally, Marcel; Jiang, Xiaoyan; Young, Robert N; Lum, Julian J; Gorski, Sharon M

    2015-01-01

    Research presented at the Vancouver Autophagy Symposium (VAS) 2014 suggests that autophagy's influence on health and disease depends on tight regulation and precision targeting of substrates. Discussions recognized a pressing need for robust biomarkers that accurately assess the clinical utility of modulating autophagy in disease contexts. Biomarker discovery could flow from investigations of context-dependent triggers, sensors, and adaptors that tailor the autophagy machinery to achieve target specificity. In his keynote address, Dr. Vojo Deretic (University of New Mexico) described the discovery of a cargo receptor family that utilizes peptide motif-based cargo recognition, a mechanism that may be more precise than generic substrate tagging. The keynote by Dr. Alec Kimmelman (Harvard Medical School) emphasized that unbiased screens for novel selective autophagy factors may accelerate the development of autophagy-based therapies. Using a quantitative proteomics screen for de novo identification of autophagosome substrates in pancreatic cancer, Kimmelman's group discovered a new type of selective autophagy that regulates bioavailable iron. Additional presentations revealed novel autophagy regulators and receptors in metabolic diseases, proteinopathies, and cancer, and outlined the development of specific autophagy inhibitors and treatment regimens that combine autophagy modulation with anticancer therapies. VAS 2014 stimulated interdisciplinary discussions focused on the development of biomarkers, drugs, and preclinical models to facilitate clinical translation of key autophagy discoveries.

  7. Anti-tumor immunity, autophagy and chemotherapy

    Institute of Scientific and Technical Information of China (English)

    Gy(o)rgyi Müzes; Ferenc Sipos

    2012-01-01

    Autophagy or self-digestion of cells is activated upon various stressful stimuli and has been found to be a survival and drug resistance pathway in cancer.However,genetic studies support that autophagy can act as a tumor suppressor.Furthermore,defective autophagy is implicated in tumorigenesis,as well.The precise impact of autophagy on malignant transformation has not yet been clarified,but recent data suggest that this complex process is mainly directed by cell types,phases,genetic background and microenvironment.Relation of autophagy to anticancer immune responses may indicate a novel aspect in cancer chemotherapy.

  8. The cellular decision between apoptosis and autophagy

    Institute of Scientific and Technical Information of China (English)

    Yong-Jun Fan; Wei-Xing Zong

    2013-01-01

    Apoptosis and autophagy are important molecular processes that maintain organismal and cellular homeostasis,respectively.While apoptosis fulfills its role through dismantling damaged or unwanted cells,autophagy maintains cellular homeostasis through recycling selective intracellular organelles and molecules.Yet in some conditions,autophagy can lead to cell death.Apoptosis and autophagy can be stimulated by the same stresses.Emerging evidence indicates an interplay between the core proteins in both pathways,which underlies the molecular mechanism of the crosstalk between apoptosis and autophagy.This review summarizes recent literature on molecules that regulate both the apoptotic and autophagic processes.

  9. AUTOPHAGY AND IL-1 FAMILY CYTOKINES

    Directory of Open Access Journals (Sweden)

    James Harris

    2013-01-01

    Full Text Available Autophagy is an important intracellular homeostatic mechanism for the targeting of cytosolic constituents, including organelles, for lysosomal degradation. Autophagy plays roles in numerous physiological processes, include immune cell responses to endogenous and exogenous pathogenic stimuli. Moreover, autophagy has a potentially pivotal role in the regulation of inflammatory responses. In particular, autophagy regulates endogenous inflammasome activators, as well as inflammasome components and pro-IL-1β. This review focuses specifically on the role autophagy plays in regulating the production, processing and secretion of IL-1 family cytokines.

  10. Autophagy in sepsis: Degradation into exhaustion?

    Science.gov (United States)

    Ho, Jeffery; Yu, Jun; Wong, Sunny H; Zhang, Lin; Liu, Xiaodong; Wong, Wai T; Leung, Czarina C H; Choi, Gordon; Wang, Maggie H T; Gin, Tony; Chan, Matthew T V; Wu, William K K

    2016-07-01

    Autophagy is one of the innate immune defense mechanisms against microbial challenges. Previous in vitro and in vivo models of sepsis demonstrated that autophagy was activated initially in sepsis, followed by a subsequent phase of impairment. Autophagy modulation appears to be protective against multiple organ injuries in these murine sepsis models. This is achieved in part by preventing apoptosis, maintaining a balance between the productions of pro- and anti-inflammatory cytokines, and preserving mitochondrial functions. This article aims to discuss the role of autophagy in sepsis and the therapeutic potential of autophagy enhancers.

  11. Cardiac-specific catalase overexpression rescues anthrax lethal toxin-induced cardiac contractile dysfunction: role of oxidative stress and autophagy

    Directory of Open Access Journals (Sweden)

    Kandadi Machender R

    2012-11-01

    Full Text Available Abstract Background Lethal and edema toxins secreted by Bacillus anthracis during anthrax infection were found to incite serious cardiovascular complications. However, the underlying mechanisms in anthrax lethal toxin-induced cardiac anomalies remain unknown. This study was designed to evaluate the impact of antioxidant enzyme catalase in anthrax lethal toxin-induced cardiomyocyte contractile dysfunction. Methods Wild type (WT and cardiac-specific catalase overexpression mice were challenged with lethal toxin (2 μg/g, intraperotineally (i.p.. Cardiomyocyte contractile and intracellular Ca2+ properties were assessed 18 h later using an IonOptix edge-detection system. Proteasome function was assessed using chymotrypsin-like and caspase-like activities. GFP-LC3 puncta and Western blot analysis were used to evaluate autophagy and protein ubiquitination. Results Lethal toxin exposure suppressed cardiomyocyte contractile function (suppressed peak shortening, maximal velocity of shortening/re-lengthening, prolonged duration of shortening/re-lengthening, and impaired intracellular Ca2+ handling, the effects of which were alleviated by catalase. In addition, lethal toxin triggered autophagy, mitochondrial and ubiquitin-proteasome defects, the effects of which were mitigated by catalase. Pretreatment of cardiomyocytes from catalase mice with the autophagy inducer rapamycin significantly attenuated or ablated catalase-offered protection against lethal toxin-induced cardiomyocyte dysfunction. On the other hand, the autophagy inhibitor 3-MA ablated or significantly attenuated lethal toxin-induced cardiomyocyte contractile anomalies. Conclusions Our results suggest that catalase is protective against anthrax lethal toxin-induced cardiomyocyte contractile and intracellular Ca2+ anomalies, possibly through regulation of autophagy and mitochondrial function.

  12. The role of STAT3 in autophagy.

    Science.gov (United States)

    You, Liangkun; Wang, Zhanggui; Li, Hongsen; Shou, Jiawei; Jing, Zhao; Xie, Jiansheng; Sui, Xinbing; Pan, Hongming; Han, Weidong

    2015-01-01

    Autophagy is an evolutionarily conserved process in eukaryotes that eliminates harmful components and maintains cellular homeostasis in response to a series of extracellular insults. However, these insults may trigger the downstream signaling of another prominent stress responsive pathway, the STAT3 signaling pathway, which has been implicated in multiple aspects of the autophagic process. Recent reports further indicate that different subcellular localization patterns of STAT3 affect autophagy in various ways. For example, nuclear STAT3 fine-tunes autophagy via the transcriptional regulation of several autophagy-related genes such as BCL2 family members, BECN1, PIK3C3, CTSB, CTSL, PIK3R1, HIF1A, BNIP3, and microRNAs with targets of autophagy modulators. Cytoplasmic STAT3 constitutively inhibits autophagy by sequestering EIF2AK2 as well as by interacting with other autophagy-related signaling molecules such as FOXO1 and FOXO3. Additionally, the mitochondrial translocation of STAT3 suppresses autophagy induced by oxidative stress and may effectively preserve mitochondria from being degraded by mitophagy. Understanding the role of STAT3 signaling in the regulation of autophagy may provide insight into the classic autophagy model and also into cancer therapy, especially for the emerging targeted therapy, because a series of targeted agents execute antitumor activities via blocking STAT3 signaling, which inevitably affects the autophagy pathway. Here, we review several of the representative studies and the current understanding in this particular field.

  13. The natural history of ubiquitin and ubiquitin-related domains.

    Science.gov (United States)

    Burroughs, Alexander Maxwell; Iyer, Lakshminarayan M; Aravind, L

    2012-01-01

    The ubiquitin (Ub) system is centered on conjugation and deconjugation of Ub and Ub-like (Ubls) proteins by a system of ligases and peptidases, respectively. Ub/Ubls contain the beta-grasp fold, also found in numerous proteins with biochemically distinct roles unrelated to the conventional Ub-system. The beta-GF underwent an early radiation spawning at least seven clades prior to the divergence of extant organisms from their last universal common ancestor, first emerging in the context of translation-related RNA-interactions and subsequently exploding to occupy various functional niches. Most beta-GF diversification occurred in prokaryotes, with the Ubl clade showing dramatic expansion in the eukaryotes. Diversification of Ubl families in eukaryotes played a major role in emergence of characteristic eukaryotic cellular sub-structures and systems. Recent comparative genomics studies indicate precursors of the eukaryotic Ub-system emerged in prokaryotes. The simplest of these combine an Ubl and an E1-like enzyme in metabolic pathways. Sampylation in archaea and Urmylation in eukaryotes appear to represent recruitment of such systems as simple protein-tagging apparatuses. However, other prokaryotic systems incorporated further components and mirror the eukaryotic condition in possessing an E2, a RING-type E3 or both of these components. Additionally, prokaryotes have evolved conjugation systems independent of Ub ligases, such as the Pup system.

  14. The natural history of ubiquitin and ubiquitin-related domains.

    Science.gov (United States)

    Burroughs, Alexander Maxwell; Iyer, Lakshminarayan M; Aravind, L

    2012-01-01

    The ubiquitin (Ub) system is centered on conjugation and deconjugation of Ub and Ub-like (Ubls) proteins by a system of ligases and peptidases, respectively. Ub/Ubls contain the beta-grasp fold, also found in numerous proteins with biochemically distinct roles unrelated to the conventional Ub-system. The beta-GF underwent an early radiation spawning at least seven clades prior to the divergence of extant organisms from their last universal common ancestor, first emerging in the context of translation-related RNA-interactions and subsequently exploding to occupy various functional niches. Most beta-GF diversification occurred in prokaryotes, with the Ubl clade showing dramatic expansion in the eukaryotes. Diversification of Ubl families in eukaryotes played a major role in emergence of characteristic eukaryotic cellular sub-structures and systems. Recent comparative genomics studies indicate precursors of the eukaryotic Ub-system emerged in prokaryotes. The simplest of these combine an Ubl and an E1-like enzyme in metabolic pathways. Sampylation in archaea and Urmylation in eukaryotes appear to represent recruitment of such systems as simple protein-tagging apparatuses. However, other prokaryotic systems incorporated further components and mirror the eukaryotic condition in possessing an E2, a RING-type E3 or both of these components. Additionally, prokaryotes have evolved conjugation systems independent of Ub ligases, such as the Pup system. PMID:22201813

  15. Autophagy in immune cell regulation and dysregulation.

    Science.gov (United States)

    Chaturvedi, Akanksha; Pierce, Susan K

    2009-09-01

    Autophagy is an ancient pathway required for cell and tissue homeostasis and differentiation. Initially thought to be a process leading to cell death, autophagy is currently viewed as a beneficial catabolic process that promotes cell survival under starvation conditions by sequestering components of the cytoplasm, including misfolded proteins, protein aggregates, and damaged organelles, and targeting them for lysosome-mediated degradation. In this way, autophagy plays a role in maintaining a balance between degradation and recycling of cellular material. The importance of autophagy is underscored by the fact that malfunctioning of this pathway results in neurodegeneration, cancer, susceptibility to microbial infection, and premature aging. Autophagy occurs in almost all cell types, including immune cells. Recent advances in the field suggest that autophagy plays a central role in regulating the immune system at multiple levels. In this review, we focus on recent developments in the area of autophagy-mediated modulation of immune responses. PMID:19671376

  16. The symphony of autophagy and calcium signaling.

    Science.gov (United States)

    Yao, Zhiyuan; Klionsky, Daniel J

    2015-01-01

    Posttranslational regulation of macroautophagy (hereafter autophagy), including phosphorylating and dephosphorylating components of the autophagy-related (Atg) core machinery and the corresponding upstream transcriptional factors, is important for the precise modulation of autophagy levels. Several kinases that are involved in phosphorylating autophagy-related proteins have been identified in both yeast and mammalian cells. However, there has been much less research published with regard to the identification of the complementary phosphatases that function in autophagy. A recent study identified PPP3/calcineurin, a calcium-dependent phosphatase, as a regulator of autophagy, and demonstrated that one of the key targets of PPP3/calcineurin is TFEB, a master transcriptional factor that controls autophagy and lysosomal function in mammalian cells.

  17. Modulation of pathogen recognition by autophagy

    Directory of Open Access Journals (Sweden)

    Ji Eun eOh

    2012-03-01

    Full Text Available Autophagy is an ancient biological process for maintaining cellular homeostasis by degradation of long-lived cytosolic proteins and organelles. Recent studies demonstrated that autophagy is availed by immune cells to regulate innate immunity. On the one hand, cells exert direct effector function by degrading intracellular pathogens; on the other hand, autophagy modulates pathogen recognition and downstream signaling for innate immune responses. Pathogen recognition via pattern recognition receptors induces autophagy. The function of phagocytic cells is enhanced by recruitment of autophagy-related proteins. Moreover, autophagy acts as a delivery system for viral replication complexes to migrate to the endosomal compartments where virus sensing occurs. In another case, key molecules of the autophagic pathway have been found to negatively regulate immune signaling, thus preventing aberrant activation of cytokine production and consequent immune responses. In this review, we focus on the recent advances in the role of autophagy in pathogen recognition and modulation of innate immune responses.

  18. Evolution of Plant HECT Ubiquitin Ligases

    OpenAIRE

    Ignacio Marín

    2013-01-01

    HECT ubiquitin ligases are key components of the ubiquitin-proteasome system, which is present in all eukaryotes. In this study, the patterns of emergence of HECT genes in plants are described. Phylogenetic and structural data indicate that viridiplantae have six main HECT subfamilies, which arose before the split that separated green algae from the rest of plants. It is estimated that the common ancestor of all plants contained seven HECT genes. Contrary to what happened in animals, the numb...

  19. Role of ubiquitination in meiotic recombination repair

    Institute of Scientific and Technical Information of China (English)

    2010-01-01

    Programmed and unprogrammed double-strand breaks (DSBs) often arise from such physiological requirements as meiotic recombination, and exogenous insults, such as ionizing radiation (IR). Due to deleterious impacts on genome stability, DSBs must be appropriately processed and repaired in a regulatory manner. Recent investigations have indicated that ubiquitination is a critical factor in DNA damage response and meiotic recombination repair. This review summarizes the effects of proteins and complexes associated with ubiquitination with regard to homologous recombination (HR)-dependent DSB repair.

  20. Historical landmarks of autophagy research

    OpenAIRE

    Ohsumi, Yoshinori

    2013-01-01

    The year of 2013 marked the 50th anniversary of C de Duve's coining of the term “autophagy” for the degradation process of cytoplasmic constituents in the lysosome/vacuole. This year we regretfully lost this great scientist, who contributed much during the early years of research to the field of autophagy. Soon after the discovery of lysosomes by de Duve, electron microscopy revealed autophagy as a means of delivering intracellular components to the lysosome. For a long time after the discove...

  1. Autophagy deficiency in myeloid cells increases susceptibility to obesity-induced diabetes and experimental colitis.

    Science.gov (United States)

    Lee, Hae-Youn; Kim, Jinyoung; Quan, Wenying; Lee, June-Chul; Kim, Min-Soo; Kim, Seok-Hyung; Bae, Jin-Woo; Hur, Kyu Yeon; Lee, Myung-Shik

    2016-08-01

    Autophagy, which is critical for the proper turnover of organelles such as endoplasmic reticulum and mitochondria, affects diverse aspects of metabolism, and its dysregulation has been incriminated in various metabolic disorders. However, the role of autophagy of myeloid cells in adipose tissue inflammation and type 2 diabetes has not been addressed. We produced mice with myeloid cell-specific deletion of Atg7 (autophagy-related 7), an essential autophagy gene (Atg7 conditional knockout [cKO] mice). While Atg7 cKO mice were metabolically indistinguishable from control mice, they developed diabetes when bred to ob/w mice (Atg7 cKO-ob/ob mice), accompanied by increases in the crown-like structure, inflammatory cytokine expression and inflammasome activation in adipose tissue. Mφs (macrophages) from Atg7 cKO mice showed significantly higher interleukin 1 β release and inflammasome activation in response to a palmitic acid plus lipopolysaccharide combination. Moreover, a decrease in the NAD(+):NADH ratio and increase in intracellular ROS content after treatment with palmitic acid in combination with lipopolysaccharide were more pronounced in Mφs from Atg7 cKO mice, suggesting that mitochondrial dysfunction in autophagy-deficient Mφs leads to an increase in lipid-induced inflammasome and metabolic deterioration in Atg7 cKO-ob/ob mice. Atg7 cKO mice were more susceptible to experimental colitis, accompanied by increased colonic cytokine expression, T helper 1 skewing and systemic bacterial invasion. These results suggest that autophagy of Mφs is important for the control of inflammasome activation in response to metabolic or extrinsic stress, and autophagy deficiency in Mφs may contribute to the progression of metabolic syndrome associated with lipid injury and colitis. PMID:27337687

  2. Autophagy contributes to regulation of nuclear dynamics during vegetative growth and hyphal fusion in Fusarium oxysporum.

    Science.gov (United States)

    Corral-Ramos, Cristina; Roca, M Gabriela; Di Pietro, Antonio; Roncero, M Isabel G; Ruiz-Roldán, Carmen

    2015-01-01

    In the fungal pathogen Fusarium oxysporum, vegetative hyphal fusion triggers nuclear mitotic division in the invading hypha followed by migration of a nucleus into the receptor hypha and degradation of the resident nucleus. Here we examined the role of autophagy in fusion-induced nuclear degradation. A search of the F. oxysporum genome database for autophagy pathway components identified putative orthologs of 16 core autophagy-related (ATG) genes in yeast, including the ubiquitin-like protein Atg8, which is required for the formation of autophagosomal membranes. F. oxysporum Foatg8Δ mutants were generated in a strain harboring H1-cherry fluorescent protein (ChFP)-labeled nuclei to facilitate analysis of nuclear dynamics. The Foatg8Δ mutants did not show MDC-positive staining in contrast to the wild type and the FoATG8-complemented (cFoATG8) strain, suggesting that FoAtg8 is required for autophagy in F. oxysporum. The Foatg8Δ strains displayed reduced rates of hyphal growth, conidiation, and fusion, and were significantly attenuated in virulence on tomato plants and in the nonvertebrate animal host Galleria mellonella. In contrast to wild-type hyphae, which are almost exclusively composed of uninucleated hyphal compartments, the hyphae of the Foatg8Δ mutants contained a significant fraction of hyphal compartments with 2 or more nuclei. The increase in the number of nuclei per hyphal compartment was particularly evident after hyphal fusion events. Time-lapse microscopy analyses revealed abnormal mitotic patterns during vegetative growth in the Foatg8Δ mutants. Our results suggest that autophagy mediates nuclear degradation after hyphal fusion and has a general function in the control of nuclear distribution in F. oxysporum.

  3. Brazilin Limits Inflammatory Responses through Induction of Prosurvival Autophagy in Rheumatoid Fibroblast-Like Synoviocytes.

    Directory of Open Access Journals (Sweden)

    Hyunji Lee

    Full Text Available Brazilin is an active compound of Caesalpinia sappan L. (Leguminosae, which possesses pro-apoptotic and anti-inflammation potentials depending on the specific cell type. However, it is largely unknown whether autophagy is implicated in the mechanism underlying its chemotherapeutic and anti-inflammatory effects in rheumatoid arthritis (RA. Here, we show that treatment of RA fibroblast-like synoviocytes (FLS with brazilin results in enhanced level of autophagic flux, evidenced by accumulation of autophagosome and increased level of lipidated LC3 (LC3-II, which is mainly mediated by enhanced production of reactive oxygen species (ROS. Interestingly, long-term exposure of brazilin was able to restore cell survival against the cytotoxity, exclusively in RA FLS, but not in normal fibroblast. Importantly, such a restoration from brazilin-induced cytotoxity in RA FLS was completely abrogated after co-treatment with autophagy inhibitors including NH4Cl or chloroquine. Furthermore, we found that the pretreatment of RA FLS with brazilin reduced LPS- or TNF-induced NF-κB activation and the secretion of inflammatory cytokines in parallel with the enhanced autophagic flux. Such anti-NF-κB potentials of brazilin were drastically masked in RA FLS when autophagy was suppressed. These results suggest that brazilin is capable of activating autophagy exclusively in RA FLS, and such inducible autophagy promotes cell survival and limits inflammatory response.

  4. Inhibition of NF-κB promotes autophagy via JNK signaling pathway in porcine granulosa cells.

    Science.gov (United States)

    Gao, Hui; Lin, Lu; Haq, Ihtesham Ul; Zeng, Shen-Ming

    2016-04-22

    The transcription factor nuclear factor-κB (NF-κB) plays an important role in diverse processes, including cell proliferation and differentiation, apoptosis and inflammation. However, the role of NF-κB in porcine follicle development is not clearly elucidated. In this study, we demonstrated that follicle stimulating hormone (FSH) increased the level of inhibitor of NF-κB (IκB) protein and promoted the cytoplasmic localization of p65, indicating that FSH inhibits the activation of NF-κB in porcine granulosa cells. Moreover, inhibition of NF-κB by FSH or another specific inhibitor of NF-κB, pyrrolidine dithiocarbamate (PDTC), could activate JNK signaling and enhance autophagic activity in porcine granulosa cells. Knockdown of RelA (p65) Subunit of NF-κB by RNA interference abrogated the activation of JNK signaling pathway and the increase of autophagic protein expression by FSH. Meanwhile, the functional significance of FSH or PDTC-mediated autophagy were further investigated. Our results demonstrated that the increased autophagy promoted progesterone secretion in porcine granulosa cells. Blockage of autophagy by chloroquine obviated the FSH or PDTC-induced progesterone production. Taken together, these results indicate that inhibition of NF-κB increased autophagy via JNK signaling, and promote steroidogenesis in porcine granulosa cells. Our results provide new insights into the regulation and function of autophagy in mammalian follicle development. PMID:27016483

  5. Autophagy, NLRP3 inflammasome and auto-inflammatory/immune diseases.

    Science.gov (United States)

    Zhong, Zhenyu; Sanchez-Lopez, Elsa; Karin, Michael

    2016-01-01

    Loss of homeostasis, as a result of pathogen invasion or self imbalance, causes tissue damage and inflammation. In addition to its well-established role in promoting clearance of pathogens or cell corpses, inflammation is also key to drive tissue repair and regeneration. Conserved from flies to humans, a transient, well-balanced inflammatory response is critical for restoration of tissue homeostasis after damage. The absence of such a response can result in failure of tissue repair, leading to the development of devastating immunopathologies and degenerative diseases. Studies in the past decade collectively suggest that a malfunction of NLRP3 inflammasome, a key tissue damage sensor, is a dominant driver of various autoinflammatory and autoimmune diseases, including gout, rheumatoid arthritis, and lupus. It is therefore crucial to understand the biology and regulation of NLRP3 inflammasome and determine its affect in the context of various diseases. Of note, various studies suggest that autophagy, a cellular waste removal and rejuvenation process, serves an important role as a macrophage-intrinsic negative regulator of NLRP3 inflammasome. Here, we review recent advances in understanding how autophagy regulates NLRP3 inflammasome activity and discuss the implications of this regulation on the pathogenesis of autoinflammatory and autoimmune diseases. PMID:27586797

  6. The recognition of ubiquitinated proteins by the proteasome.

    Science.gov (United States)

    Grice, Guinevere L; Nathan, James A

    2016-09-01

    The ability of ubiquitin to form up to eight different polyubiquitin chain linkages generates complexity within the ubiquitin proteasome system, and accounts for the diverse roles of ubiquitination within the cell. Understanding how each type of ubiquitin linkage is correctly interpreted by ubiquitin binding proteins provides important insights into the link between chain recognition and cellular fate. A major function of ubiquitination is to signal degradation of intracellular proteins by the 26S proteasome. Lysine-48 (K48) linked polyubiquitin chains are well established as the canonical signal for proteasomal degradation, but recent studies show a role for other ubiquitin linked chains in facilitating degradation by the 26S proteasome. Here, we review how different types of polyubiquitin linkage bind to ubiquitin receptors on the 26S proteasome, how they signal degradation and discuss the implications of ubiquitin chain linkage in regulating protein breakdown by the proteasome. PMID:27137187

  7. Activation of Adenosine 2A receptor inhibits neutrophil apoptosis in an autophagy-dependent manner in mice with systemic inflammatory response syndrome.

    Science.gov (United States)

    Liu, Yang-Wuyue; Yang, Ting; Zhao, Li; Ni, Zhenhong; Yang, Nan; He, Fengtian; Dai, Shuang-Shuang

    2016-01-01

    Systemic inflammatory response syndrome (SIRS) is an overwhelming whole body inflammation caused by infectious diseases or sterile insults. Neutrophils are the dominant participants during inflammation, and their survival and death determine the initiation as well as resolution of SIRS. Apoptosis and autophagy are two fundamental cellular processes that modulating cell fate, but their correlation and regulators in neutrophils under SIRS condition have not been elucidated. In this study, we demonstrated that high dose of LPS induced both apoptosis and autophagy of neutrophils in a mouse SIRS model and LPS-stimulated neutrophils in vitro. Moreover, we found that the adenosine 2A receptor (A2AR), a known anti-inflammatory G protein-coupled receptor (GPCR), could inhibit LPS-induced neutrophil apoptosis by suppressing the LPS-induced autophagy. Activation of A2AR suppressed LPS-induced autophagy by inhibiting the ROS-JNK pathway as well as promoting GPCR βϒ subunit-AKT signaling. The A2AR-inhibited autophagy suppressed apoptosis of neutrophils by blocking caspase8, caspase3 and PARP signaling. These findings not only increase our understandings of neutrophils' fate and function in response to systemic inflammation, but also identify a novel anti-inflammatory role of A2AR in modulating neutrophils' survival during inflammation. PMID:27647162

  8. Regulation of DNA double-strand break repair by ubiquitin and ubiquitin-like modifiers

    DEFF Research Database (Denmark)

    Schwertman, Petra; Bekker-Jensen, Simon; Mailand, Niels

    2016-01-01

    DNA double-strand breaks (DSBs) are highly cytotoxic DNA lesions. The swift recognition and faithful repair of such damage is crucial for the maintenance of genomic stability, as well as for cell and organismal fitness. Signalling by ubiquitin, SUMO and other ubiquitin-like modifiers (UBLs...

  9. Sucrose induces vesicle accumulation and autophagy.

    Science.gov (United States)

    Higuchi, Takahiro; Nishikawa, Jun; Inoue, Hiroko

    2015-04-01

    It has been shown that the treatment of mammalian cells with sucrose leads to vacuole accumulation associated with lysosomes and upregulation of lysosomal enzyme expression and activity. Autophagy is an evolutionarily conserved homeostatic process by which cells deliver cytoplasmic material for degradation into lysosomes, thus it is probable that sucrose affects the autophagic activity. The role of sucrose in autophagy is unknown; however, another disaccharide, trehalose has been shown to induce autophagy. In the current study, we used mouse embryonic fibroblasts to investigate whether sucrose induces autophagy and whether vesicle formation is associated with autophagy. The results showed that sucrose induces autophagy while being accumulated within the endosomes/lysosomes. These vesicles were swollen and packed within the cytoplasm. Furthermore, trehalose and the trisaccharide raffinose, which are not hydrolyzed in mammalian cells, increased the rate of vesicles accumulation and LC3-II level (a protein marker of autophagy). However, fructose and maltose did not show the same effects. The correlation between the two processes, vesicle accumulation and autophagy induction, was confirmed by treatment of cells with sucrose plus invertase, or maltose plus acarbose-the α-glucosidase inhibitor-and by sucrose deprivation. Results also showed that vesicle accumulation was not affected by autophagy inhibition. Therefore, the data suggest that sucrose-induced autophagy through accumulation of sucrose-containing vesicles is caused by the absence of hydrolysis enzymes.

  10. Sucrose induces vesicle accumulation and autophagy.

    Science.gov (United States)

    Higuchi, Takahiro; Nishikawa, Jun; Inoue, Hiroko

    2015-04-01

    It has been shown that the treatment of mammalian cells with sucrose leads to vacuole accumulation associated with lysosomes and upregulation of lysosomal enzyme expression and activity. Autophagy is an evolutionarily conserved homeostatic process by which cells deliver cytoplasmic material for degradation into lysosomes, thus it is probable that sucrose affects the autophagic activity. The role of sucrose in autophagy is unknown; however, another disaccharide, trehalose has been shown to induce autophagy. In the current study, we used mouse embryonic fibroblasts to investigate whether sucrose induces autophagy and whether vesicle formation is associated with autophagy. The results showed that sucrose induces autophagy while being accumulated within the endosomes/lysosomes. These vesicles were swollen and packed within the cytoplasm. Furthermore, trehalose and the trisaccharide raffinose, which are not hydrolyzed in mammalian cells, increased the rate of vesicles accumulation and LC3-II level (a protein marker of autophagy). However, fructose and maltose did not show the same effects. The correlation between the two processes, vesicle accumulation and autophagy induction, was confirmed by treatment of cells with sucrose plus invertase, or maltose plus acarbose-the α-glucosidase inhibitor-and by sucrose deprivation. Results also showed that vesicle accumulation was not affected by autophagy inhibition. Therefore, the data suggest that sucrose-induced autophagy through accumulation of sucrose-containing vesicles is caused by the absence of hydrolysis enzymes. PMID:25389129

  11. Crohn's disease-associated adherent-invasive E. coli are selectively favoured by impaired autophagy to replicate intracellularly.

    Science.gov (United States)

    Lapaquette, Pierre; Glasser, Anne-Lise; Huett, Alan; Xavier, Ramnik J; Darfeuille-Michaud, Arlette

    2010-01-01

    Ileal lesions in Crohn's disease (CD) patients are colonized by pathogenic adherent-invasive Escherichia coli (AIEC) able to invade and to replicate within intestinal epithelial cells. Recent genome-wide association studies have highlighted the autophagy pathway as being associated with CD risk. In the present study we investigated whether defects in autophagy enhance replication of commensal and pathogenic Escherichia coli and CD-associated AIEC. We show that functional autophagy limits intracellular AIEC replication and that a subpopulation of the intracellular bacteria is located within LC3-positive autophagosomes. In IRGM and ATG16L1 deficient cells intracellular AIEC LF82 bacteria have enhanced replication. Surprisingly autophagy deficiency did not interfere with the ability of intracellular bacteria to survive and/or replicate for any other E. coli strains tested, including non-pathogenic, environmental, commensal, or pathogenic strains involved in gastro enteritis. Together these findings demonstrate a central role for autophagy restraining Adherent-Invasive E. coli strains associated with ileal CD. AIEC infection in patients with polymorphisms in autophagy genes may have a significant impact on the outcome of intestinal inflammation. PMID:19747213

  12. Ubiquitination of specific mitochondrial matrix proteins.

    Science.gov (United States)

    Lehmann, Gilad; Ziv, Tamar; Braten, Ori; Admon, Arie; Udasin, Ronald G; Ciechanover, Aaron

    2016-06-17

    Several protein quality control systems in bacteria and/or mitochondrial matrix from lower eukaryotes are absent in higher eukaryotes. These are transfer-messenger RNA (tmRNA), The N-end rule ATP-dependent protease ClpAP, and two more ATP-dependent proteases, HslUV and ClpXP (in yeast). The lost proteases resemble the 26S proteasome and the role of tmRNA and the N-end rule in eukaryotic cytosol is performed by the ubiquitin proteasome system (UPS). Therefore, we hypothesized that the UPS might have substituted these systems - at least partially - in the mitochondrial matrix of higher eukaryotes. Using three independent experimental approaches, we demonstrated the presence of ubiquitinated proteins in the matrix of isolated yeast mitochondria. First, we show that isolated mitochondria contain ubiquitin (Ub) conjugates, which remained intact after trypsin digestion. Second, we demonstrate that the mitochondrial soluble fraction contains Ub-conjugates, several of which were identified by mass spectrometry and are localized to the matrix. Third, using immunoaffinity enrichment by specific antibodies recognizing digested ubiquitinated peptides, we identified a group of Ub-modified matrix proteins. The modification was further substantiated by separation on SDS-PAGE and immunoblots. Last, we attempted to identify the ubiquitin ligase(s) involved, and identified Dma1p as a trypsin-resistant protein in our mitochondrial preparations. Taken together, these data suggest a yet undefined role for the UPS in regulation of the mitochondrial matrix proteins. PMID:27157140

  13. Dengue Virus Genome Uncoating Requires Ubiquitination

    Directory of Open Access Journals (Sweden)

    Laura A. Byk

    2016-06-01

    Full Text Available The process of genome release or uncoating after viral entry is one of the least-studied steps in the flavivirus life cycle. Flaviviruses are mainly arthropod-borne viruses, including emerging and reemerging pathogens such as dengue, Zika, and West Nile viruses. Currently, dengue virus is one of the most significant human viral pathogens transmitted by mosquitoes and is responsible for about 390 million infections every year around the world. Here, we examined for the first time molecular aspects of dengue virus genome uncoating. We followed the fate of the capsid protein and RNA genome early during infection and found that capsid is degraded after viral internalization by the host ubiquitin-proteasome system. However, proteasome activity and capsid degradation were not necessary to free the genome for initial viral translation. Unexpectedly, genome uncoating was blocked by inhibiting ubiquitination. Using different assays to bypass entry and evaluate the first rounds of viral translation, a narrow window of time during infection that requires ubiquitination but not proteasome activity was identified. In this regard, ubiquitin E1-activating enzyme inhibition was sufficient to stabilize the incoming viral genome in the cytoplasm of infected cells, causing its retention in either endosomes or nucleocapsids. Our data support a model in which dengue virus genome uncoating requires a nondegradative ubiquitination step, providing new insights into this crucial but understudied viral process.

  14. Cellular contractility requires ubiquitin mediated proteolysis.

    Directory of Open Access Journals (Sweden)

    Yuval Cinnamon

    Full Text Available BACKGROUND: Cellular contractility, essential for cell movement and proliferation, is regulated by microtubules, RhoA and actomyosin. The RhoA dependent kinase ROCK ensures the phosphorylation of the regulatory Myosin II Light Chain (MLC Ser19, thereby activating actomyosin contractions. Microtubules are upstream inhibitors of contractility and their depolymerization or depletion cause cells to contract by activating RhoA. How microtubule dynamics regulates RhoA remains, a major missing link in understanding contractility. PRINCIPAL FINDINGS: We observed that contractility is inhibited by microtubules not only, as previously reported, in adherent cells, but also in non-adhering interphase and mitotic cells. Strikingly we observed that contractility requires ubiquitin mediated proteolysis by a Cullin-RING ubiquitin ligase. Inhibition of proteolysis, ubiquitination and neddylation all led to complete cessation of contractility and considerably reduced MLC Ser19 phosphorylation. CONCLUSIONS: Our results imply that cells express a contractility inhibitor that is degraded by ubiquitin mediated proteolysis, either constitutively or in response to microtubule depolymerization. This degradation seems to depend on a Cullin-RING ubiquitin ligase and is required for cellular contractions.

  15. Dengue Virus Genome Uncoating Requires Ubiquitination

    Science.gov (United States)

    Byk, Laura A.; Iglesias, Néstor G.; De Maio, Federico A.; Gebhard, Leopoldo G.; Rossi, Mario

    2016-01-01

    ABSTRACT The process of genome release or uncoating after viral entry is one of the least-studied steps in the flavivirus life cycle. Flaviviruses are mainly arthropod-borne viruses, including emerging and reemerging pathogens such as dengue, Zika, and West Nile viruses. Currently, dengue virus is one of the most significant human viral pathogens transmitted by mosquitoes and is responsible for about 390 million infections every year around the world. Here, we examined for the first time molecular aspects of dengue virus genome uncoating. We followed the fate of the capsid protein and RNA genome early during infection and found that capsid is degraded after viral internalization by the host ubiquitin-proteasome system. However, proteasome activity and capsid degradation were not necessary to free the genome for initial viral translation. Unexpectedly, genome uncoating was blocked by inhibiting ubiquitination. Using different assays to bypass entry and evaluate the first rounds of viral translation, a narrow window of time during infection that requires ubiquitination but not proteasome activity was identified. In this regard, ubiquitin E1-activating enzyme inhibition was sufficient to stabilize the incoming viral genome in the cytoplasm of infected cells, causing its retention in either endosomes or nucleocapsids. Our data support a model in which dengue virus genome uncoating requires a nondegradative ubiquitination step, providing new insights into this crucial but understudied viral process. PMID:27353759

  16. Ubiquitination of specific mitochondrial matrix proteins.

    Science.gov (United States)

    Lehmann, Gilad; Ziv, Tamar; Braten, Ori; Admon, Arie; Udasin, Ronald G; Ciechanover, Aaron

    2016-06-17

    Several protein quality control systems in bacteria and/or mitochondrial matrix from lower eukaryotes are absent in higher eukaryotes. These are transfer-messenger RNA (tmRNA), The N-end rule ATP-dependent protease ClpAP, and two more ATP-dependent proteases, HslUV and ClpXP (in yeast). The lost proteases resemble the 26S proteasome and the role of tmRNA and the N-end rule in eukaryotic cytosol is performed by the ubiquitin proteasome system (UPS). Therefore, we hypothesized that the UPS might have substituted these systems - at least partially - in the mitochondrial matrix of higher eukaryotes. Using three independent experimental approaches, we demonstrated the presence of ubiquitinated proteins in the matrix of isolated yeast mitochondria. First, we show that isolated mitochondria contain ubiquitin (Ub) conjugates, which remained intact after trypsin digestion. Second, we demonstrate that the mitochondrial soluble fraction contains Ub-conjugates, several of which were identified by mass spectrometry and are localized to the matrix. Third, using immunoaffinity enrichment by specific antibodies recognizing digested ubiquitinated peptides, we identified a group of Ub-modified matrix proteins. The modification was further substantiated by separation on SDS-PAGE and immunoblots. Last, we attempted to identify the ubiquitin ligase(s) involved, and identified Dma1p as a trypsin-resistant protein in our mitochondrial preparations. Taken together, these data suggest a yet undefined role for the UPS in regulation of the mitochondrial matrix proteins.

  17. Molecular mechanism and regulation of autophagy

    Institute of Scientific and Technical Information of China (English)

    Ya-ping YANG; Zhong-qin LIANG; Zhen-lun GU; Zheng-hong QIN

    2005-01-01

    Autophagy is a major cellular pathway for the degradation of long-lived proteins and cytoplasmic organelles in eukaryotic cells. A large number of intracellular/extracellular stimuli, including amino acid starvation and invasion of microorganisms, are able to induce the autophagic response in cells. The discovery of the ATG genes in yeast has greatly advanced our understanding of the molecular mechanisms participating in autophagy and the genes involved in regulating the autophagic pathway. Many yeast genes have mammalian homologs,suggesting that the basic machinery for autophagy has been evolutionarily conserved along the eukaryotic phylum. The regulation of autophagy is a very complex process. Many signaling pathways, including target of rapamycin (TOR) or mammalian target of rapamycin (mTOR), phosphatidylinositol 3-kinase-I (PI3K-I)/PKB, GTPases, calcium and protein synthesis all play important roles in regulating autophagy. The molecular mechanisms and regulation of autophagy are discussed in this review.

  18. Feedback regulation between autophagy and PKA.

    Science.gov (United States)

    Torres-Quiroz, Francisco; Filteau, Marie; Landry, Christian R

    2015-01-01

    Protein kinase A (PKA) controls diverse cellular processes and homeostasis in eukaryotic cells. Many processes and substrates of PKA have been described and among them are direct regulators of autophagy. The mechanisms of PKA regulation and how they relate to autophagy remain to be fully understood. We constructed a reporter of PKA activity in yeast to identify genes affecting PKA regulation. The assay systematically measures relative protein-protein interactions between the regulatory and catalytic subunits of the PKA complex in a systematic set of genetic backgrounds. The candidate PKA regulators we identified span multiple processes and molecular functions (autophagy, methionine biosynthesis, TORC signaling, protein acetylation, and DNA repair), which themselves include processes regulated by PKA. These observations suggest the presence of many feedback loops acting through this key regulator. Many of the candidate regulators include genes involved in autophagy, suggesting that not only does PKA regulate autophagy but that autophagy also sends signals back to PKA.

  19. Autophagy and IL-1 family cytokines

    Directory of Open Access Journals (Sweden)

    James eHarris

    2013-04-01

    Full Text Available Autophagy is an important intracellular homeostatic mechanism for the targeting of cytosolic constituents, including organelles, for lysosomal degradation. Autophagy plays roles in numerous physiological processes, including immune cell responses to endogenous and exogenous pathogenic stimuli. Moreover, autophagy has a potentially pivotal role to play in the regulation of inflammatory responses. In particular, autophagy regulates endogenous inflammasome activators, as well as inflammasome components and pro-IL-1β. As a result, autophagy acts a key modulator of IL-1β and IL-18, as well as IL-1α, release. This review focuses specifically on the role autophagy plays in regulating the production, processing and secretion of IL-1 and IL-18 and the consequences of this important function.

  20. Autophagy and IL-1 Family Cytokines.

    Science.gov (United States)

    Harris, James

    2013-01-01

    Autophagy is an important intracellular homeostatic mechanism for the targeting of cytosolic constituents, including organelles, for lysosomal degradation. Autophagy plays roles in numerous physiological processes, including immune cell responses to endogenous and exogenous pathogenic stimuli. Moreover, autophagy has a potentially pivotal role to play in the regulation of inflammatory responses. In particular, autophagy regulates endogenous inflammasome activators, as well as inflammasome components and pro-IL-1β. As a result, autophagy acts a key modulator of IL-1β and IL-18, as well as IL-1α, release. This review focuses specifically on the role autophagy plays in regulating the production, processing, and secretion of IL-1 and IL-18 and the consequences of this important function.

  1. Heat shock induced change in protein ubiquitination in Chlamydomonas

    International Nuclear Information System (INIS)

    Ubiquitin was purified from pea (Pisum sativum L.) and its antibody was produced. Western blot analysis showed that the antibody cross-reacted with ubiquitins from a green alga Chlamydomonas reinhardtii, a brown alga Laminaria angustata and a red alga Porphyridium cruentum but not with ubiquitin from a blue-green alga Synechococcus sp. In Chlamydomonas, the antibody also reacted with some ubiquitinated proteins including 28- and 31-kDa polypeptides. The isoelectric points of Chlamydomonas ubiquitin and the 28- and 31-kDa ubiquitinated proteins were 8.0, 8.9 and 10.3, respectively. The ubiquitinated proteins, including the 28- and 31-kDa polypeptides were detected after in vitro ATP-dependent ubiquitination of Chlamydomonas cell extract with l25I-labeled bovine ubiquitin. Heat treatment of Chlamydomonas cells (>40°C) caused drastic increase of ubiquitinated proteins with high mol wt (>60kDa), and coordinated redistribution or decrease of other ubiquitinated proteins and free ubiquitin. Quantitative analysis revealed that the 28- and 31-kDa ubiquitinated proteins showed different responses against heat stress, i.e. the former being more sensitive than the latter. (author)

  2. Investigating regulatory signatures of human autophagy related gene 5 (ATG5 through functional in silico analysis

    Directory of Open Access Journals (Sweden)

    Avni Vij

    2016-09-01

    Full Text Available Autophagy is an essential, homeostatic process which removes damaged cellular proteins and organelles for cellular renewal. ATG5, a part of E3 ubiquitin ligase-like complex (Atg12-Atg5/Atg16L1, is a key regulator involved in autophagosome formation - a crucial phase of autophagy. In this study, we used different in silico methods for comprehensive analysis of ATG5 to investigate its less explored regulatory activity. We have predicted various physico-chemical parameters and two possible transmembrane models that helped in exposing its functional regions. Twenty four PTM sites and 44 TFBS were identified which could be targeted to modulate the autophagy pathway. Furthermore, LD analysis identified 3 blocks of genotyped SNPs and 2 deleterious nsSNPs that may have damaging impact on protein function and thus could be employed for carrying genome-wide association studies. In conclusion, the information obtained in this study could be helpful for better understanding of regulatory roles of ATG5 and provides a base for its implication in population-based studies.

  3. FGF signalling regulates bone growth through autophagy.

    Science.gov (United States)

    Cinque, Laura; Forrester, Alison; Bartolomeo, Rosa; Svelto, Maria; Venditti, Rossella; Montefusco, Sandro; Polishchuk, Elena; Nusco, Edoardo; Rossi, Antonio; Medina, Diego L; Polishchuk, Roman; De Matteis, Maria Antonietta; Settembre, Carmine

    2015-12-10

    Skeletal growth relies on both biosynthetic and catabolic processes. While the role of the former is clearly established, how the latter contributes to growth-promoting pathways is less understood. Macroautophagy, hereafter referred to as autophagy, is a catabolic process that plays a fundamental part in tissue homeostasis. We investigated the role of autophagy during bone growth, which is mediated by chondrocyte rate of proliferation, hypertrophic differentiation and extracellular matrix (ECM) deposition in growth plates. Here we show that autophagy is induced in growth-plate chondrocytes during post-natal development and regulates the secretion of type II collagen (Col2), the major component of cartilage ECM. Mice lacking the autophagy related gene 7 (Atg7) in chondrocytes experience endoplasmic reticulum storage of type II procollagen (PC2) and defective formation of the Col2 fibrillary network in the ECM. Surprisingly, post-natal induction of chondrocyte autophagy is mediated by the growth factor FGF18 through FGFR4 and JNK-dependent activation of the autophagy initiation complex VPS34-beclin-1. Autophagy is completely suppressed in growth plates from Fgf18(-/-) embryos, while Fgf18(+/-) heterozygous and Fgfr4(-/-) mice fail to induce autophagy during post-natal development and show decreased Col2 levels in the growth plate. Strikingly, the Fgf18(+/-) and Fgfr4(-/-) phenotypes can be rescued in vivo by pharmacological activation of autophagy, pointing to autophagy as a novel effector of FGF signalling in bone. These data demonstrate that autophagy is a developmentally regulated process necessary for bone growth, and identify FGF signalling as a crucial regulator of autophagy in chondrocytes. PMID:26595272

  4. Oxidative Stress and Autophagy in Cardiovascular Homeostasis

    OpenAIRE

    Morales, Cyndi R.; Pedrozo, Zully; Lavandero, Sergio; Hill, Joseph A.

    2014-01-01

    Significance: Autophagy is an evolutionarily ancient process of intracellular protein and organelle recycling required to maintain cellular homeostasis in the face of a wide variety of stresses. Dysregulation of reactive oxygen species (ROS) and reactive nitrogen species (RNS) leads to oxidative damage. Both autophagy and ROS/RNS serve pathological or adaptive roles within cardiomyocytes, depending on the context. Recent Advances: ROS/RNS and autophagy communicate with each other via both tra...

  5. Autophagy and oxidative stress in cardiovascular diseases

    OpenAIRE

    Mei, Yu; Thompson, Melissa D.; Cohen, Richard A.; Tong, XiaoYong

    2014-01-01

    Autophagy is a highly conserved degradation process by which intracellular components, including soluble macromolecules (e.g. nucleic acids, proteins, carbohydrates, and lipids) and dysfunctional organelles (e.g. mitochondria, ribosomes, peroxisomes, and endoplasmic reticulum) are degraded by the lysosome. Autophagy is orchestrated by the autophagy related protein (Atg) composed protein complexes to form autophagosomes, which fuse with lysosomes to generate autolysosomes where the contents ar...

  6. The role of autophagy in Parkinson's disease

    Institute of Scientific and Technical Information of China (English)

    Lei Zhang; Yaru Dong; Xiaoheng Xu; Zhong Xu

    2012-01-01

    Although Parkinson's disease is the most common neurodegenerative movement disorder, the mechanisms of pathogenesis remain poorly understood. Recent findings have shown that deregulation of the autophagy-lysosome pathway is involved in the pathogenesis of Parkinson's disease. This review summarizes the most recent findings and discusses the unique role of the autophagy-lysosome pathway in Parkinson's disease to highlight the possibility of Parkinson's disease treatment strategies that incorporate autophagy-lysosome pathway modulation.

  7. Autophagy gets in on the regulatory act

    Institute of Scientific and Technical Information of China (English)

    Steven K. Backues; Daniel J. Klionsky

    2011-01-01

    Autophagy down-regulates the Wnt signal transduction pathway via targeted degradation of a key signaling protein. This may provide an explanation for autophagy's role in tumor suppression.%@@ The eukaryotic cell has at its disposal two primary methods for getting rid of unwanted proteins: the proteasome and autophagy.The proteasome is a large protein complex comprising regulatory and proteolytic subunits whose core function is the degradation of damaged or misfolded proteins.

  8. The Molecular Basis for Ubiquitin and Ubiquitin-like Specificities in Bacterial Effector Proteases.

    Science.gov (United States)

    Pruneda, Jonathan N; Durkin, Charlotte H; Geurink, Paul P; Ovaa, Huib; Santhanam, Balaji; Holden, David W; Komander, David

    2016-07-21

    Pathogenic bacteria rely on secreted effector proteins to manipulate host signaling pathways, often in creative ways. CE clan proteases, specific hydrolases for ubiquitin-like modifications (SUMO and NEDD8) in eukaryotes, reportedly serve as bacterial effector proteins with deSUMOylase, deubiquitinase, or, even, acetyltransferase activities. Here, we characterize bacterial CE protease activities, revealing K63-linkage-specific deubiquitinases in human pathogens, such as Salmonella, Escherichia, and Shigella, as well as ubiquitin/ubiquitin-like cross-reactive enzymes in Chlamydia, Rickettsia, and Xanthomonas. Five crystal structures, including ubiquitin/ubiquitin-like complexes, explain substrate specificities and redefine relationships across the CE clan. Importantly, this work identifies novel family members and provides key discoveries among previously reported effectors, such as the unexpected deubiquitinase activity in Xanthomonas XopD, contributed by an unstructured ubiquitin binding region. Furthermore, accessory domains regulate properties such as subcellular localization, as exemplified by a ubiquitin-binding domain in Salmonella Typhimurium SseL. Our work both highlights and explains the functional adaptations observed among diverse CE clan proteins. PMID:27425412

  9. HERC 1 ubiquitin ligase mutation affects neocortical, CA3 hippocampal and spinal cord projection neurons. An ultrastructural study

    Directory of Open Access Journals (Sweden)

    Rocío eRuiz

    2016-04-01

    Full Text Available The spontaneous mutation tambaleante is caused by the Gly483Glu substitution in the highly conserved N terminal RCC1-like domain of the HERC1 protein, which leads to the increase of mutated protein levels responsible for cerebellar Purkinje cell death by autophagy. Until now, Purkinje cells have been the only central nervous neurons reported as being targeted by the mutation, and their degeneration elicits an ataxic syndrome in adult mutant mice. However, the ultrastructural analysis performed here demonstrates that signs of autophagy, such as autophagosomes, lysosomes, and altered mitochondria, are present in neocortical pyramidal, CA3 hippocampal pyramidal, and spinal cord motor neurons. The main difference is that the reduction in the number of neurons affected in the tambaleante mutation in the neocortex, the hippocampus, and the spinal cord is not so evident as the dramatic loss of cerebellar Purkinje cells. Interestingly, signs of autophagy are absent in both interneurons and neuroglia cells. Affected neurons have in common that they are projection neurons which receive strong and varied synaptic inputs, and possess the highest degree of neuronal activity. Therefore, because the integrity of the ubiquitin-proteasome system is essential for protein degradation and, hence, for normal protein turnover, it could be hypothesized that the deleterious effects of the misrouting of these pathways would depend directly on the neuronal activity.

  10. HERC 1 Ubiquitin Ligase Mutation Affects Neocortical, CA3 Hippocampal and Spinal Cord Projection Neurons: An Ultrastructural Study.

    Science.gov (United States)

    Ruiz, Rocío; Pérez-Villegas, Eva María; Bachiller, Sara; Rosa, José Luis; Armengol, José Angel

    2016-01-01

    The spontaneous mutation tambaleante is caused by the Gly483Glu substitution in the highly conserved N terminal RCC1-like domain of the HERC1 protein, which leads to the increase of mutated protein levels responsible for cerebellar Purkinje cell death by autophagy. Until now, Purkinje cells have been the only central nervous neurons reported as being targeted by the mutation, and their degeneration elicits an ataxic syndrome in adult mutant mice. However, the ultrastructural analysis performed here demonstrates that signs of autophagy, such as autophagosomes, lysosomes, and altered mitochondria, are present in neocortical pyramidal, CA3 hippocampal pyramidal, and spinal cord motor neurons. The main difference is that the reduction in the number of neurons affected in the tambaleante mutation in the neocortex, the hippocampus, and the spinal cord is not so evident as the dramatic loss of cerebellar Purkinje cells. Interestingly, signs of autophagy are absent in both interneurons and neuroglia cells. Affected neurons have in common that they are projection neurons which receive strong and varied synaptic inputs, and possess the highest degree of neuronal activity. Therefore, because the integrity of the ubiquitin-proteasome system is essential for protein degradation and hence, for normal protein turnover, it could be hypothesized that the deleterious effects of the misrouting of these pathways would depend directly on the neuronal activity. PMID:27147983

  11. Modulating autophagy: a strategy for cancer therapy

    Institute of Scientific and Technical Information of China (English)

    Jun-Lin Li; Shao-Liang Han; Xia Fan

    2011-01-01

    Autophagy is a process in which long-lived proteins,damaged cell organelles,and other cellular particles are sequestered and degraded.This process is important for maintaining the cellular microenvironment when the cell is under stress.Many studies have shown that autophagy plays a complex role in human diseases,especially in cancer,where it is known to have paradoxical effects.Namely,autophagy provides the energy for metabolism and tumor growth and leads to cell death that promotes tumor suppression.The link between autophagy and cancer is also evident in that some of the genes that regulate carcinogenesis,oncogenes and tumor suppressor genes,participate in or impact the autophagy process.Therefore,modulating autophagy will be a valuable topic for cancer therapy.Many studies have shown that autophagy can inhibit the tumor growth when autophagy modulators are combined with radiotherapy and/or chemotherapy.These findings suggest that autophagy may be a potent target for cancer therapy.

  12. Autophagy and the nutritional signaling pathway

    Directory of Open Access Journals (Sweden)

    Long HE,Shabnam ESLAMFAM,Xi MA,Defa LI

    2016-09-01

    Full Text Available During their growth and development, animals adapt to tremendous changes in order to survive. These include responses to both environmental and physiological changes and autophagy is one of most important adaptive and regulatory mechanisms. Autophagy is defined as an autolytic process to clear damaged cellular organelles and recycle the nutrients via lysosomic degradation. The process of autophagy responds to special conditions such as nutrient withdrawal. Once autophagy is induced, phagophores form and then elongate and curve to form autophagosomes. Autophagosomes then engulf cargo, fuse with endosomes, and finally fuse with lysosomes for maturation. During the initiation process, the ATG1/ULK1 (unc-51-like kinase 1 and VPS34 (which encodes a class III phosphatidylinositol (PtdIns 3-kinase complexes are critical in recruitment and assembly of other complexes required for autophagy. The process of autophagy is regulated by autophagy related genes (ATGs. Amino acid and energy starvation mediate autophagy by activating mTORC1 (mammalian target of rapamycin and AMP-activated protein kinase (AMPK. AMPK is the energy status sensor, the core nutrient signaling component and the metabolic kinase of cells. This review mainly focuses on the mechanism of autophagy regulated by nutrient signaling especially for the two important complexes, ULK1 and VPS34.

  13. Homocysteine thiolactone affects protein ubiquitination in yeast.

    Science.gov (United States)

    Bretes, Ewa; Zimny, Jarosław

    2013-01-01

    The formation of homocysteine thiolactone (HcyTl) from homocysteine occurs in all examined so far organisms including bacteria, yeast, and humans. Protein N-homocysteinylation at the ε-amino group of lysine is an adverse result of HcyTl accumulation. Since tagging of proteins by ubiquitination before their proteasomal degradation takes place at the same residue, we wondered how N-homocysteinylation may affect the ubiquitination of proteins. We used different yeast strains carrying mutations in genes involved in the homocysteine metabolism. We found positive correlation between the concentration of endogenous HcyTl and the concentration of ubiquitinated proteins. This suggests that N-homocysteinylation of proteins apparently does not preclude but rather promotes their decomposition. PMID:24051443

  14. 自噬调节与肿瘤治疗%Autophagy modulation in cancer therapy

    Institute of Scientific and Technical Information of China (English)

    杨晓亮

    2011-01-01

    自噬是一个细胞质成分被双层膜的囊泡包裹并与溶酶体融合降解的过程.肿瘤细胞利用自噬过程存活于代谢应激下,因此推测自噬抑制剂在肿瘤治疗中具有临床应用价值.与此相矛盾的是在人类乳腺肿瘤、卵巢肿瘤、前列腺肿瘤中,重要的自噬基因beclinl等位性丢失发生很频繁,表明自噬的受损促使肿瘤的发生.与自噬缺陷和对代谢应激损伤耐受有关的肿瘤,其发生的可能原因包括增加炎症反应导致的细胞死亡及细胞因子的产生,染色体组的损伤.这表明自噬诱导剂通过限制细胞基因组损伤、细胞死亡及炎症来预防肿瘤可能具有价值.因为自噬是细胞应激反应的重要组成部分,所以通过调节自噬过程进行肿瘤的预防及治疗将是一个很有前景的新领域.%Autophagy is a process by which cytoplasmic components are sequestered in double membrane vesicles and degraded upon fusion with lysosomal compartments. Autophagy is utilized by tumor cells to survive meta bolic stress and as such autophagy inhibitors are predicted to have clinical value in the setting of cancer therapy. Paradoxically, allelic loss of the essential autophagy gene beclinl is found with high frequency in human breast, ovarian and prostate cancers, suggesting that impaired autophagy promotes cancer. Probable causes of cancer associated with defects in autophagy and impaired tolerance to stress include enhanced cell death resulting in an inflammatory response and cytokine production, and also genome damage. This suggests that autophagy inducers would also be valuable in the setting of cancer prevention to limit cellular genome damage, cell death and inflammation. As autophagy is a major component of the cellular stress response, modulating the autophagy pathway is a promising new area in cancer prevention and therapy.

  15. Carboxyl terminus of Hsp70-interacting protein (CHIP) is required to modulate cardiac hypertrophy and attenuate autophagy during exercise.

    Science.gov (United States)

    Willis, Monte S; Min, Jin-Na; Wang, Shaobin; McDonough, Holly; Lockyer, Pamela; Wadosky, Kristine M; Patterson, Cam

    2013-12-01

    The carboxyl terminus of Hsp70-interacting protein (CHIP) is a ubiquitin ligase/cochaperone critical for the maintenance of cardiac function. Mice lacking CHIP (CHIP-/-) suffer decreased survival, enhanced myocardial injury and increased arrhythmias compared with wild-type controls following challenge with cardiac ischaemia reperfusion injury. Recent evidence implicates a role for CHIP in chaperone-assisted selective autophagy, a process that is associated with exercise-induced cardioprotection. To determine whether CHIP is involved in cardiac autophagy, we challenged CHIP-/- mice with voluntary exercise. CHIP-/- mice respond to exercise with an enhanced autophagic response that is associated with an exaggerated cardiac hypertrophy phenotype. No impairment of function was identified in the CHIP-/- mice by serial echocardiography over the 5 weeks of running, indicating that the cardiac hypertrophy was physiologic not pathologic in nature. It was further determined that CHIP plays a role in inhibiting Akt signalling and autophagy determined by autophagic flux in cardiomyocytes and in the intact heart. Taken together, cardiac CHIP appears to play a role in regulating autophagy during the development of cardiac hypertrophy, possibly by its role in supporting Akt signalling, induced by voluntary running in vivo.

  16. Mechanistically linking age-related diseases and dietary carbohydrate via autophagy and the ubiquitin proteolytic systems

    Science.gov (United States)

    Epidemiological data indicate that consuming diets that deliver sugar to the blood rapidly (called high glycemic index, GI) is associated with enhanced risk for age-related diseases such as cardiovascular disease, type 2 diabetes, cataract and age-related macular degeneration (AMD). These debilities...

  17. Autophagy Constitutes a Protective Mechanism against Ethanol Toxicity in Mouse Astrocytes and Neurons.

    Science.gov (United States)

    Pla, Antoni; Pascual, María; Guerri, Consuelo

    2016-01-01

    Ethanol induces brain damage and neurodegeneration by triggering inflammatory processes in glial cells through activation of Toll-like receptor 4 (TLR4) signaling. Recent evidence indicates the role of protein degradation pathways in neurodegeneration and alcoholic liver disease, but how these processes affect the brain remains elusive. We have demonstrated that chronic ethanol consumption impairs proteolytic pathways in mouse brain, and the immune response mediated by TLR4 receptors participates in these dysfunctions. We evaluate the in vitro effects of an acute ethanol dose on the autophagy-lysosome pathway (ALP) on WT and TLR4-/- mouse astrocytes and neurons in primary culture, and how these changes affect cell survival. Our results show that ethanol induces overexpression of several autophagy markers (ATG12, LC3-II, CTSB), and increases the number of lysosomes in WT astrocytes, effects accompanied by a basification of lysosomal pH and by lowered phosphorylation levels of autophagy inhibitor mTOR, along with activation of complexes beclin-1 and ULK1. Notably, we found only minor changes between control and ethanol-treated TLR4-/- mouse astroglial cells. Ethanol also triggers the expression of the inflammatory mediators iNOS and COX-2, but induces astroglial death only slightly. Blocking autophagy by using specific inhibitors increases both inflammation and cell death. Conversely, in neurons, ethanol down-regulates the autophagy pathway and triggers cell death, which is partially recovered by using autophagy enhancers. These results support the protective role of the ALP against ethanol-induced astroglial cell damage in a TLR4-dependent manner, and provide new insight into the mechanisms that underlie ethanol-induced brain damage and are neuronal sensitive to the ethanol effects.

  18. Role of autophagy in the bimodal stage after spinal cord ischemia reperfusion injury in rats.

    Science.gov (United States)

    Fang, Bo; Li, Xiao-Qian; Bao, Na-Ren; Tan, Wen-Fei; Chen, Feng-Shou; Pi, Xiao-Li; Zhang, Ying; Ma, Hong

    2016-07-22

    Autophagy plays an important role in spinal cord ischemia reperfusion (I/R) injury, but its neuroprotective or neurodegenerative role remains controversial. The extent and persistence of autophagy activation may be the critical factor to explain the opposing effects. In this study, the different roles and action mechanisms of autophagy in the early and later stages after I/R injury were investigated in rats. Thespinal cord I/R injury was induced by 14-min occlusion of the aortic arch, after which rats were treated with autophagic inhibitor (3-methyladenine, 3-MA) or agonist (rapamycin) immediately or 48h following the injury. Autophagy markers, microtubule-associated protein light chain 3-II (LC3-II) and Beclin 1 increased and peaked at the early stage (8h) and the later stage (72h) after spinal cord I/R injury. Beclin 1 was mostly expressed in neurons, but was also expressed to an extent in astrocytes, microglia and vascular endothelial cells. 8h after injury, rats treated with 3-MA showed a decrease in the hind-limb Basso-Beattie-Bresnahan (BBB) motor function scores, surviving motor neurons, and B-cell lymphoma-2 (Bcl-2) expression, and increase in the terminal deoxynucleotidyl transferase (TdT)-mediated dUTP-biotin nick end labeling (TUNEL)-positive cells, Bcl-2-associated X protein (Bax), tumor necrosis factor-α (TNF-α), interleukin-1β (IL-1β) expression, and activation of microglia, while those treated with rapamycin showed opposing effects. However, 72h after injury, rats treated with 3-MA improved the BBB scores, and the surviving motor neurons, and reduced the autophagic cell death, while those treated with rapamycin had adverse effects. These findings provide the first evidence that early activated autophagy alleviates spinal cord I/R injury via inhibiting apoptosis and inflammation; however later excessively elevated autophagy aggravates I/R injury through inducing autophagic cell death. PMID:27109922

  19. Ghrelin Attenuated Lipotoxicity via Autophagy Induction and Nuclear Factor-κB Inhibition

    Directory of Open Access Journals (Sweden)

    Yuqing Mao

    2015-09-01

    Full Text Available Background/Aims: Nonalcoholic fatty liver disease (NAFLD is the most common chronic liver disease worldwide. Autophagy is associated with NAFLD. Ghrelin is a gut hormone with various functions including energy metabolism and inflammation inhibition. We investigated the therapeutic effect of ghrelin on NAFLD and its association with autophagy. Methods: C57bl/6 mice were fed a high-fat diet for 8 weeks to induce a model of chronic NAFLD, with ghrelin (10 µg/kg administrated subcutaneously twice weekly from weeks 6 to 8. LO2 cells were pretreated with ghrelin (10-8 M before stimulation with free fatty acid (palmitic and oleic acids; 1 mM. Lipid droplets were identified by hematoxylin and eosin and Red O staining and quantified by triglyceride test kits. LC3I/II, an important biomarker protein of autophagy was detected by western blotting, real-time polymerase chain reaction, immunohistochemistry and immunofluorescence. Tumor necrosis factor (TNF-a and interleukin (IL-6 were detected by ELISA and immunohistochemistry. Nuclear factor (NF-κB p65 was detected by western blotting and immunofluorescence. AMP-activated protein kinase (AMPK and mammalian target of rapamycin (mTOR were detected by western blotting. Results: Ghrelin reduced the triglyceride content in high fat diet (HFD group in vivo and free fatty acid (FFA group in vitro. TNF-a and IL-6 were significantly reduced in the ghrelin-treated mice compared with the control group. Autophagy induction was accompanied with intracellular lipid reduction in ghrelin-treated mice. Ghrelin upregulated autophagy via AMPK/mTOR restoration and inhibited translocation of NF-κB into the nucleus. Conclusions: The results indicate that ghrelin attenuates lipotoxicity by autophagy stimulation and NF-κB inhibition.

  20. Atorvastatin Protects Vascular Smooth Muscle Cells From TGF-β1-Stimulated Calcification by Inducing Autophagy via Suppression of the β-Catenin Pathway

    Directory of Open Access Journals (Sweden)

    Demin Liu

    2014-01-01

    Full Text Available Background: Arterial calcification is a major event in the progression of atherosclerosis. It is reported that statins exhibit various protective effects against vascular smooth muscle cell (VSMC inflammation and proliferation in cardiovascular remodeling. Although statins counteract atherosclerosis, the molecular mechanisms of statins on the calcium release from VSMCs have not been clearly elucidated. Methods: Calcium content of VSMCs was measured using enzyme-linked immunosorbent assay (ELISA. The expression of proteins involved in cellular transdifferentiation was analyzed by western blot. Cell autophagy was measured by fluorescence microscopic analysis for acridine orange staining and transmission electron microscopy analysis. The autophagic inhibitors (3-MA, chloroquine, NH4Cl and bafilomycin A1 and β-catenin inhibitor JW74 were used to assess the effects of atorvastatin on autophagy and the involvement of β-catenin on cell calcification respectively. Furthermore, cell transfection was performed to overexpress β-catenin. Results: In VSMCs, atorvastatin significantly suppressed transforming growth factor-β1 (TGF-β1-stimulated calcification, accompanied by the induction of autophagy. Downregulation of autophagy with autophagic inhibitors significantly suppressed the inhibitory effect of atorvastatin on cell calcification. Moreover, the beneficial effect of atorvastatin on calcification and autophagy was reversed by β-catenin overexpression. Conversely, JW74 supplement enhanced this effect. Conclusion: These data demonstrated that atorvastatin protect VSMC from TGF-β1-stimulated calcification by inducing autophagy through suppression of the β-catenin pathway, identifying autophagy induction might be a therapeutic strategy for use in vascular calcification.

  1. Coffee induces autophagy in vivo.

    Science.gov (United States)

    Pietrocola, Federico; Malik, Shoaib Ahmad; Mariño, Guillermo; Vacchelli, Erika; Senovilla, Laura; Chaba, Kariman; Niso-Santano, Mireia; Maiuri, Maria Chiara; Madeo, Frank; Kroemer, Guido

    2014-01-01

    Epidemiological studies and clinical trials revealed that chronic consumption coffee is associated with the inhibition of several metabolic diseases as well as reduction in overall and cause-specific mortality. We show that both natural and decaffeinated brands of coffee similarly rapidly trigger autophagy in mice. One to 4 h after coffee consumption, we observed an increase in autophagic flux in all investigated organs (liver, muscle, heart) in vivo, as indicated by the increased lipidation of LC3B and the reduction of the abundance of the autophagic substrate sequestosome 1 (p62/SQSTM1). These changes were accompanied by the inhibition of the enzymatic activity of mammalian target of rapamycin complex 1 (mTORC1), leading to the reduced phosphorylation of p70(S6K), as well as by the global deacetylation of cellular proteins detectable by immunoblot. Immunohistochemical analyses of transgenic mice expressing a GFP-LC3B fusion protein confirmed the coffee-induced relocation of LC3B to autophagosomes, as well as general protein deacetylation. Altogether, these results indicate that coffee triggers 2 phenomena that are also induced by nutrient depletion, namely a reduction of protein acetylation coupled to an increase in autophagy. We speculate that polyphenols contained in coffee promote health by stimulating autophagy.

  2. Coffee induces autophagy in vivo

    Science.gov (United States)

    Pietrocola, Federico; Malik, Shoaib Ahmad; Mariño, Guillermo; Vacchelli, Erika; Senovilla, Laura; Chaba, Kariman; Niso-Santano, Mireia; Maiuri, Maria Chiara; Madeo, Frank; Kroemer, Guido

    2014-01-01

    Epidemiological studies and clinical trials revealed that chronic consumption coffee is associated with the inhibition of several metabolic diseases as well as reduction in overall and cause-specific mortality. We show that both natural and decaffeinated brands of coffee similarly rapidly trigger autophagy in mice. One to 4 h after coffee consumption, we observed an increase in autophagic flux in all investigated organs (liver, muscle, heart) in vivo, as indicated by the increased lipidation of LC3B and the reduction of the abundance of the autophagic substrate sequestosome 1 (p62/SQSTM1). These changes were accompanied by the inhibition of the enzymatic activity of mammalian target of rapamycin complex 1 (mTORC1), leading to the reduced phosphorylation of p70S6K, as well as by the global deacetylation of cellular proteins detectable by immunoblot. Immunohistochemical analyses of transgenic mice expressing a GFP–LC3B fusion protein confirmed the coffee-induced relocation of LC3B to autophagosomes, as well as general protein deacetylation. Altogether, these results indicate that coffee triggers 2 phenomena that are also induced by nutrient depletion, namely a reduction of protein acetylation coupled to an increase in autophagy. We speculate that polyphenols contained in coffee promote health by stimulating autophagy. PMID:24769862

  3. Characterization of the autophagy marker protein Atg8 reveals atypical features of autophagy in Plasmodium falciparum.

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    Rahul Navale

    Full Text Available Conventional autophagy is a lysosome-dependent degradation process that has crucial homeostatic and regulatory functions in eukaryotic organisms. As malaria parasites must dispose a number of self and host cellular contents, we investigated if autophagy in malaria parasites is similar to the conventional autophagy. Genome wide analysis revealed a partial autophagy repertoire in Plasmodium, as homologs for only 15 of the 33 yeast autophagy proteins could be identified, including the autophagy marker Atg8. To gain insights into autophagy in malaria parasites, we investigated Plasmodium falciparum Atg8 (PfAtg8 employing techniques and conditions that are routinely used to study autophagy. Atg8 was similarly expressed and showed punctate localization throughout the parasite in both asexual and sexual stages; it was exclusively found in the pellet fraction as an integral membrane protein, which is in contrast to the yeast or mammalian Atg8 that is distributed among cytosolic and membrane fractions, and suggests for a constitutive autophagy. Starvation, the best known autophagy inducer, decreased PfAtg8 level by almost 3-fold compared to the normally growing parasites. Neither the Atg8-associated puncta nor the Atg8 expression level was significantly altered by treatment of parasites with routinely used autophagy inhibitors (cysteine (E64 and aspartic (pepstatin protease inhibitors, the kinase inhibitor 3-methyladenine, and the lysosomotropic agent chloroquine, indicating an atypical feature of autophagy. Furthermore, prolonged inhibition of the major food vacuole protease activity by E64 and pepstatin did not cause accumulation of the Atg8-associated puncta in the food vacuole, suggesting that autophagy is primarily not meant for degradative function in malaria parasites. Atg8 showed partial colocalization with the apicoplast; doxycycline treatment, which disrupts apicoplast, did not affect Atg8 localization, suggesting a role, but not exclusive, in

  4. Cytochrome P450 3A Conjugation to Ubiquitin in a Process Distinct from Classical Ubiquitination Pathway

    Energy Technology Data Exchange (ETDEWEB)

    Zangar, Richard C.(BATTELLE (PACIFIC NW LAB)); Kimzey, Amy L.(ASSOC WESTERN UNIVERSITY); Okita, Janice R.(Washington State University); Wunschel, David S.(BATTELLE (PACIFIC NW LAB)); Edwards, Robert J.(Imperial College School of Medical, Hammersmith Campus); Kim, Hyesook (Wayne State University); Okita, Richard T.(Washington State University)

    2001-12-01

    We characterize a novel microsome system that forms high-molecular-mass (HMM) CYP3A, CYP2E1, and ubiquitin conjugates, but does not alter CYP4A or most other microsomal proteins. The formation of the HMM bands was observed in hepatic microsomes isolated from rats treated 1 week or more with high doses (50 mg/kg/day) of nicardipine, clotrimazole, or pregnenolone 16alpha-carbonitrile, but not microsomes from control, dexamethasone-, nifedipine-, or diltiazem-treated rats. Extensive washing of the microsomes to remove loosely attached proteins or cytosolic contaminants did not prevent the conjugation reaction. In contrast to prototypical ubiquitination pathways, this reaction did not require addition of ubiquitin, ATP, Mg(2+), or cytosol. Addition of cytosol did result in the degradation of the HMM CYP3A bands in a process that was not blocked by proteasome inhibitors. Immunoprecipitated CYP3A contained HMM ubiquitin. Even so, mass spectrometric analysis of tryptic peptides indicated that the HMM CYP3A was in molar excess to ubiquitin, suggesting that the formation of the HMM CYP3A may have resulted from conjugation to itself or a diffuse pool of ubiquitinated proteins already present in the microsomes. Addition of CYP3A substrates inhibited the formation of the HMM CYP3A and the cytosol-dependent degradation of HMM CYP3A. These results suggest that after extended periods of elevated CYP3A expression, microsomal factors are induced that catalyze the formation of HMM CYP3A conjugates that contain ubiquitin. This conjugation reaction, however, seems to be distinct from the classical ubiquitination pathway but may be related to the substrate-dependent stabilization of CYP3A observed in vivo.

  5. Autophagy facilitates secretion and protects against degeneration of the Harderian gland.

    Science.gov (United States)

    Koenig, Ulrich; Fobker, Manfred; Lengauer, Barbara; Brandstetter, Marlene; Resch, Guenter P; Gröger, Marion; Plenz, Gabriele; Pammer, Johannes; Barresi, Caterina; Hartmann, Christine; Rossiter, Heidemarie

    2015-01-01

    The epithelial derived Harderian gland consists of 2 types of secretory cells. The more numerous type A cells are responsible for the secretion of lipid droplets, while type B cells produce dark granules of multilamellar bodies. The process of autophagy is constitutively active in the Harderian gland, as confirmed by our analysis of LC3 processing in GFP-LC3 transgenic mice. This process is compromised by epithelial deletion of Atg7. Morphologically, the Atg7 mutant glands are hypotrophic and degenerated, with highly vacuolated cells and pyknotic nuclei. The mutant glands accumulate lipid droplets coated with PLIN2 (perilipin 2) and contain deposits of cholesterol, ubiquitinated proteins, SQSTM1/p62 (sequestosome 1) positive aggregates and other metabolic products such as porphyrin. Immunofluorescence stainings show that distinct cells strongly aggregate both proteins and lipids. Electron microscopy of the Harderian glands reveals that its organized structure is compromised, and the presence of large intracellular lipid droplets and heterologous aggregates. We attribute the occurrence of large vacuoles to a malfunction in the formation of multilamellar bodies found in the less abundant type B Harderian gland cells. This defect causes the formation of large tertiary lysosomes of heterologous content and is accompanied by the generation of tight lamellar stacks of endoplasmic reticulum in a pseudo-crystalline form. To test the hypothesis that lipid and protein accumulation is the cause for the degeneration in autophagy-deficient Harderian glands, epithelial cells were treated with a combination of the proteasome inhibitor and free fatty acids, to induce aggregation of misfolded proteins and lipid accumulation, respectively. The results show that lipid accumulation indeed enhanced the toxicity of misfolded proteins and that this was even more pronounced in autophagy-deficient cells. Thus, we conclude autophagy controls protein and lipid catabolism and anabolism to

  6. Autophagy- An emerging target for melanoma therapy

    Science.gov (United States)

    Ndoye, Abibatou; Weeraratna, Ashani T.

    2016-01-01

    Melanoma accounts for only 5% of all cancers but is the leading cause of skin cancer death due to its high metastatic potential. Patients with metastatic melanoma have a 10-year survival rate of less than 10%. While the clinical landscape for melanoma is evolving rapidly, lack of response to therapies, as well as resistance to therapy remain critical obstacles for treatment of this disease. In recent years, a myriad of therapy resistance mechanisms have been unravelled, one of which is autophagy, the focus of this review. In advanced stages of malignancy, melanoma cells hijack the autophagy machinery in order to alleviate drug-induced and metabolic stress in the tumor microenvironment, thereby promoting resistance to multiple therapies, tumor cell survival, and progression.  Autophagy is an essential cellular process that maintains cellular homeostasis through the recycling of intracellular constituents. Early studies on the role of autophagy in cancer generated controversy as to whether autophagy was pro- or anti-tumorigenic. Currently, there is a consensus that autophagy is tumor-suppressive in the early stages of cancer and tumor-promoting in established tumors.  This review aims to highlight current understandings on the role of autophagy in melanoma malignancy, and specifically therapy resistance; as well as to evaluate recent strategies for therapeutic autophagy modulation. PMID:27583134

  7. Autophagy in allografts rejection: A new direction?

    Science.gov (United States)

    Sun, Hukui; Cheng, Dayan; Ma, Yuanyuan; Wang, Huaiquan; Liang, Ting; Hou, Guihua

    2016-03-18

    Despite the introduction of new and effective immunosuppressive drugs, acute cellular graft rejection is still a major risk for graft survival. Modulating the dosage of immunosuppressive drugs is not a good choice for all patients, new rejection mechanisms discovery are crucial to limit the inflammatory process and preserve the function of the transplant. Autophagy, a fundamental cellular process, can be detected in all subsets of lymphocytes and freshly isolated naive T lymphocytes. It is required for the homeostasis and function of T lymphocytes, which lead to cell survival or cell death depending on the context. T cell receptor (TCR) stimulation and costimulator signals induce strong autophagy, and autophagy deficient T cells leads to rampant apoptosis upon TCR stimulation. Autophagy has been proved to be activated during ischemia-reperfusion (I/R) injury and associated with grafts dysfunction. Furthermore, Autophagy has also emerged as a key mechanism in orchestrating innate and adaptive immune response to self-antigens, which relates with negative selection and Foxp3(+) Treg induction. Although, the role of autophagy in allograft rejection is unknown, current data suggest that autophagy indeed sweeps across both in the graft organs and recipients lymphocytes after transplantation. This review presents the rationale for the hypothesis that targeting the autophagy pathway could be beneficial in promoting graft survival after transplantation.

  8. Tumor Suppression and Promotion by Autophagy

    Directory of Open Access Journals (Sweden)

    Yenniffer Ávalos

    2014-01-01

    Full Text Available Autophagy is a highly regulated catabolic process that involves lysosomal degradation of proteins and organelles, mostly mitochondria, for the maintenance of cellular homeostasis and reduction of metabolic stress. Problems in the execution of this process are linked to different pathological conditions, such as neurodegeneration, aging, and cancer. Many of the proteins that regulate autophagy are either oncogenes or tumor suppressor proteins. Specifically, tumor suppressor genes that negatively regulate mTOR, such as PTEN, AMPK, LKB1, and TSC1/2 stimulate autophagy while, conversely, oncogenes that activate mTOR, such as class I PI3K, Ras, Rheb, and AKT, inhibit autophagy, suggesting that autophagy is a tumor suppressor mechanism. Consistent with this hypothesis, the inhibition of autophagy promotes oxidative stress, genomic instability, and tumorigenesis. Nevertheless, autophagy also functions as a cytoprotective mechanism under stress conditions, including hypoxia and nutrient starvation, that promotes tumor growth and resistance to chemotherapy in established tumors. Here, in this brief review, we will focus the discussion on this ambiguous role of autophagy in the development and progression of cancer.

  9. Keeping autophagy in cheCK1

    Science.gov (United States)

    Cheong, Jit Kong; Virshup, David M.

    2016-01-01

    Abstract Mutant RAS-driven cancer cells cope with proliferative stress by increasing basal autophagy to maintain protein/organelle and energy homeostasis. We recently demonstrated that casein kinase 1 alpha (CK1α), a therapeutically tractable enzyme, is critical for fine-tuning the transcriptional regulation of mutant RAS-induced autophagy and the development of mutant RAS-driven cancers. PMID:27314070

  10. Autophagy- An emerging target for melanoma therapy.

    Science.gov (United States)

    Ndoye, Abibatou; Weeraratna, Ashani T

    2016-01-01

    Melanoma accounts for only 5% of all cancers but is the leading cause of skin cancer death due to its high metastatic potential. Patients with metastatic melanoma have a 10-year survival rate of less than 10%. While the clinical landscape for melanoma is evolving rapidly, lack of response to therapies, as well as resistance to therapy remain critical obstacles for treatment of this disease. In recent years, a myriad of therapy resistance mechanisms have been unravelled, one of which is autophagy, the focus of this review. In advanced stages of malignancy, melanoma cells hijack the autophagy machinery in order to alleviate drug-induced and metabolic stress in the tumor microenvironment, thereby promoting resistance to multiple therapies, tumor cell survival, and progression.  Autophagy is an essential cellular process that maintains cellular homeostasis through the recycling of intracellular constituents. Early studies on the role of autophagy in cancer generated controversy as to whether autophagy was pro- or anti-tumorigenic. Currently, there is a consensus that autophagy is tumor-suppressive in the early stages of cancer and tumor-promoting in established tumors.  This review aims to highlight current understandings on the role of autophagy in melanoma malignancy, and specifically therapy resistance; as well as to evaluate recent strategies for therapeutic autophagy modulation. PMID:27583134

  11. Wwp2, an E3 Ubiquitin Ligase That Targets Transcription Factor Oct-4 for Ubiquitination

    Institute of Scientific and Technical Information of China (English)

    HuiMingXu; BingLiao; QianJunZhang; BeiBeiWang; Hui,Li; XiaoMinZhong; HuiZhenSheng; YingXinZhao; YingMingZhao; YingJin

    2005-01-01

    The POU transcription factor Oct-4 is a master regulator affecting the fate of pluripotent embryonic stem cells. However, the precise mechanisms by which the activation and expression of Oct-4 are regulated still remain to be elucidated. We describe here a novel murine ubiquitin ligase, Wwp2, that specifically interacts with Oct-4 and promotes its ubiquitination both in vivo and in vitro. Remarkably, the expression of a catalytically inactive point mutant of Wwp2 abolishes Oct-4 ubiquitination. Moreover, Wwp2 promotes Oct-4 degradation in the presence of overexpressed ubiquitin. The degradation is blocked by treatment with proteasome inhibitor. Fusion of a single ubiquitin to Oct-4 inactivates its transcriptional activity in a heterologous Oct-4-driven reporter system. Furthermore, overexpression of Wwp2 in embryonic stem cells significantly reduces the Oct-4-transcriptional activities. Collectively, we demonstrate for the first time that Oct-4 can be posttranslationatly modified by ubiquitination and that this modification dramatically suppresses its transcriptional activity. These results reveal that the functional status of Oct-4, in addition to its expression level, dictates its transcriptional activity, and the results open up a new avenue to understand how Oct-4 defines the fate of embryonic stem cells.

  12. Emerging connections between RNA and autophagy

    DEFF Research Database (Denmark)

    Frankel, Lisa B; Lubas, Michal; Lund, Anders H

    2016-01-01

    Macroautophagy/autophagy is a key catabolic process, essential for maintaining cellular homeostasis and survival through the removal and recycling of unwanted cellular material. Emerging evidence has revealed intricate connections between the RNA and autophagy research fields. While a majority...... of studies have focused on protein, lipid and carbohydrate catabolism via autophagy, accumulating data supports the view that several types of RNA and associated ribonucleoprotein complexes are specifically recruited to phagophores (precursors to autophagosomes) and subsequently degraded in the lysosome....../vacuole. Moreover, recent studies have revealed a substantial number of novel autophagy regulators with RNA-related functions, indicating roles for RNA and associated proteins not only as cargo, but also as regulators of this process. In this review, we discuss widespread evidence of RNA catabolism via autophagy...

  13. Stress management by autophagy: Implications for chemoresistance.

    Science.gov (United States)

    Huang, Zhao; Zhou, Li; Chen, Zhibin; Nice, Edouard C; Huang, Canhua

    2016-07-01

    Development of chemoresistance, which limits the efficiency of anticancer agents, has long been a major problem in cancer therapy and urgently needs to be solved to improve clinical outcomes. Factors contributing to chemoresistance are various, but a key factor is the cell's capability for stress management. Autophagy, a favored survival strategy that organisms employ to get over many kinds of stress, is emerging as a crucial player in drug resistance. It has been shown that autophagy facilitates the resistance of tumor cells to anticancer agents, and abrogation of autophagy could be therapeutically beneficial in some cases, suggesting autophagy could be a promising target for cancer treatments. Thus, defining the roles of autophagy in chemoresistance, and the mechanisms involved, will be critical to enhance the efficiency of chemotherapy and develop novel anticancer strategy interventions.

  14. The putative role of proteolytic pathways in the pathogenesis of Type 1 diabetes mellitus: the 'autophagy' hypothesis.

    Science.gov (United States)

    Fierabracci, Alessandra

    2014-05-01

    Autoimmune diseases are a heterogeneous group of disorders affecting different organs and tissues. New tools, such as genome-wide association studies, have provided evidence for new susceptibility loci and candidate genes in the disease process including common susceptibility genes involved in the immunological synapse and T cell activation. Close linkages have been found in a number of diseases, including ankylosing spondylitis, multiple sclerosis, Crohn's disease and insulin-dependent diabetes mellitus (Type 1 diabetes mellitus). Evidence for some association with Type 1 diabetes was previously found in the region containing 5q15/ERAP1 (endoplasmic reticulum aminopeptidase 1) (rs30187, ARTS1). Recent data suggest that in eukaryotic cells in addition to the ubiquitin/proteasome system another proteolytic pathway may have a significant role in the autoimmunity process, i.e. the autophagic pathway which constitutes the principal regulated catabolic process mediated by lysosomes. Autophagy could play a role in MHC class I and class II self-antigen presentation at the basis of the autoimmunity process. Furthermore cross-talk among different proteolytic pathways was recently highlighted i.e. components processed in the ubiquitin/proteasome system possibly engaged in autophagic pathways. T1D is an autoimmune disease characterised by the destruction of pancreatic beta cells by autoreactive T cells. Immunological abnormalities can precede months to years the initial symptoms and clinical diagnosis. Our hypothesis suggests that in the autoimmune process autophagy can intervene at different levels, during the thymic selection process of T lymphocytes causing escape of autoreactive T cells, at the initiation stage of the disease, in the preclinical period or subsequently to the disease onset having a role at the level of perpetuation of the autoimmunity process. Supporting evidence derives from the already reported discovery of polymorphisms in autophagy-related genes in

  15. Cooperativity of the SUMO and Ubiquitin Pathways in Genome Stability

    Directory of Open Access Journals (Sweden)

    Minghua Nie

    2016-02-01

    Full Text Available Covalent attachment of ubiquitin (Ub or SUMO to DNA repair proteins plays critical roles in maintaining genome stability. These structurally related polypeptides can be viewed as distinct road signs, with each being read by specific protein interaction motifs. Therefore, via their interactions with selective readers in the proteome, ubiquitin and SUMO can elicit distinct cellular responses, such as directing DNA lesions into different repair pathways. On the other hand, through the action of the SUMO-targeted ubiquitin ligase (STUbL family proteins, ubiquitin and SUMO can cooperate in the form of a hybrid signal. These mixed SUMO-ubiquitin chains recruit “effector” proteins such as the AAA+ ATPase Cdc48/p97-Ufd1-Npl4 complex that contain both ubiquitin and SUMO interaction motifs. This review will summarize recent key findings on collaborative and distinct roles that ubiquitin and SUMO play in orchestrating DNA damage responses.

  16. The ubiquitin proteasome system and efficacy of proteasome inhibitors in diseases.

    Science.gov (United States)

    Chitra, Selvarajan; Nalini, Ganesan; Rajasekhar, Gopalakrishnan

    2012-06-01

    In eukaryotes the ubiquitin proteasome pathway plays an important role in cellular homeostasis and also it exerts a critical role in regulating a wide variety of cellular pathways, including cell growth and proliferation, apoptosis, DNA repair, transcription and immune response. Defects in these pathways have been implicated in a number of human pathologies. Inhibition of the ubiquitin proteasome pathway by proteasome inhibitors may be a rational therapeutic approach for various diseases, such as cancer and inflammatory diseases. Many of the critical cytokine and chemokine mediators of the progression of rheumatoid arthritis are regulated by nuclear factor kappa B (NF-κB). In peptidoglycan/polysaccharide-induced polyarthritis, proteasome inhibitors limit the overall inflammation, reduce NF-κB activation, decrease cellular adhesion molecule expression, inhibit nitric oxide synthase, attenuate circulating levels of proinflammatory cytokine interleukin-6 and reduce the arthritis index and swelling in the joints of the animals. Since proteasome inhibitors exhibit anti-inflammatory and anti proliferative effects, diseases characterized by both of these processes such as rheumatoid arthritis might also represent clinical opportunities for such drugs. The regulation of the proteasomal complex by proteasome inhibitors also has implications and potential benefits for the treatment of rheumatoid arthritis. This review summarizes the ubiquitin proteasome pathway, the structure of 26S proteasomes and types of proteasome inhibitors, with their actions, and clinical applications of proteasome inhibitors in various diseases. PMID:22709487

  17. Ubiquitin reference technique and its use in ubiquitin-lacking prokaryotes.

    Directory of Open Access Journals (Sweden)

    Konstantin Piatkov

    Full Text Available In a pulse-chase assay, the in vivo degradation of a protein is measured through a brief labeling of cells with, for example, a radioactive amino acid, followed by cessation of labeling and analysis of cell extracts prepared at different times afterward ("chase", using immunoprecipitation, electrophoresis and autoradiography of a labeled protein of interest. A conventional pulse-chase assay is fraught with sources of data scatter, as the efficacy of labeling and immunoprecipitation can vary, and sample volumes can vary as well. The ubiquitin reference technique (URT, introduced in 1996, addresses these problems. In eukaryotes, a DNA-encoded linear fusion of ubiquitin to another protein is cleaved by deubiquitylases at the ubiquitin-protein junction. A URT assay uses a fusion in which the ubiquitin moiety is located between a downstream polypeptide (test protein and an upstream polypeptide (a long-lived reference protein. The cotranslational cleavage of a URT fusion by deubiquitylases after the last residue of ubiquitin produces, at the initially equimolar ratio, a test protein with a desired N-terminal residue and a reference protein containing C-terminal ubiquitin moiety. In addition to being more accurate than pulse-chases without a reference, URT makes it possible to detect and measure the degradation of a test protein during the pulse (before the chase. Because prokaryotes, including Gram-negative bacteria such as, for example, Escherichia coli and Vibrio vulnificus, lack the ubiquitin system, the use of URT in such cells requires ectopic expression of a deubiquitylase. We describe designs and applications of plasmid vectors that coexpress, in bacteria, both a URT-type fusion and Ubp1, a deubiquitylase of the yeast Saccharomyces cerevisiae. This single-plasmid approach extends the accuracy-enhancing URT assay to studies of protein degradation in prokaryotes.

  18. Radiation inhibits proteasomes and increases ubiquitinated proteins

    International Nuclear Information System (INIS)

    Full text: Exposure of cells to ionizing radiation results in accumulation of a number of short lived proteins that mediate cell survival/death, proliferation, repair, and differentiation. Expression of most of these proteins, including p53, mdm2, p21, c-jun, IkB-a, bcl-2, bax, cyclins A, B, E, Cdc25A, DNA-PKcs, and caspase-3 is regulated at the post-transcriptional level through ubiquitin/26S proteasome pathway. Several previous studies have shown that inhibition of proteasome activity by drugs leads to accumulation of ubiquitinated proteins. In this study we show that irradiation can do the same due to its inhibitory effect on 26S, but not 20S, proteasome activity. Two prostate cancer cell lines, murine TRAMP-C1 and human PC3, were used to examine the effect of ionizing radiation on the catalytic activity of the 26S proteasome. Cells were irradiated with different doses ranging from 0.25 to 20 Gy and lysed at different time points after irradiation. Crude extracts of both cell lines showed a rapid 30-50% decrease in chymotryptic activity of the 26S proteasome, as measured by a fluorogenic assay. The same level of inhibition was observed if purified 26S proteasomes were themselves irradiated, indicating that radiation has direct effects on this multicatalytic enzyme complex. Neither direct irradiation of proteasomes or cells had effect on 20S catalytic activity, suggesting that radiation selectively acts on 26S structure. Next, we examined whether this partial inhibition had any effect on ability of 26S proteasome to efficiently remove ubiquitinated proteins. Cells were irradiated with 10Gy and lysed at different time points. Ubiquitinated proteins were precipitated and examined by Western blot. Levels of ubiquitinated conjugates slowly increased over time and peaked at 7h post-irradiation. Accumulation of ubiquitinated conjugates has been shown to lead to formation of protein aggregates which can induce cell death. It has also been shown that monoubiquitination

  19. Autophagy in granular corneal dystrophy type 2.

    Science.gov (United States)

    Choi, Seung-Il; Kim, Eung Kweon

    2016-03-01

    Autophagy is a lysosomal degradative process that is essential for cellular homeostasis and metabolic stress adaptation. Defective autophagy is involved in the pathogenesis of many diseases including granular corneal dystrophy type 2 (GCD2). GCD2 is an autosomal dominant disorder caused by substitution of histidine for arginine at codon 124 (R124H) in the transforming growth factor β-induced gene (TGFBI) on chromosome 5q31. Transforming growth factor β-induced protein (TGFBIp) is degraded by autophagy, but mutant-TGFBIp accumulates in autophagosomes and/or lysosomes, despite significant activation of basal autophagy, in GCD2 corneal fibroblasts. Furthermore, inhibition of autophagy induces cell death of GCD2 corneal fibroblasts through active caspase-3. As there is currently no pharmacological treatment for GCD2, development of novel therapies is required. A potential strategy for preventing cytoplasmic accumulation of mutant-TGFBIp in GCD2 corneal fibroblasts is to enhance mutant-TGFBIp degradation. This could be achieved by activation of the autophagic pathway. Here, we will consider the role and the potential therapeutic benefits of autophagy in GCD2, with focus on TGFBIp degradation, in light of the recently established role of autophagy in protein degradation.

  20. Autophagy: for better or for worse

    Institute of Scientific and Technical Information of China (English)

    Ellen Wirawan; Tom Vanden Berghe; Saskia Lippens; Patrizia Agostinis; Peter Vandenabeele

    2012-01-01

    Autophagy is a lysosomal degradation pathway that degrades damaged or superfluous cell components into basic biomolecules,which are then recycled back into the cytosol.In this respect,autophagy drives a flow of biomolecules in a continuous degradation-regeneration cycle.Autophagy is generally considered a pro-survival mechanism protecting cells under stress or poor nutrient conditions.Current research clearly shows that autophagy fulfills numerous functions in vital biological processes.It is implicated in development,differentiation,innate and adaptive immunity,ageing and cell death.In addition,accumulating evidence demonstrates interesting links between autophagy and several human diseases and tumor development.Therefore,autophagy seems to be an important player in the life and death of cells and organisms.Despite the mounting knowledge about autophagy,the mechanisms through which the autophagic machinery regulates these diverse processes are not entirely understood.In this review,we give a comprehensive overview of the autophagic signaling pathway,its role in general cellular processes and its connection to cell death.In addition,we present a brief overview of the possible contribution of defective autophagic signaling to disease.

  1. Guidelines for monitoring autophagy in Caenorhabditis elegans.

    Science.gov (United States)

    Zhang, Hong; Chang, Jessica T; Guo, Bin; Hansen, Malene; Jia, Kailiang; Kovács, Attila L; Kumsta, Caroline; Lapierre, Louis R; Legouis, Renaud; Lin, Long; Lu, Qun; Meléndez, Alicia; O'Rourke, Eyleen J; Sato, Ken; Sato, Miyuki; Wang, Xiaochen; Wu, Fan

    2015-01-01

    The cellular recycling process of autophagy has been extensively characterized with standard assays in yeast and mammalian cell lines. In multicellular organisms, numerous external and internal factors differentially affect autophagy activity in specific cell types throughout the stages of organismal ontogeny, adding complexity to the analysis of autophagy in these metazoans. Here we summarize currently available assays for monitoring the autophagic process in the nematode C. elegans. A combination of measuring levels of the lipidated Atg8 ortholog LGG-1, degradation of well-characterized autophagic substrates such as germline P granule components and the SQSTM1/p62 ortholog SQST-1, expression of autophagic genes and electron microscopy analysis of autophagic structures are presently the most informative, yet steady-state, approaches available to assess autophagy levels in C. elegans. We also review how altered autophagy activity affects a variety of biological processes in C. elegans such as L1 survival under starvation conditions, dauer formation, aging, and cell death, as well as neuronal cell specification. Taken together, C. elegans is emerging as a powerful model organism to monitor autophagy while evaluating important physiological roles for autophagy in key developmental events as well as during adulthood.

  2. Autophagy in human embryonic stem cells.

    Directory of Open Access Journals (Sweden)

    Thien Tra

    Full Text Available Autophagy (macroautophagy is a degradative process that involves the sequestration of cytosolic material including organelles into double membrane vesicles termed autophagosomes for delivery to the lysosome. Autophagy is essential for preimplantation development of mouse embryos and cavitation of embryoid bodies. The precise roles of autophagy during early human embryonic development, remain however largely uncharacterized. Since human embryonic stem cells constitute a unique model system to study early human embryogenesis we investigated the occurrence of autophagy in human embryonic stem cells. We have, using lentiviral transduction, established multiple human embryonic stem cell lines that stably express GFP-LC3, a fluorescent marker for the autophagosome. Each cell line displays both a normal karyotype and pluripotency as indicated by the presence of cell types representative of the three germlayers in derived teratomas. GFP expression and labelling of autophagosomes is retained after differentiation. Baseline levels of autophagy detected in cultured undifferentiated hESC were increased or decreased in the presence of rapamycin and wortmannin, respectively. Interestingly, autophagy was upregulated in hESCs induced to undergo differentiation by treatment with type I TGF-beta receptor inhibitor SB431542 or removal of MEF secreted maintenance factors. In conclusion we have established hESCs capable of reporting macroautophagy and identify a novel link between autophagy and early differentiation events in hESC.

  3. Autophagy in Mycobacterium tuberculosis and HIV infections

    Directory of Open Access Journals (Sweden)

    Lucile eEspert

    2015-06-01

    Full Text Available Human Immunodeficiency Virus (HIV and Mycobacterium tuberculosis (M.tb are among the most lethal human pathogens worldwide, each being responsible for around 1.5 million deaths annually. Moreover, synergy between acquired immune deficiency syndrome (AIDS and tuberculosis (TB has turned HIV/M.tb co-infection into a major public health threat in developing countries. In the past decade, autophagy, a lysosomal catabolic process, has emerged as a major host immune defense mechanism against infectious agents like M.tb and HIV. Nevertheless, in some instances, autophagy machinery appears to be instrumental for HIV infection. Finally, there is mounting evidence that both pathogens deploy various countermeasures to thwart autophagy. This mini-review proposes an overview of the roles and regulations of autophagy in HIV and M.tb infections with an emphasis on microbial factors. We also discuss the role of autophagy manipulation in the context of HIV/M.tb co-infection. In future, a comprehensive understanding of autophagy interaction with these pathogens will be critical for development of autophagy-based prophylactic and therapeutic interventions for AIDS and TB.

  4. The Ubiquitin System and Jasmonate Signaling

    Directory of Open Access Journals (Sweden)

    Astrid Nagels Durand

    2016-01-01

    Full Text Available The ubiquitin (Ub system is involved in most, if not all, biological processes in eukaryotes. The major specificity determinants of this system are the E3 ligases, which bind and ubiquitinate specific sets of proteins and are thereby responsible for target recruitment to the proteasome or other cellular processing machineries. The Ub system contributes to the regulation of the production, perception and signal transduction of plant hormones. Jasmonic acid (JA and its derivatives, known as jasmonates (JAs, act as signaling compounds regulating plant development and plant responses to various biotic and abiotic stress conditions. We provide here an overview of the current understanding of the Ub system involved in JA signaling.

  5. A novel effect of thalidomide and its analogs: suppression of cereblon ubiquitination enhances ubiquitin ligase function.

    Science.gov (United States)

    Liu, Yaobin; Huang, Xiangao; He, Xian; Zhou, Yanqing; Jiang, Xiaogang; Chen-Kiang, Selina; Jaffrey, Samie R; Xu, Guoqiang

    2015-12-01

    The immunomodulatory drug (IMiD) thalidomide and its structural analogs lenalidomide and pomalidomide are highly effective in treating clinical indications. Thalidomide binds to cereblon (CRBN), a substrate receptor of the cullin-4 really interesting new gene (RING) E3 ligase complex. Here, we examine the effect of thalidomide and its analogs on CRBN ubiquitination and its functions in human cell lines. We find that the ubiquitin modification of CRBN includes K48-linked polyubiquitin chains and that thalidomide blocks the formation of CRBN-ubiquitin conjugates. Furthermore, we show that ubiquitinated CRBN is targeted for proteasomal degradation. Treatment of human myeloma cell lines such as MM1.S, OPM2, and U266 with thalidomide (100 μM) and its structural analog lenalidomide (10 μM) results in stabilization of CRBN and elevation of CRBN protein levels. This in turn leads to the reduced level of CRBN target proteins and enhances the sensitivity of human multiple myeloma cells to IMiDs. Our results reveal a novel mechanism by which thalidomide and its analogs modulate the CRBN function in cells. Through inhibition of CRBN ubiquitination, thalidomide and its analogs allow CRBN to accumulate, leading to the increased cullin-4 RING E3 ligase-mediated degradation of target proteins.

  6. Inflammation and Heart Disease

    Science.gov (United States)

    ... Pressure High Blood Pressure Tools & Resources Stroke More Inflammation and Heart Disease Updated:Apr 18,2016 Understand the risks of inflammation. Although it is not proven that inflammation causes ...

  7. The increasing complexity of the ubiquitin code.

    Science.gov (United States)

    Yau, Richard; Rape, Michael

    2016-05-27

    Ubiquitylation is essential for signal transduction as well as cell division and differentiation in all eukaryotes. Substrate modifications range from a single ubiquitin molecule to complex polymeric chains, with different types of ubiquitylation often eliciting distinct outcomes. The recent identification of novel chain topologies has improved our understanding of how ubiquitylation establishes precise communication within cells. Here, we discuss how the increasing complexity of ubiquitylation is employed to ensure robust and faithful signal transduction in eukaryotic cells. PMID:27230526

  8. Degradation of Activated Protein Kinases by Ubiquitination

    OpenAIRE

    Lu, Zhimin; Hunter, Tony

    2009-01-01

    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.

  9. Effect of Autophagy Over Liver Diseases

    Institute of Scientific and Technical Information of China (English)

    Dong-qian Yi; Xue-feng Yang; Duan-fang Liao; Qing Wu; Nian Fu; Yang Hu; Ting Cao

    2016-01-01

    Abstract In recent years, increasingly evidences show that autophagy plays an important role in the pathogenesis and development of liver diseases, and the relationship between them has increasingly become a focus of concern. Autophagy refers to the process through which the impaired organelles, misfolded protein, and intruding microorganisms is degraded by lysosomes to maintain stability inside cells. This article states the effect of autophagy on liver diseases (hepatic fibrosis, fatty liver, viral hepatitis, and liver cancer), which aims to provide a new direction for the treatment of liver diseases.

  10. Autophagy mitigates metabolic stress and genome damage in mammary tumorigenesis

    Science.gov (United States)

    Karantza-Wadsworth, Vassiliki; Patel, Shyam; Kravchuk, Olga; Chen, Guanghua; Mathew, Robin; Jin, Shengkan; White, Eileen

    2007-01-01

    Autophagy is a catabolic process involving self-digestion of cellular organelles during starvation as a means of cell survival; however, if it proceeds to completion, autophagy can lead to cell death. Autophagy is also a haploinsufficient tumor suppressor mechanism for mammary tumorigenesis, as the essential autophagy regulator beclin1 is monoallelically deleted in breast carcinomas. However, the mechanism by which autophagy suppresses breast cancer remains elusive. Here we show that allelic loss of beclin1 and defective autophagy sensitized mammary epithelial cells to metabolic stress and accelerated lumen formation in mammary acini. Autophagy defects also activated the DNA damage response in vitro and in mammary tumors in vivo, promoted gene amplification, and synergized with defective apoptosis to promote mammary tumorigenesis. Therefore, we propose that autophagy limits metabolic stress to protect the genome, and that defective autophagy increases DNA damage and genomic instability that ultimately facilitate breast cancer progression. PMID:17606641

  11. Oxidative stress-induced autophagy: Role in pulmonary toxicity

    Energy Technology Data Exchange (ETDEWEB)

    Malaviya, Rama [Department of Pharmacology and Toxicology, Ernest Mario School of Pharmacy, Rutgers University, Piscataway, NJ 08854 (United States); Laskin, Jeffrey D. [Department of Environmental and Occupational Medicine, Robert Wood Johnson Medical School, Rutgers University, Piscataway, NJ 08854 (United States); Laskin, Debra L., E-mail: laskin@eohsi.rutgers.edu [Department of Pharmacology and Toxicology, Ernest Mario School of Pharmacy, Rutgers University, Piscataway, NJ 08854 (United States)

    2014-03-01

    Autophagy is an evolutionarily conserved catabolic process important in regulating the turnover of essential proteins and in elimination of damaged organelles and protein aggregates. Autophagy is observed in the lung in response to oxidative stress generated as a consequence of exposure to environmental toxicants. Whether autophagy plays role in promoting cell survival or cytotoxicity is unclear. In this article recent findings on oxidative stress-induced autophagy in the lung are reviewed; potential mechanisms initiating autophagy are also discussed. A better understanding of autophagy and its role in pulmonary toxicity may lead to the development of new strategies to treat lung injury associated with oxidative stress. - Highlights: • Exposure to pulmonary toxicants is associated with oxidative stress. • Oxidative stress is known to induce autophagy. • Autophagy is upregulated in the lung following exposure to pulmonary toxicants. • Autophagy may be protective or pathogenic.

  12. Histone deacetylase 2 is phosphorylated, ubiquitinated, and degraded by cigarette smoke.

    Science.gov (United States)

    Adenuga, David; Yao, Hongwei; March, Thomas H; Seagrave, Jeanclare; Rahman, Irfan

    2009-04-01

    Cigarette smoke (CS)-induced lung inflammation involves the reduction of histone deacetylase 2 (HDAC2) abundance, which is associated with steroid resistance in patients with chronic obstructive pulmonary disease and in individuals with severe asthma who smoke cigarettes. However, the molecular mechanism of CS-mediated reduction of HDAC2 is not clearly known. We hypothesized that HDAC2 is phosphorylated and subsequently degraded by the proteasome in vitro in macrophages (MonoMac6), human bronchial and primary small airway epithelial cells, and in vivo in mouse lungs in response to chronic CS exposure. Cigarette smoke extract (CSE) exposure in MonoMac6 and in bronchial and airway epithelial cells led to phosphorylation of HDAC2 on serine/threonine residues by a protein kinase CK2-mediated mechanism, decreased HDAC2 activity, and increased ubiquitin-proteasome-dependent HDAC2 degradation. CK2 and proteasome inhibitors reversed CSE-mediated HDAC2 degradation, whereas serine/threonine phosphatase inhibitor, okadaic acid, caused phosphorylation and subsequent ubiquitination of HDAC2. CS-induced HDAC2 phosphorylation was detected in mouse lungs from 2 weeks to 4 months of CS exposure, and mice showed significantly lower lung HDAC2 levels. Thus, CS-mediated down-regulation of HDAC2 in human macrophages and lung epithelial cells in vitro and in mouse lung in vivo involves the induction of serine/threonine phosphorylation and proteasomal degradation, which may have implications for steroid resistance and abnormal inflammation caused by cigarette smoke. PMID:18927347

  13. ORMDL3 contributes to the risk of atherosclerosis in Chinese Han population and mediates oxidized low-density lipoprotein-induced autophagy in endothelial cells.

    Science.gov (United States)

    Ma, Xiaochun; Qiu, Rongfang; Dang, Jie; Li, Jiangxia; Hu, Qin; Shan, Shan; Xin, Qian; Pan, Wenying; Bian, Xianli; Yuan, Qianqian; Long, Feng; Liu, Na; Li, Yan; Gao, Fei; Zou, Chengwei; Gong, Yaoqin; Liu, Qiji

    2015-11-25

    ORMDL sphingolipid biosynthesis regulator 3 (ORMDL3) is a universally confirmed susceptibility gene for asthma and has recently emerged as a crucial modulator in lipid metabolism, inflammation and endoplasmic reticulum (ER) stress-the mechanisms also closely involved in atherosclerosis (AS). Here we first presented the evidence of two single nucleotide polymorphisms regulating ORMDL3 expression (rs7216389 and rs9303277) significantly associated with AS risk and the evidence of increased ORMDL3 expression in AS cases compared to controls, in Chinese Han population. Following the detection of its statistical correlation with AS, we further explored the functional relevance of ORMDL3 and hypothesized a potential role mediating autophagy as autophagy is activated upon modified lipid, inflammation and ER stress. Our results demonstrated that in endothelial cells oxidized low-density lipoprotein (ox-LDL) up-regulated ORMDL3 expression and knockdown of ORMDL3 alleviated not only ox-LDL-induced but also basal autophagy. BECN1 is essential for autophagy initiation and silencing of ORMDL3 suppressed ox-LDL-induced as well as basal BECN1 expression. In addition, deletion of ORMDL3 resulted in greater sensitivity to ox-LDL-induced cell death. Taken together, ORMDL3 might represent a causal gene mediating autophagy in endothelial cells in the pathogenesis of AS.

  14. Nanomaterial-modulated autophagy: underlying mechanisms and functional consequences.

    Science.gov (United States)

    Zheng, Wei; Wei, Min; Li, Song; Le, Weidong

    2016-06-01

    Autophagy is an essential lysosome-dependent process that controls the quality of the cytoplasm and maintains cellular homeostasis, and dysfunction of this protein degradation system is correlated with various disorders. A growing body of evidence suggests that nanomaterials (NMs) have autophagy-modulating effects, thus predicting a valuable and promising application potential of NMs in the diagnosis and treatment of autophagy-related diseases. NMs exhibit unique physical, chemical and biofunctional properties, which may endow NMs with capabilities to modulate autophagy via various mechanisms. The present review highlights the impacts of various NMs on autophagy and their functional consequences. The possible underlying mechanisms for NM-modulated autophagy are also discussed.

  15. Ubiquitination directly enhances activity of the deubiquitinating enzyme ataxin-3

    OpenAIRE

    Todi, Sokol V.; Winborn, Brett J; Scaglione, K Matthew; Blount, Jessica R.; Travis, Sue M.; Paulson, Henry L.

    2009-01-01

    Deubiquitinating enzymes (DUBs) control the ubiquitination status of proteins in various cellular pathways. Regulation of the activity of DUBs, which is critically important to cellular homoeostasis, can be achieved at the level of gene expression, protein complex formation, or degradation. Here, we report that ubiquitination also directly regulates the activity of a DUB, ataxin-3, a polyglutamine disease protein implicated in protein quality control pathways. Ubiquitination enhances ubiquiti...

  16. Promoters active in interphase are bookmarked during mitosis by ubiquitination

    OpenAIRE

    Arora, Mansi; Jie ZHANG; Heine, George F.; Ozer, Gulcin; Liu, Hui-Wen; Huang, Kun; Parvin, Jeffrey D.

    2012-01-01

    We analyzed modification of chromatin by ubiquitination in human cells and whether this mark changes through the cell cycle. HeLa cells were synchronized at different stages and regions of the genome with ubiquitinated chromatin were identified by affinity purification coupled with next-generation sequencing. During interphase, ubiquitin marked the chromatin on the transcribed regions of ∼70% of highly active genes and deposition of this mark was sensitive to transcriptional inhibition. Promo...

  17. N-Methylcysteine-Mediated Total Chemical Synthesis of Ubiquitin Thioester

    OpenAIRE

    Erlich, Lesly A.; Ajish Kumar, K. S.; Haj-Yahya, Mahmood; Dawson, Philip E.; Brik, Ashraf

    2010-01-01

    Ubiquitin thioester is a key intermediate in the ubiquitylation of proteins and is formed enzymatically through the activation of α-COOH of ubiquitin in an ATP dependent manner using the E1 enzyme. The current methods used for the preparation of ubiquitin thioester rely on either the enzymatic machinery or on expressed protein ligation technology. In this article, we report a new chemical strategy, combining native chemical ligation and N-methylcysteine containing peptides, to chemically prep...

  18. RNF20 Links Histone H2B Ubiquitylation with Inflammation and Inflammation-Associated Cancer

    Science.gov (United States)

    Tarcic, Ohad; Pateras, Ioannis S.; Cooks, Tomer; Shema, Efrat; Kanterman, Julia; Ashkenazi, Hadas; Boocholez, Hana; Hubert, Ayala; Rotkopf, Ron; Baniyash, Michal; Pikarsky, Eli; Gorgoulis, Vassilis G.; Oren, Moshe

    2016-01-01

    Summary Factors linking inflammation and cancer are of great interest. We now report that the chromatin-targeting E3 ubiquitin ligase RNF20/RNF40, driving histone H2B monoubiquitylation (H2Bub1), modulates inflammation and inflammation-associated cancer in mice and humans. Downregulation of RNF20 and H2Bub1 favors recruitment of p65-containing nuclear factor κB (NF-κB) dimers over repressive p50 homodimers and decreases the heterochromatin mark H3K9me3 on a subset of NF-κB target genes to augment their transcription. Concordantly, RNF20+/− mice are predisposed to acute and chronic colonic inflammation and inflammation-associated colorectal cancer, with excessive myeloid-derived suppressor cells (MDSCs) that may quench antitumoral T cell activity. Notably, colons of human ulcerative colitis patients, as well as colorectal tumors, reveal downregulation of RNF20/RNF40 and H2Bub1 in both epithelium and stroma, supporting the clinical relevance of our tissue culture and mouse model findings. PMID:26854224

  19. RNF20 Links Histone H2B Ubiquitylation with Inflammation and Inflammation-Associated Cancer

    Directory of Open Access Journals (Sweden)

    Ohad Tarcic

    2016-02-01

    Full Text Available Factors linking inflammation and cancer are of great interest. We now report that the chromatin-targeting E3 ubiquitin ligase RNF20/RNF40, driving histone H2B monoubiquitylation (H2Bub1, modulates inflammation and inflammation-associated cancer in mice and humans. Downregulation of RNF20 and H2Bub1 favors recruitment of p65-containing nuclear factor κB (NF-κB dimers over repressive p50 homodimers and decreases the heterochromatin mark H3K9me3 on a subset of NF-κB target genes to augment their transcription. Concordantly, RNF20+/− mice are predisposed to acute and chronic colonic inflammation and inflammation-associated colorectal cancer, with excessive myeloid-derived suppressor cells (MDSCs that may quench antitumoral T cell activity. Notably, colons of human ulcerative colitis patients, as well as colorectal tumors, reveal downregulation of RNF20/RNF40 and H2Bub1 in both epithelium and stroma, supporting the clinical relevance of our tissue culture and mouse model findings.

  20. The role of autophagy in Parkinson's disease☆

    OpenAIRE

    Zhang, Lei; Dong, Yaru; Xu, Xiaoheng; Xu, Zhong

    2012-01-01

    Although Parkinson's disease is the most common neurodegenerative movement disorder, the mechanisms of pathogenesis remain poorly understood. Recent findings have shown that deregulation of the autophagy-lysosome pathway is involved in the pathogenesis of Parkinson's disease. This review summarizes the most recent findings and discusses the unique role of the autophagy-lysosome pathway in Parkinson's disease to highlight the possibility of Parkinson's disease treatment strategies that incorpo...

  1. Skeletal Muscle Autophagy: A New Metabolic Regulator

    OpenAIRE

    Neel, Brian A.; Lin, Yuxi; Pessin, Jeffrey E.

    2013-01-01

    Autophagy classically functions as a physiological process to degrade cytoplasmic components, protein aggregates, and/or organelles, as a mechanism for nutrient breakdown, and as a regulator of cellular architecture. Proper autophagic flux is vital for both functional skeletal muscle, which controls support and movement of the skeleton, and muscle metabolism. The role of autophagy as a metabolic regulator in muscle has been previously studied; however, the underlying molecular mechanisms that...

  2. Mechanisms of mitochondria and autophagy crosstalk

    OpenAIRE

    Rambold, Angelika S.; Lippincott-Schwartz, Jennifer

    2011-01-01

    Autophagy is a cellular survival pathway that recycles intracellular components to compensate for nutrient depletion and ensures the appropriate degradation of organelles. Mitochondrial number and health are regulated by mitophagy, a process by which excessive or damaged mitochondria are subjected to autophagic degradation. Autophagy is thus a key determinant for mitochondrial health and proper cell function. Mitophagic malfunction has been recently proposed to contribute to progressive neuro...

  3. RUFY4: Immunity piggybacking on autophagy?

    Science.gov (United States)

    Terawaki, Seigo; Camosseto, Voahirana; Pierre, Philippe; Gatti, Evelina

    2016-01-01

    Although autophagy is a highly conserved mechanism among species and cell types, few are the molecules involved with the autophagic process that display cell- or tissue- specific expression. We have unraveled the positive regulatory role on autophagy of RUFY4 (RUN and FYVE domain containing 4), which is expressed in subsets of immune cells, including dendritic cells (DCs). DCs orchestrate the eradication of pathogens by coordinating the action of the different cell types involved in microbe recognition and destruction during the immune response. To fulfill this function, DC display particular regulation of their endocytic and autophagy pathways in response to the immune environment. Autophagy flux is downmodulated in DCs upon microbe sensing, but is remarkably augmented, when cells are differentiated in the presence of the pleiotropic cytokine IL4 (interleukin 4). From gene expression studies aimed at comparing the impact of IL4 on DC differentiation, we identified RUFY4, as a novel regulator that augments autophagy flux and, when overexpressed, induces drastic membrane redistribution and strongly tethers lysosomes. RUFY4 is therefore one of the few known positive regulators of autophagy that is expressed in a cell-specific manner or under specific immunological conditions associated with IL4 expression such as allergic asthma.

  4. Autophagy in the control of food intake.

    Science.gov (United States)

    Singh, Rajat

    2012-04-01

    The cellular nutrient sensing apparatus detects nutritional depletion and transmits this information to downstream effectors that generate energy from alternate sources. Autophagy is a crucial catabolic pathway that turns over redundant cytoplasmic components in lysosomes to provide energy to the starved cell. Recent studies have described a role for hypothalamic autophagy in the control of food intake and energy balance. Activated autophagy in hypothalamic neurons during starvation mobilized neuron-intrinsic lipids to generate free fatty acids that increased AgRP levels. AgRP neuron-specific inhibition of autophagy decreased fasting-induced increases in AgRP levels and food intake. Deletion of autophagy in AgRP neurons led to constitutive increases in levels of proopiomelanocortin and its active processed product, α-melanocyte stimulating hormone that contributed to reduced adiposity in these rodents. The current manuscript discusses these new findings and raises additional questions that may help understand how hypothalamic autophagy controls food intake and energy balance. These studies may have implications for designing new therapies against obesity and insulin resistance. PMID:23700515

  5. Autophagy in stem and progenitor cells.

    Science.gov (United States)

    Rodolfo, Carlo; Di Bartolomeo, Sabrina; Cecconi, Francesco

    2016-02-01

    Autophagy is a highly conserved cellular process, responsible for the degradation and recycling of damaged and/or outlived proteins and organelles. This is the major cellular pathway, acting throughout the formation of cytosolic vesicles, called autophagosomes, for the delivering to lysosome. Recycling of cellular components through autophagy is a crucial step for cell homeostasis as well as for tissue remodelling during development. Impairment of this process has been related to the pathogenesis of various diseases, such as cancer and neurodegeneration, to the response to bacterial and viral infections, and to ageing. The ability of stem cells to self-renew and differentiate into the mature cells of the body renders this unique type of cell highly crucial to development and tissue renewal, not least in various diseases. During the last two decades, extensive knowledge about autophagy roles and regulation in somatic cells has been acquired; however, the picture about the role and the regulation of autophagy in the different types of stem cells is still largely unknown. Autophagy is a major player in the quality control and maintenance of cellular homeostasis, both crucial factors for stem cells during an organism's life. In this review, we have highlighted the most significant advances in the comprehension of autophagy regulation in embryonic and tissue stem cells, as well as in cancer stem cells and induced pluripotent cells.

  6. Macrophage specific caspase-1/11 deficiency protects against cholesterol crystallization and hepatic inflammation in hyperlipidemic mice.

    Directory of Open Access Journals (Sweden)

    Tim Hendrikx

    Full Text Available While non-alcoholic steatohepatitis (NASH is characterized by hepatic steatosis combined with inflammation, the mechanisms triggering hepatic inflammation are unknown. In Ldlr(-/- mice, we have previously shown that lysosomal cholesterol accumulation in Kupffer cells (KCs correlates with hepatic inflammation and cholesterol crystallization. Previously, cholesterol crystals have been shown to induce the activation of inflammasomes. Inflammasomes are protein complexes that induce the processing and release of pro-inflammatory cytokines IL-1b and IL-18 via caspase-1 activation. Whereas caspase-1 activation is independent of caspase-11 in the canonical pathway of inflammasome activation, caspase-11 was found to trigger caspase-1-dependent IL-1b and IL-18 in response to non-canonical inflammasome activators. So far, it has not been investigated whether inflammasome activation stimulates the formation of cholesterol crystals. We hypothesized that inflammasome activation in KCs stimulates cholesterol crystallization, thereby leading to hepatic inflammation.Ldlr (-/- mice were transplanted (tp with wild-type (Wt or caspase-1/11(-/- (dKO bone marrow and fed either regular chow or a high-fat, high-cholesterol (HFC diet for 12 weeks. In vitro, bone marrow derived macrophages (BMDM from wt or caspase-1/11(-/- mice were incubated with oxLDL for 24h and autophagy was assessed.In line with our hypothesis, caspase-1/11(-/--tp mice had less severe hepatic inflammation than Wt-tp animals, as evident from liver histology and gene expression analysis in isolated KCs. Mechanistically, KCs from caspase-1/11(-/--tp mice showed less cholesterol crystals, enhanced cholesterol efflux and increased autophagy. In wt BMDM, oxLDL incubation led to disturbed autophagy activity whereas BMDM from caspase-1/11(-/- mice had normal autophagy activity.Altogether, these data suggest a vicious cycle whereby disturbed autophagy and decreased cholesterol efflux leads to newly formed

  7. Origin and diversification of TRIM ubiquitin ligases.

    Directory of Open Access Journals (Sweden)

    Ignacio Marín

    Full Text Available Most proteins of the TRIM family (also known as RBCC family are ubiquitin ligases that share a peculiar protein structure, characterized by including an N-terminal RING finger domain closely followed by one or two B-boxes. Additional protein domains found at their C termini have been used to classify TRIM proteins into classes. TRIMs are involved in multiple cellular processes and many of them are essential components of the innate immunity system of animal species. In humans, it has been shown that mutations in several TRIM-encoding genes lead to diverse genetic diseases and contribute to several types of cancer. They had been hitherto detected only in animals. In this work, by comprehensively analyzing the available diversity of TRIM and TRIM-like protein sequences and evaluating their evolutionary patterns, an improved classification of the TRIM family is obtained. Members of one of the TRIM subfamilies defined, called Subfamily A, turn to be present not only in animals, but also in many other eukaryotes, such as fungi, apusozoans, alveolates, excavates and plants. The rest of subfamilies are animal-specific and several of them originated only recently. Subfamily A proteins are characterized by containing a MATH domain, suggesting a potential evolutionary connection between TRIM proteins and a different type of ubiquitin ligases, known as TRAFs, which contain quite similar MATH domains. These results indicate that the TRIM family emerged much earlier than so far thought and contribute to our understanding of its origin and diversification. The structural and evolutionary links with the TRAF family of ubiquitin ligases can be experimentally explored to determine whether functional connections also exist.

  8. The DUB-ious lack of ALIS in Salmonella infection: a Salmonella deubiquitinase regulates the autophagy of protein aggregates.

    Science.gov (United States)

    Thomas, Mair; Mesquita, Francisco S; Holden, David W

    2012-12-01

    Ubiquitinated aggregates are formed in eukaryotic cells in response to several external stimuli, including exposure to bacterial lipopolysaccharide (LPS). Although Salmonella enterica serovar Typhimurium (S. Typhimurium) LPS has been shown to induce aggresome-like induced structures (ALIS) in macrophages, these have not been described in S. Typhimurium-infected macrophages. Given that LPS is present in infection, this suggests that S. Typhimurium might suppress the formation of ALIS. We found that S. Typhimurium induces the formation of ubiquitinated aggregates in epithelial cells and macrophages, but that their presence is masked by the deubiquitinase (DUB) activity of the S. Typhimurium virulence protein, SseL. SseL deubiquitinates SQSTM1/p62-bound proteins found in S. Typhimurium-induced aggregates and ALIS, and reduces the recruitment of autophagic components. While the functions of ALIS and other ubiquitinated aggregates remain unclear, they serve to sequester cytosolic proteins under a variety of stress conditions and are suggested to be involved in host immune defense. During infection, the deubiquitinase activity of SseL reduces autophagic flux in infected cells and favors bacterial replication. This is a new example of how a bacterial pathogen counteracts the autophagy pathway through the action of a translocated virulence protein.

  9. Autophagy in acute brain injury: feast, famine, or folly?

    Science.gov (United States)

    Smith, Craig M; Chen, Yaming; Sullivan, Mara L; Kochanek, Patrick M; Clark, Robert S B

    2011-07-01

    In the central nervous system, increased autophagy has now been reported after traumatic brain and spinal cord injury, cerebral ischemia, intracerebral hemorrhage, and seizures. This increase in autophagy could be physiologic, converting damaged or dysfunctional proteins, lipids, and/or organelles to their amino acid and fatty acid components for recycling. On the other hand, this increase in autophagy could be supraphysiologic, perhaps consuming and eliminating functional proteins, lipids, and/or organelles as well. Whether an increase in autophagy is beneficial (feast) or detrimental (famine) in brain likely depends on both the burden of intracellular substrate targeted for autophagy and the capacity of the cell's autophagic machinery. Of course, increased autophagy observed after brain injury could also simply be an epiphenomenon (folly). These divergent possibilities have clear ramifications for designing therapeutic strategies targeting autophagy after acute brain injury and are the subject of this review. This article is part of a Special Issue entitled "Autophagy and protein degradation in neurological diseases."

  10. Regulation of the Tumor-Suppressor Function of the Class III Phosphatidylinositol 3-Kinase Complex by Ubiquitin and SUMO

    Energy Technology Data Exchange (ETDEWEB)

    Reidick, Christina [Biochemie Intrazellulärer Transportprozesse, Ruhr-Universität Bochum, Bochum 44801 (Germany); El Magraoui, Fouzi; Meyer, Helmut E. [Biomedical Research, Human Brain Proteomics II, Leibniz-Institut für Analytische Wissenschaften-ISAS, Dortmund 44139 (Germany); Stenmark, Harald [Department of Biochemistry, Institute for Cancer Research, Oslo University Hospital, Montebello, Oslo 0310 (Norway); Platta, Harald W., E-mail: harald.platta@rub.de [Biochemie Intrazellulärer Transportprozesse, Ruhr-Universität Bochum, Bochum 44801 (Germany)

    2014-12-23

    The occurrence of cancer is often associated with a dysfunction in one of the three central membrane-involution processes—autophagy, endocytosis or cytokinesis. Interestingly, all three pathways are controlled by the same central signaling module: the class III phosphatidylinositol 3-kinase (PI3K-III) complex and its catalytic product, the phosphorylated lipid phosphatidylinositol 3-phosphate (PtdIns3P). The activity of the catalytic subunit of the PI3K-III complex, the lipid-kinase VPS34, requires the presence of the membrane-targeting factor VPS15 as well as the adaptor protein Beclin 1. Furthermore, a growing list of regulatory proteins associates with VPS34 via Beclin 1. These accessory factors define distinct subunit compositions and thereby guide the PI3K-III complex to its different cellular and physiological roles. Here we discuss the regulation of the PI3K-III complex components by ubiquitination and SUMOylation. Especially Beclin 1 has emerged as a highly regulated protein, which can be modified with Lys11-, Lys48- or Lys63-linked polyubiquitin chains catalyzed by distinct E3 ligases from the RING-, HECT-, RBR- or Cullin-type. We also point out other cross-links of these ligases with autophagy in order to discuss how these data might be merged into a general concept.

  11. Canine hereditary ataxia in old english sheepdogs and gordon setters is associated with a defect in the autophagy gene encoding RAB24.

    Directory of Open Access Journals (Sweden)

    Caryline Agler

    2014-02-01

    Full Text Available Old English Sheepdogs and Gordon Setters suffer from a juvenile onset, autosomal recessive form of canine hereditary ataxia primarily affecting the Purkinje neuron of the cerebellar cortex. The clinical and histological characteristics are analogous to hereditary ataxias in humans. Linkage and genome-wide association studies on a cohort of related Old English Sheepdogs identified a region on CFA4 strongly associated with the disease phenotype. Targeted sequence capture and next generation sequencing of the region identified an A to C single nucleotide polymorphism (SNP located at position 113 in exon 1 of an autophagy gene, RAB24, that segregated with the phenotype. Genotyping of six additional breeds of dogs affected with hereditary ataxia identified the same polymorphism in affected Gordon Setters that segregated perfectly with phenotype. The other breeds tested did not have the polymorphism. Genome-wide SNP genotyping of Gordon Setters identified a 1.9 MB region with an identical haplotype to affected Old English Sheepdogs. Histopathology, immunohistochemistry and ultrastructural evaluation of the brains of affected dogs from both breeds identified dramatic Purkinje neuron loss with axonal spheroids, accumulation of autophagosomes, ubiquitin positive inclusions and a diffuse increase in cytoplasmic neuronal ubiquitin staining. These findings recapitulate the changes reported in mice with induced neuron-specific autophagy defects. Taken together, our results suggest that a defect in RAB24, a gene associated with autophagy, is highly associated with and may contribute to canine hereditary ataxia in Old English Sheepdogs and Gordon Setters. This finding suggests that detailed investigation of autophagy pathways should be undertaken in human hereditary ataxia.

  12. The Ca{sup 2+} channel TRPML3 specifically interacts with the mammalian ATG8 homologue GATE16 to regulate autophagy

    Energy Technology Data Exchange (ETDEWEB)

    Choi, Suzy; Kim, Hyun Jin, E-mail: kimhyunjin@skku.edu

    2014-01-03

    Highlights: •Split-ubiquitin MY2H screen identified GATE16 as an interacting protein of TRPML3. •TRPML3 specifically binds to a mammalian ATG8 homologue GATE16, not to LC3B. •The interaction of TRPML3 with GATE16 facilitates autophagosome formation. •GATE16 is expressed in both autophagosome and extra-autophagosomal compartments. -- Abstract: TRPML3 is a Ca{sup 2+} permeable cation channel expressed in multiple intracellular compartments. Although TRPML3 is implicated in autophagy, how TRPML3 can regulate autophagy is not understood. To search interacting proteins with TRPML3 in autophagy, we performed split-ubiquitin membrane yeast two-hybrid (MY2H) screening with TRPML3-loop as a bait and identified GATE16, a mammalian ATG8 homologue. GST pull-down assay revealed that TRPML3 and TRPML3-loop specifically bind to GATE16, not to LC3B. Co-immunoprecipitation (co-IP) experiments showed that TRPML3 and TRPML3-loop pull down only the lipidated form of GATE16, indicating that the interaction occurs exclusively at the organellar membrane. The interaction of TRPML3 with GATE16 and GATE16-positive vesicle formation were increased in starvation induced autophagy, suggesting that the interaction facilitates the function of GATE16 in autophagosome formation. However, GATE16 was not required for TRPML3 trafficking to autophagosomes. Experiments using dominant-negative (DN) TRPML3(D458K) showed that GATE16 is localized not only in autophagosomes but also in extra-autophagosomal compartments, by contrast with LC3B. Since GATE16 acts at a later stage of the autophagosome biogenesis, our results suggest that TRPML3 plays a role in autophagosome maturation through the interaction with GATE16, by providing Ca{sup 2+} in the fusion process.

  13. HUWE1 and TRIP12 collaborate in degradation of ubiquitin-fusion proteins and misframed ubiquitin.

    Directory of Open Access Journals (Sweden)

    Esben G Poulsen

    Full Text Available In eukaryotic cells an uncleavable ubiquitin moiety conjugated to the N-terminus of a protein signals the degradation of the fusion protein via the proteasome-dependent ubiquitin fusion degradation (UFD pathway. In yeast the molecular mechanism of the UFD pathway has been well characterized. Recently the human E3 ubiquitin-protein ligase TRIP12 was connected with the UFD pathway, but little is otherwise known about this system in mammalian cells. In the present work, we utilized high-throughput imaging on cells transfected with a targeted siRNA library to identify components involved in degradation of the UFD substrate Ub(G76V-YFP. The most significant hits from the screen were the E3 ubiquitin-protein ligase HUWE1, as well as PSMD7 and PSMD14 that encode proteasome subunits. Accordingly, knock down of HUWE1 led to an increase in the steady state level and a retarded degradation of the UFD substrate. Knock down of HUWE1 also led to a stabilization of the physiological UFD substrate UBB(+1. Precipitation experiments revealed that HUWE1 is associated with both the Ub(G76V-YFP substrate and the 26S proteasome, indicating that it functions late in the UFD pathway. Double knock down of HUWE1 and TRIP12 resulted in an additive stabilization of the substrate, suggesting that HUWE1 and TRIP12 function in parallel during UFD. However, even when both HUWE1 and TRIP12 are downregulated, ubiquitylation of the UFD substrate was still apparent, revealing functional redundancy between HUWE1, TRIP12 and yet other ubiquitin-protein ligases.

  14. The influence of autophagy on mouse inflammatory responses caused by Salmonella enterica serovar Typhimurium with spv genes

    Institute of Scientific and Technical Information of China (English)

    LI Yuan-Yuan; WU Shu-Yan; CHU Yuan-Yuan; LIAO LI; LIQiong; HUANG Rui

    2011-01-01

    An investigation into the effects of Salmonella plasmid virulence genes (spv) on autophagy,apoptosis,and inflammation was carried out in mice,using a strain of Salmonella enterica serovar Typhimurium (S.typhimurium) SR-11 carrying spv.Strain BRD509 without spy was used as a control.Results showed that the expression of autophagy protein Beclin-1 in the livers and spleens in the SR-11 group was lower than that in the BRD509 group,while the apoptosis protein,Caspase-3,was higher in the SR-11 group.Inflammatory cytokine levels [interleukin 12 (IL-12) and interferon γ (IFN-γ)] were higher in the SR-11 group compared with those in the BRD509 group since 4 d post-infection.In addition,we found an increase in severe pathological changes and larger viable bacterial amounts in livers and spleens in the SR-11 group.After intervention with autophagy agonist rapamycin (RAPA),Beclin-1 expression increased in both groups,while Caspase-3 expression was different between the two groups: Caspase-3 decreased in the SR-11 group but increased in the BRD509 group.Moreover,RAPA decreased cytokine levels,bacterial quantity and organ-related injury in the SR-11 group whereas RAPA increased cytokine levels and aggravated organ injury in the BRD509 group.Results from these studies suggest that S.typhimurium with spv genes may exacerbate infection by inhibiting autophagy and affecting the production of inflammatory cytokines.RAPA-enhanced autophagy may improve the secretion of cytokines in order to protect the host from damaging by Salmonella infection.Our study suggests that the regulation of cellular autophagy may play a role in the prevention and control of certain infectious diseases.

  15. Knockdown of GSK3β increases basal autophagy and AMPK signalling in nutrient-laden human aortic endothelial cells

    Science.gov (United States)

    Weikel, Karen A.; Cacicedo, José M.; Ruderman, Neil B.; Ido, Yasuo

    2016-01-01

    High concentrations of glucose and palmitate increase endothelial cell inflammation and apoptosis, events that often precede atherogenesis. They may do so by decreasing basal autophagy and AMP-activated protein kinase (AMPK) activity, although the mechanisms by which this occurs are not clear. Decreased function of the lysosome, an organelle required for autophagy and AMPK, have been associated with hyperactivity of glycogen synthase kinase 3β (GSK3β). To determine whether GSK3β affects nutrient-induced changes in autophagy and AMPK activity, we used a primary human aortic endothelial cell (HAEC) model of type 2 diabetes that we had previously characterized with impaired AMPK activity and autophagy [Weikel et al. (2015) Am. J. Phys. Cell Physiol. 308, C249–C263]. Presently, we found that incubation of HAECs with excess nutrients (25 mM glucose and 0.4 mM palmitate) increased GSK3β activity and impaired lysosome acidification. Suppression of GSK3β in these cells by treatment with a chemical inhibitor or overexpression of kinase-dead GSK3β attenuated these lysosomal changes. Under control and excess nutrient conditions, knockdown of GSK3β increased autophagosome formation, forkhead box protein O1 (FOXO1) activity and AMPK signalling and decreased Akt signalling. Similar changes in autophagy, AMPK and Akt signalling were observed in aortas from mice treated with the GSK3β inhibitor CHIR 99021. Thus, increasing basal autophagy and AMPK activity by inhibiting GSK3β may be an effective strategy in the setting of hyperglycaemia and dyslipidaemia for restoring endothelial cell health and reducing atherogenesis. PMID:27534430

  16. Dual role of autophagy in HIV-1 replication and pathogenesis

    OpenAIRE

    Killian M

    2012-01-01

    Abstract Autophagy, the major mechanism for degrading long-lived intracellular proteins and organelles, is essential for eukaryotic cell homeostasis. Autophagy also defends the cell against invasion by microorganisms and has important roles in innate and adaptive immunity. Increasingly evident is that HIV-1 replication is dependent on select components of autophagy. Fittingly, HIV-1 proteins are able to modulate autophagy to maximize virus production. At the same time, HIV-1 proteins appear t...

  17. Bone Cell Autophagy Is Regulated by Environmental Factors

    OpenAIRE

    Zahm, Adam M.; Bohensky, Jolene; Adams, Christopher S.; Shapiro, Irving M.; Srinivas, Vickram

    2011-01-01

    The goal of this investigation was to ascertain whether bone cells undergo autophagy and to determine if this process is regulated by environmental factors. We showed that osteocytes in both murine and human cortical bone display a punctuate distribution of microtubule-associated protein light chain 3, indicative of autophagy. In addition, we noted a basal level of autophagy in preosteocyte-like murine long bone-derived osteocytic (MLO)-A5 cells. Autophagy was upregulated following nutrient d...

  18. Ubiquitination-dependent mechanisms regulate synaptic growth and function.

    Science.gov (United States)

    DiAntonio, A; Haghighi, A P; Portman, S L; Lee, J D; Amaranto, A M; Goodman, C S

    2001-07-26

    The covalent attachment of ubiquitin to cellular proteins is a powerful mechanism for controlling protein activity and localization. Ubiquitination is a reversible modification promoted by ubiquitin ligases and antagonized by deubiquitinating proteases. Ubiquitin-dependent mechanisms regulate many important processes including cell-cycle progression, apoptosis and transcriptional regulation. Here we show that ubiquitin-dependent mechanisms regulate synaptic development at the Drosophila neuromuscular junction (NMJ). Neuronal overexpression of the deubiquitinating protease fat facets leads to a profound disruption of synaptic growth control; there is a large increase in the number of synaptic boutons, an elaboration of the synaptic branching pattern, and a disruption of synaptic function. Antagonizing the ubiquitination pathway in neurons by expression of the yeast deubiquitinating protease UBP2 (ref. 5) also produces synaptic overgrowth and dysfunction. Genetic interactions between fat facets and highwire, a negative regulator of synaptic growth that has structural homology to a family of ubiquitin ligases, suggest that synaptic development may be controlled by the balance between positive and negative regulators of ubiquitination.

  19. The Ubiquitin-Proteasome Pathway and Synaptic Plasticity

    Science.gov (United States)

    Hegde, Ashok N.

    2010-01-01

    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…

  20. Lysine Ubiquitination and Acetylation of Human Cardiac 20S Proteasomes

    Science.gov (United States)

    Lau, Edward; Choi, Howard JH; Ng, Dominic CM; Meyer, David; Fang, Caiyun; Li, Haomin; Wang, Ding; Zelaya, Ivette M; Yates, John R; Lam, Maggie PY

    2016-01-01

    Purpose Altered proteasome functions are associated with multiple cardiomyopathies. While the proteasome targets poly-ubiquitinated proteins for destruction, it itself is modifiable by ubiquitination. We aim to identify the exact ubiquitination sites on cardiac proteasomes and examine whether they are also subject to acetylations. Experimental design Assembled cardiac 20S proteasome complexes were purified from five human hearts with ischemic cardiomyopathy, then analyzed by high-resolution MS to identify ubiquitination and acetylation sites. We developed a library search strategy that may be used to complement database search in identifying PTM in different samples. Results We identified 63 ubiquitinated lysines from intact human cardiac 20S proteasomes. In parallel, 65 acetylated residues were also discovered, 39 of which shared with ubiquitination sites. Conclusion and clinical relevance This is the most comprehensive characterization of cardiac proteasome ubiquitination to-date. There are significant overlaps between the discovered ubiquitination and acetylation sites, permitting potential crosstalk in regulating proteasome functions. The information presented here will aid future therapeutic strategies aimed at regulating the functions of cardiac proteasomes. PMID:24957502

  1. From the urea cycle to autophagy: Alfred J. Meijer

    NARCIS (Netherlands)

    D.J. Klionsky; A.J. Meijer

    2011-01-01

    Now that many of the components of the autophagy machinery have been identified, in particular the autophagy-related (Atg) proteins, increasing focus is being directed toward the role of autophagy in health and disease. Accordingly, it is of ever-greater importance to understand the central role of

  2. Autophagy modulates the Mycobacterium tuberculosis-induced cytokine response

    NARCIS (Netherlands)

    Kleinnijenhuis, J.; Oosting, M.; Plantinga, T.S.; Meer, J.W.M. van der; Joosten, L.A.B.; Crevel, R. van; Netea, M.G.

    2011-01-01

    Both autophagy and pro-inflammatory cytokines are involved in the host defence against mycobacteria, but little is known regarding the effect of autophagy on Mycobacterium tuberculosis (MTB)-induced cytokine production. In the present study, we assessed the effect of autophagy on production of monoc

  3. Autophagy and Retromer Components in Plant Innate Immunity

    DEFF Research Database (Denmark)

    Munch, David

    -hormone salicylic acid. Here, I present data that make it clear that NPR1 does not directly regulate autophagy, but instead control stress responses that indirectly activate autophagy. The observations presented will also clarify why autophagy has been described as being both a pro-death and pro-life pathway under...

  4. COMMD1-mediated ubiquitination regulates CFTR trafficking.

    Directory of Open Access Journals (Sweden)

    Loïc Drévillon

    Full Text Available The CFTR (cystic fibrosis transmembrane conductance regulator protein is a large polytopic protein whose biogenesis is inefficient. To better understand the regulation of CFTR processing and trafficking, we conducted a genetic screen that identified COMMD1 as a new CFTR partner. COMMD1 is a protein associated with multiple cellular pathways, including the regulation of hepatic copper excretion, sodium uptake through interaction with ENaC (epithelial sodium channel and NF-kappaB signaling. In this study, we show that COMMD1 interacts with CFTR in cells expressing both proteins endogenously. This interaction promotes CFTR cell surface expression as assessed by biotinylation experiments in heterologously expressing cells through regulation of CFTR ubiquitination. In summary, our data demonstrate that CFTR is protected from ubiquitination by COMMD1, which sustains CFTR expression at the plasma membrane. Thus, increasing COMMD1 expression may provide an approach to simultaneously inhibit ENaC absorption and enhance CFTR trafficking, two major issues in cystic fibrosis.

  5. High Fat Diet-Induced Skeletal Muscle Wasting Is Decreased by Mesenchymal Stem Cells Administration: Implications on Oxidative Stress, Ubiquitin Proteasome Pathway Activation, and Myonuclear Apoptosis

    Science.gov (United States)

    Aravena, Javier; Cabrera, Daniel; Simon, Felipe; Ezquer, Fernando

    2016-01-01

    Obesity can lead to skeletal muscle atrophy, a pathological condition characterized by the loss of strength and muscle mass. A feature of muscle atrophy is a decrease of myofibrillar proteins as a result of ubiquitin proteasome pathway overactivation, as evidenced by increased expression of the muscle-specific ubiquitin ligases atrogin-1 and MuRF-1. Additionally, other mechanisms are related to muscle wasting, including oxidative stress, myonuclear apoptosis, and autophagy. Stem cells are an emerging therapy in the treatment of chronic diseases such as high fat diet-induced obesity. Mesenchymal stem cells (MSCs) are a population of self-renewable and undifferentiated cells present in the bone marrow and other mesenchymal tissues of adult individuals. The present study is the first to analyze the effects of systemic MSC administration on high fat diet-induced skeletal muscle atrophy in the tibialis anterior of mice. Treatment with MSCs reduced losses of muscle strength and mass, decreases of fiber diameter and myosin heavy chain protein levels, and fiber type transitions. Underlying these antiatrophic effects, MSC administration also decreased ubiquitin proteasome pathway activation, oxidative stress, and myonuclear apoptosis. These results are the first to indicate that systemically administered MSCs could prevent muscle wasting associated with high fat diet-induced obesity and diabetes. PMID:27579157

  6. High Fat Diet-Induced Skeletal Muscle Wasting Is Decreased by Mesenchymal Stem Cells Administration: Implications on Oxidative Stress, Ubiquitin Proteasome Pathway Activation, and Myonuclear Apoptosis

    Directory of Open Access Journals (Sweden)

    Johanna Abrigo

    2016-01-01

    Full Text Available Obesity can lead to skeletal muscle atrophy, a pathological condition characterized by the loss of strength and muscle mass. A feature of muscle atrophy is a decrease of myofibrillar proteins as a result of ubiquitin proteasome pathway overactivation, as evidenced by increased expression of the muscle-specific ubiquitin ligases atrogin-1 and MuRF-1. Additionally, other mechanisms are related to muscle wasting, including oxidative stress, myonuclear apoptosis, and autophagy. Stem cells are an emerging therapy in the treatment of chronic diseases such as high fat diet-induced obesity. Mesenchymal stem cells (MSCs are a population of self-renewable and undifferentiated cells present in the bone marrow and other mesenchymal tissues of adult individuals. The present study is the first to analyze the effects of systemic MSC administration on high fat diet-induced skeletal muscle atrophy in the tibialis anterior of mice. Treatment with MSCs reduced losses of muscle strength and mass, decreases of fiber diameter and myosin heavy chain protein levels, and fiber type transitions. Underlying these antiatrophic effects, MSC administration also decreased ubiquitin proteasome pathway activation, oxidative stress, and myonuclear apoptosis. These results are the first to indicate that systemically administered MSCs could prevent muscle wasting associated with high fat diet-induced obesity and diabetes.

  7. Lipid raft-dependent FcepsilonRI ubiquitination regulates receptor endocytosis through the action of ubiquitin binding adaptors.

    Directory of Open Access Journals (Sweden)

    Rosa Molfetta

    Full Text Available The best characterized role for ubiquitination of membrane receptors is to negatively regulate signaling by targeting receptors for lysosomal degradation. The high affinity receptor for IgE (FcepsilonRI expressed on mast cells and basophils is rapidly ubiquitinated upon antigen stimulation. However, the nature and the role of this covalent modification are still largelly unknown. Here, we show that FcepsilonRI subunits are preferentially ubiquitinated at multiple sites upon stimulation, and provide evidence for a role of ubiquitin as an internalization signal: under conditions of impaired receptor ubiquitination a decrease of receptor entry is observed by FACS analysis and fluorescence microscopy. We also used biochemical approaches combined with fluorescence microscopy, to demonstrate that receptor endocytosis requires the integrity of specific membrane domains, namely lipid rafts. Additionally, by RNA interference we demonstrate the involvement of ubiquitin-binding endocytic adaptors in FcepsilonRI internalization and sorting. Notably, the triple depletion of Eps15, Eps15R and Epsin1 negatively affects the early steps of Ag-induced receptor endocytosis, whereas Hrs depletion retains ubiquitinated receptors into early endosomes and partially prevents their sorting into lysosomes for degradation. Our results are compatible with a scenario in which the accumulation of engaged receptor subunits into lipid rafts is required for receptor ubiquitination, a prerequisite for efficient receptor internalization, sorting and delivery to a lysosomal compartment.

  8. PKC-Dependent GlyT1 Ubiquitination Occurs Independent of Phosphorylation: Inespecificity in Lysine Selection for Ubiquitination.

    Directory of Open Access Journals (Sweden)

    Susana P Barrera

    Full Text Available Neurotransmitter transporter ubiquitination is emerging as the main mechanism for endocytosis and sorting of cargo into lysosomes. In this study, we demonstrate PKC-dependent ubiquitination of three different isoforms of the glycine transporter 1 (GlyT1. Incubation of cells expressing transporter with the PKC activator phorbol ester induced a dramatic, time-dependent increase in GlyT1 ubiquitination, followed by accumulation of GlyT1 in EEA1 positive early endosomes. This occurred via a mechanism that was abolished by inhibition of PKC. GlyT1 endocytosis was confirmed in both retinal sections and primary cultures of mouse amacrine neurons. Replacement of only all lysines in the N-and C-termini to arginines prevented ubiquitination and endocytosis, displaying redundancy in the mechanism of ubiquitination. Interestingly, a 40-50% reduction in glycine uptake was detected in phorbol-ester stimulated cells expressing the WT-GlyT1, whereas no significant change was for the mutant protein, demonstrating that endocytosis participates in the reduction of uptake. Consistent with previous findings for the dopamine transporter DAT, ubiquitination of GlyT1 tails functions as sorting signal to deliver transporter into the lysosome and removal of ubiquitination sites dramatically attenuated the rate of GlyT1 degradation. Finally, we showed for the first time that PKC-dependent GlyT1 phosphorylation was not affected by removal of ubiquitination sites, suggesting separate PKC-dependent signaling events for these posttranslational modifications.

  9. Amino acid metabolism inhibits antibody-driven kidney injury by inducing autophagy.

    Science.gov (United States)

    Chaudhary, Kapil; Shinde, Rahul; Liu, Haiyun; Gnana-Prakasam, Jaya P; Veeranan-Karmegam, Rajalakshmi; Huang, Lei; Ravishankar, Buvana; Bradley, Jillian; Kvirkvelia, Nino; McMenamin, Malgorzata; Xiao, Wei; Kleven, Daniel; Mellor, Andrew L; Madaio, Michael P; McGaha, Tracy L

    2015-06-15

    Inflammatory kidney disease is a major clinical problem that can result in end-stage renal failure. In this article, we show that Ab-mediated inflammatory kidney injury and renal disease in a mouse nephrotoxic serum nephritis model was inhibited by amino acid metabolism and a protective autophagic response. The metabolic signal was driven by IFN-γ-mediated induction of indoleamine 2,3-dioxygenase 1 (IDO1) enzyme activity with subsequent activation of a stress response dependent on the eIF2α kinase general control nonderepressible 2 (GCN2). Activation of GCN2 suppressed proinflammatory cytokine production in glomeruli and reduced macrophage recruitment to the kidney during the incipient stage of Ab-induced glomerular inflammation. Further, inhibition of autophagy or genetic ablation of Ido1 or Gcn2 converted Ab-induced, self-limiting nephritis to fatal end-stage renal disease. Conversely, increasing kidney IDO1 activity or treating mice with a GCN2 agonist induced autophagy and protected mice from nephritic kidney damage. Finally, kidney tissue from patients with Ab-driven nephropathy showed increased IDO1 abundance and stress gene expression. Thus, these findings support the hypothesis that the IDO-GCN2 pathway in glomerular stromal cells is a critical negative feedback mechanism that limits inflammatory renal pathologic changes by inducing autophagy.

  10. Autophagy gene polymorphism is associated with susceptibility to leprosy by affecting inflammatory cytokines.

    Science.gov (United States)

    Yang, Degang; Chen, Jia; Shi, Chao; Jing, Zhichun; Song, Ningjing

    2014-04-01

    Autophagy and inflammation closely interact with each other, and together, they play critical roles in bacterial infection. Leprosy is caused by the infection of Mycobacterium leprae (M. leprae). The objective of the study was to investigate the association between polymorphisms in IRGM, an autophagy gene, and susceptibility to leprosy, and identify possible functions of the polymorphism in the infection of M. leprae. Two polymorphisms in IRGM, rs4958842 and rs13361189, were tested in 412 leprosy cases and 432 healthy controls. Levels of inflammatory cytokines including interleukin 1 beta, IL-4, IL-6, and interferon gamma (INF-γ) were measured after the infection of M. leprae in the peripheral blood mononuclear cell (PBMC) of subjects with different genotypes of rs13361189. Data showed that prevalence of rs13361189TC and CC genotypes were significantly higher in leprosy patients than in healthy controls (odds ratio (OR) = 1.49, 95 % confidence interval (CI) 1.09-2.04, P = 0.012; OR = 2.58, 95 % CI 1.65-4.05, P autophagy gene polymorphism was associated with the increased risk of leprosy by affecting inflammatory cytokines.

  11. Autophagy and Liver Ischemia-Reperfusion Injury

    Directory of Open Access Journals (Sweden)

    Raffaele Cursio

    2015-01-01

    Full Text Available Liver ischemia-reperfusion (I-R injury occurs during liver resection, liver transplantation, and hemorrhagic shock. The main mode of liver cell death after warm and/or cold liver I-R is necrosis, but other modes of cell death, as apoptosis and autophagy, are also involved. Autophagy is an intracellular self-digesting pathway responsible for removal of long-lived proteins, damaged organelles, and malformed proteins during biosynthesis by lysosomes. Autophagy is found in normal and diseased liver. Although depending on the type of ischemia, warm and/or cold, the dynamic process of liver I-R results mainly in adenosine triphosphate depletion and in production of reactive oxygen species (ROS, leads to both, a local ischemic insult and an acute inflammatory-mediated reperfusion injury, and results finally in cell death. This process can induce liver dysfunction and can increase patient morbidity and mortality after liver surgery and hemorrhagic shock. Whether autophagy protects from or promotes liver injury following warm and/or cold I-R remains to be elucidated. The present review aims to summarize the current knowledge in liver I-R injury focusing on both the beneficial and the detrimental effects of liver autophagy following warm and/or cold liver I-R.

  12. Autophagy and Autoimmunity CrossTalks

    Directory of Open Access Journals (Sweden)

    Abhisek eBhattacharya

    2013-04-01

    Full Text Available Autophagy, initially viewed as a conserved bulk-degradation mechanism, has emerged as a central player in a multitude of immune functions. Autophagy is important in host defense against intracellular and extracellular pathogens, metabolic syndromes, immune cell homeostasis, antigen processing and presentation and maintenance of tolerance. The observation that the above processes are implicated in triggering or exacerbating autoimmunity raises the possibility that the autophagy pathway is involved in mediating autoimmune processes, either directly or as a consequence of innate or adaptive functions mediated by the pathway. Genome-wide association studies have shown association between single nucleotide polymorphisms (SNPs in autophagy related gene 5 (Atg5, and Atg16l1 with susceptibility to systemic lupus erythematous (SLE and Crohn’s disease, respectively. Enhanced expression of Atg5 was also reported in blood of mice with experimental autoimmune encephalomyelitis (EAE, a mouse model of multiple sclerosis (MS, and in T cells isolated from blood or brain tissues from patients with active relapse of MS. This review explores the roles of autophagy pathway in the innate and adaptive immune systems on regulating or mediating the onset, progression or exacerbation of autoimmune processes.

  13. Mechanistic studies on activation of ubiquitin and di-ubiquitin-like protein, FAT10, by ubiquitin-like modifier activating enzyme 6, Uba6.

    Science.gov (United States)

    Gavin, James M; Chen, Jesse J; Liao, Hua; Rollins, Neil; Yang, Xiaofeng; Xu, Qing; Ma, Jingya; Loke, Huay-Keng; Lingaraj, Trupti; Brownell, James E; Mallender, William D; Gould, Alexandra E; Amidon, Benjamin S; Dick, Lawrence R

    2012-05-01

    Uba6 is a homolog of the ubiquitin-activating enzyme, Uba1, and activates two ubiquitin-like proteins (UBLs), ubiquitin and FAT10. In this study, biochemical and biophysical experiments were performed to understand the mechanisms of how Uba6 recognizes two distinct UBLs and catalyzes their activation and transfer. Uba6 is shown to undergo a three-step activation process and form a ternary complex with both UBLs, similar to what has been observed for Uba1. The catalytic mechanism of Uba6 is further supported by inhibition studies using a mechanism-based E1 inhibitor, Compound 1, which forms covalent adducts with both ubiquitin and FAT10. In addition, pre-steady state kinetic analysis revealed that the rates of UBL-adenylate (step 1) and thioester (step 2) formation are similar between ubiquitin and FAT10. However, distinct kinetic behaviors were also observed for ubiquitin and FAT10. FAT10 binds Uba6 with much higher affinity than ubiquitin while demonstrating lower catalytic activity in both ATP-PP(i) exchange and E1-E2 transthiolation assays. Also, Compound 1 is less potent with FAT10 as the UBL compared with ubiquitin in ATP-PP(i) exchange assays, and both a slow rate of covalent adduct formation and weak adduct binding to Uba6 contribute to the diminished potency observed for FAT10. Together with expression level analysis in IM-9 cells, this study sheds light on the potential role of cytokine-induced FAT10 expression in regulating Uba6 pathways.

  14. The molecular mechanism and significance of autophagy in immune response to mycobacterium tuberculosis%结核病免疫应答中自噬现象的分子机制和意义

    Institute of Scientific and Technical Information of China (English)

    黄丹丹; 鲍朗

    2012-01-01

    A growing body of evidence shows that autophagy is an essential component in the immune response to Mycobacterium tuberculosis (Mtb).Autophagy,a direct mechanism of killing intracellular Mycobacterium tuberculosis,acts as a modulator of proinflammatory cytokine secretion and plays a key role in antigen presentation.Autophagy coordinate with other host immune strategies against mycobacteria,including vitamin D-mediated innate immunity,ubiquitin-and intlammasome-involved pathways.On the other hand,Mtb has its strategies to modulate the autophagy in macrophage.Autophagy offers an attractive therapeutic target.Autophagy-promoting agents could have a clinical application as adjunctive treatment for drug-resistant tuberculosis.Moreover,vaccines which effectively induce autophagy could be more successful in providing better immune protection.%越来越多的证据表明,自噬是结核免疫反应的重要组成部分.自噬可以杀灭结核分枝杆菌、调节促炎细胞因子的分泌、增加抗原递呈功能.自噬与其他抗菌途径如维生素D3、炎性体、泛素系统存在协同作用.另一方面,结核分枝杆菌可以调控巨噬细胞的自噬.目前,自噬已成为临床重要的诊疗靶点.其能诱导自噬的药物,可以作为佐剂治疗耐药性结核;能有效诱导自噬的疫苗,可能提供更好的免疫保护作用.

  15. The ubiquitin proteasomal system: a potential target for the management of Alzheimer's disease.

    Science.gov (United States)

    Gadhave, Kundlik; Bolshette, Nityanand; Ahire, Ashutosh; Pardeshi, Rohit; Thakur, Krishan; Trandafir, Cristiana; Istrate, Alexandru; Ahmed, Sahabuddin; Lahkar, Mangala; Muresanu, Dafin F; Balea, Maria

    2016-07-01

    The cellular quality control system degrades abnormal or misfolded proteins and consists of three different mechanisms: the ubiquitin proteasomal system (UPS), autophagy and molecular chaperones. Any disturbance in this system causes proteins to accumulate, resulting in neurodegenerative diseases such as amyotrophic lateral sclerosis, Alzheimer's disease (AD), Parkinson's disease, Huntington's disease and prion or polyglutamine diseases. Alzheimer's disease is currently one of the most common age-related neurodegenerative diseases. However, its exact cause and pathogenesis are unknown. Currently approved medications for AD provide symptomatic relief; however, they fail to influence disease progression. Moreover, the components of the cellular quality control system represent an important focus for the development of targeted and potent therapies for managing AD. This review aims to evaluate whether existing evidence supports the hypothesis that UPS impairment causes the early pathogenesis of neurodegenerative disorders. The first part presents basic information about the UPS and its molecular components. The next part explains how the UPS is involved in neurodegenerative disorders. Finally, we emphasize how the UPS influences the management of AD. This review may help in the design of future UPS-related therapies for AD.

  16. Exercise induces autophagy in peripheral tissues and in the brain

    OpenAIRE

    He, Congcong; Sumpter, Jr., Rhea; Levine, Beth

    2012-01-01

    We recently identified physical exercise as a newly defined inducer of autophagy in vivo. Exercise induced autophagy in multiple organs involved in metabolic regulation, such as muscle, liver, pancreas and adipose tissue. To study the physiological role of exercise-induced autophagy, we generated mice with a knock-in nonphosphorylatable mutation in BCL2 (Thr69Ala, Ser70Ala and Ser84Ala) (BCL2 AAA) that are defective in exercise- and starvation-induced autophagy but not in basal autophagy. We ...

  17. Inflammation of the Penis

    Science.gov (United States)

    ... the Penis Medical Dictionary Additional Content Medical News Inflammation of the Penis By Patrick J. Shenot, MD ... Testicular Disorders Introduction to Penile and Testicular Disorders Inflammation of the Penis Phimosis and Paraphimosis Urethral Stricture ...

  18. Inflammation of the Orbit

    Science.gov (United States)

    ... Diagnosis Treatment Medical Dictionary Additional Content Medical News Inflammation of the Orbit (Inflammatory Orbital Pseudotumor) By James ... Introduction to Eye Socket Disorders Cavernous Sinus Thrombosis Inflammation of the Orbit Orbital Cellulitis Preseptal Cellulitis Tumors ...

  19. Idiopathic sclerosing orbital inflammation

    NARCIS (Netherlands)

    J.D. Hsuan; D. Selva; A.A. McNab; T.J. Sullivan; P. Saeed; B.A. O'Donnell

    2006-01-01

    Objective: To perform a multicenter review of the clinical features and treatment of 31 patients with idiopathic sclerosing orbital inflammation. Methods: We included all patients with histologically confirmed idiopathic sclerosing orbital inflammation from 5 regional orbital centers. We reviewed th

  20. MicroRNA regulation of Autophagy

    DEFF Research Database (Denmark)

    Frankel, Lisa B; Lund, Anders H

    2012-01-01

    Macroautophagy (hereafter referred to as autophagy) is a tightly regulated intracellular catabolic pathway involving the lysosomal degradation of cytoplasmic organelles and proteins. Central to this process is the formation of the autophagosome, a double membrane-bound vesicle, which is responsible...... for the delivery of cytoplasmic cargo to the lysosomes. Autophagy levels are constantly changing, allowing adaptation to both immediate and long-term needs of the cell, underlining why tight control of this process is essential in order to prevent the development of pathological disorders. Substantial progress has...... recently contributed to our understanding of the molecular mechanisms of the autophagy machinery, yet several gaps remain in our knowledge of this process. The discovery of microRNAs (miRNAs) established a new paradigm of post-transcriptional gene regulation and during the past decade these small non...

  1. What to Eat: Evidence for Selective Autophagy in Plants

    Institute of Scientific and Technical Information of China (English)

    Brice E.Floyd; Stephanie C.Morriss; Gustavo C.Maclntosh; Diane C.Bassham

    2012-01-01

    Autophagy is a macromolecular degradation pathway by which cells recycle their contents as a developmental process,house-keeping mechanism,and response to environmental stress.In plants,autophagy involves the sequestration of cargo to be degraded,transport to the cell vacuole in a double-membrane bound autophagosome,and subsequent degradation by lytic enzymes.Autophagy has generally been considered to be a non-selective mechanism of degradation.However,studies in yeast and animals have found numerous examples of selective autophagy,with cargo including proteins,protein aggregates,and organelles.Recent work has also provided evidence for several types of selective autophagy in plants.The degradation of protein aggregates was the first selective autophagy described in plants,and,more recently,a hybrid protein of the mammalian selective autophagy adaptors p62 and NBR1,which interacts with the autophagy machinery and may function in autophagy of protein aggregates,was described in plants.Other intracellular components have been suggested to be selectively targeted by autophagy in plants,but the current evidence is limited.Here,we discuss recent findings regarding the selective targeting of cell components by autophagy in plants.

  2. The Journal of Inflammation

    OpenAIRE

    Punchard Neville A; Whelan Cliff J; Adcock Ian

    2004-01-01

    Abstract Welcome to the Journal of Inflammation, the first open-access, peer-reviewed, online journal to focus on all aspects of the study of inflammation and inflammatory conditions. While research into inflammation has resulted in great progress in the latter half of the 20th century, the rate of progress is rapidly accelerating. Thus there is a need for a vehicle through which this very diverse research can be made readily available to the scientific community. The Journal of Inflammation,...

  3. Prostaglandins and chronic inflammation

    OpenAIRE

    Aoki, Tomohiro; Narumiya, Shuh

    2012-01-01

    Chronic inflammation is the basis of various chronic illnesses including cancer and vascular diseases. However, much has yet to be learned how inflammation becomes chronic. Prostaglandins (PGs) are well established as mediators of acute inflammation, and recent studies in experimental animals have provided evidence that they also function in transition to and maintenance of chronic inflammation. One role PGs play in such processes is amplification of cytokine signaling. As such, PGs can facil...

  4. Autophagy: A double-edged sword in intervertebral disk degeneration.

    Science.gov (United States)

    Zhang, Shu-Jun; Yang, Wei; Wang, Cheng; He, Wen-Si; Deng, Hai-Yang; Yan, Yi-Guo; Zhang, Jian; Xiang, Yong-Xiao; Wang, Wen-Jun

    2016-06-01

    Autophagy is a homeostatic mechanism through which intracellular damaged organelles and proteins are degraded and recycled in response to increased metabolic demands or stresses. Although primarily cytoprotective, dysfunction of autophagy is often associated with many degenerative diseases, including intervertebral disc (IVD) degeneration (IDD). As a main contributing factor to low back pain, IDD is the pathological basis for various debilitating spinal diseases. Either higher or lower levels of autophagy are observed in degenerative IVD cells. Despite the precise role of autophagy in disc degeneration that is still controversial, with difference from protection to aggravation, targeting autophagy has shown promise for mitigating disc degeneration. In the current review, we summarize the changes of autophagy in degenerative IVD cells and mainly discuss the relationship between autophagy and IDD. With continued efforts, modulation of the autophagic process could be a potential and attractive therapeutic strategy for degenerative disc disease. PMID:27018178

  5. Autophagy-associated immune responses and cancer immunotherapy

    Science.gov (United States)

    Xu, Yinghua; Han, Weidong; Lou, Fang; Fei, Weiqiang; Liu, Shuiping; Jing, Zhao; Sui, Xinbing

    2016-01-01

    Autophagy is an evolutionarily conserved catabolic process by which cellular components are sequestered into a double-membrane vesicle and delivered to the lysosome for terminal degradation and recycling. Accumulating evidence suggests that autophagy plays a critical role in cell survival, senescence and homeostasis, and its dysregulation is associated with a variety of diseases including cancer, cardiovascular disease, neurodegeneration. Recent studies show that autophagy is also an important regulator of cell immune response. However, the mechanism by which autophagy regulates tumor immune responses remains elusive. In this review, we will describe the role of autophagy in immune regulation and summarize the possible molecular mechanisms that are currently well documented in the ability of autophagy to control cell immune response. In addition, the scientific and clinical hurdles regarding the potential role of autophagy in cancer immunotherapy will be discussed. PMID:26788909

  6. HIV-1 differentially modulates autophagy in neurons and astrocytes.

    Science.gov (United States)

    Mehla, Rajeev; Chauhan, Ashok

    2015-08-15

    Autophagy, a lysosomal degradative pathway that maintains cellular homeostasis, has emerged as an innate immune defense against pathogens. The role of autophagy in the deregulated HIV-infected central nervous system (CNS) is unclear. We have found that HIV-1-induced neuro-glial (neurons and astrocytes) damage involves modulation of the autophagy pathway. Neuro-glial stress induced by HIV-1 led to biochemical and morphological dysfunctions. X4 HIV-1 produced neuro-glial toxicity coupled with suppression of autophagy, while R5 HIV-1-induced toxicity was restricted to neurons. Rapamycin, a specific mTOR inhibitor (autophagy inducer) relieved the blockage of the autophagy pathway caused by HIV-1 and resulted in neuro-glial protection. Further understanding of the regulation of autophagy by cytokines and chemokines or other signaling events may lead to recognition of therapeutic targets for neurodegenerative diseases.

  7. Autophagy-associated immune responses and cancer immunotherapy.

    Science.gov (United States)

    Pan, Hongming; Chen, Liuxi; Xu, Yinghua; Han, Weidong; Lou, Fang; Fei, Weiqiang; Liu, Shuiping; Jing, Zhao; Sui, Xinbing

    2016-04-19

    Autophagy is an evolutionarily conserved catabolic process by which cellular components are sequestered into a double-membrane vesicle and delivered to the lysosome for terminal degradation and recycling. Accumulating evidence suggests that autophagy plays a critical role in cell survival, senescence and homeostasis, and its dysregulation is associated with a variety of diseases including cancer, cardiovascular disease, neurodegeneration. Recent studies show that autophagy is also an important regulator of cell immune response. However, the mechanism by which autophagy regulates tumor immune responses remains elusive. In this review, we will describe the role of autophagy in immune regulation and summarize the possible molecular mechanisms that are currently well documented in the ability of autophagy to control cell immune response. In addition, the scientific and clinical hurdles regarding the potential role of autophagy in cancer immunotherapy will be discussed.

  8. Effect of inhibition of the Ubiquitin-Proteasome System and Hsp90 on growth and survival of Rhabdomyosarcoma cells in vitro

    Directory of Open Access Journals (Sweden)

    Peron Marica

    2012-06-01

    Full Text Available Abstract Background The ubiquitin-proteasome system (UPS and the heat shock response (HSR are two critical regulators of cell homeostasis, as their inhibition affects growth and survival of normal cells, as well as stress response and invasiveness of cancer cells. We evaluated the effects of the proteasome inhibitor Bortezomib and of 17-DMAG, a competitive inhibitor of Hsp90, in rhabdomyosarcoma (RMS cells, and analyzed the efficacy of single-agent exposures with combination treatments. Methods To assess cytotoxicity induced by Bortezomib and 17-DMAG in RMS cells, viability was measured by MTT assay after 24, 48 and 72 hours. Western blotting and immunofluorescence analyses were carried out to elucidate the mechanisms of action. Apoptosis was measured by FACS with Annexin-V-FITC and Propidium Iodide. Results Bortezomib and 17-DMAG, when combined at single low-toxic concentrations, enhanced growth inhibition of RMS cells, with signs of autophagy that included intensive cytoplasmic vacuolization and conversion of cytosolic LC3-I protein to its autophagosome-associated form. Treatment with lysosomal inhibitor chloroquine facilitates apoptosis, whereas stimulation of autophagy by rapamycin prevents LC3-I conversion and cell death, suggesting that autophagy is a resistance mechanism in RMS cells exposed to proteotoxic drugs. However, combination treatment also causes caspase-dependent apoptosis, PARP cleavage and Annexin V staining, as simultaneous inhibition of both UPS and HSR systems limits cytoprotective autophagy, exacerbating stress resulting from accumulation of misfolded proteins. Conclusion The combination of proteasome inhibitor Bortezomib with Hsp90 inhibitor 17-DMAG, appears to have important therapeutic advantages in the treatment of RMS cells compared with single-agent exposure, because compensatory survival mechanisms that occur as side effects of treatment may be prevented.

  9. Puromycin induces SUMO and ubiquitin redistribution upon proteasome inhibition.

    Science.gov (United States)

    Matsumoto, Hotaru; Saitoh, Hisato

    2016-07-29

    We have previously reported the co-localization of O-propargyl-puromycin (OP-Puro) with SUMO-2/3 and ubiquitin at promyelocytic leukemia-nuclear bodies (PML-NBs) in the presence of the proteasome inhibitor MG132, implying a role for the ubiquitin family in sequestering OP-puromycylated immature polypeptides to the nucleus during impaired proteasome activity. Here, we found that as expected puromycin induced SUMO-1/2/3 accumulation with ubiquitin at multiple nuclear foci in HeLa cells when co-exposed to MG132. Co-administration of puromycin and MG132 also facilitated redistribution of PML and the SUMO-targeted ubiquitin ligase RNF4 concurrently with SUMO-2/3. As removal of the drugs from the medium led to disappearance of the SUMO-2/3-ubiquitin nuclear foci, our findings indicated that nuclear assembly/disassembly of SUMO-2/3 and ubiquitin was pharmacologically manipulable, supporting our previous observation on OP-Puro, which predicted the ubiquitin family function in sequestrating aberrant proteins to the nucleus. PMID:27181354

  10. A cotranslational ubiquitination pathway for quality control of misfolded proteins.

    Science.gov (United States)

    Wang, Feng; Durfee, Larissa A; Huibregtse, Jon M

    2013-05-01

    Previous studies have indicated that 6%-30% of newly synthesized proteins are rapidly degraded by the ubiquitin-proteasome system; however, the relationship of ubiquitination to translation for these proteins has been unclear. We report that cotranslational ubiquitination (CTU) is a robust process, with 12%-15% of nascent polypeptides being ubiquitinated in human cells. CTU products contained primarily K48-linked polyubiquitin chains, consistent with a proteasomal targeting function. While nascent chains have been shown previously to be ubiquitinated within stalled complexes (CTU(S)), the majority of nascent chain ubiquitination occurred within active translation complexes (CTU(A)). CTU(A) was increased in response to agents that induce protein misfolding, while CTU(S) was increased in response to agents that lead to translational errors or stalling. These results indicate that ubiquitination of nascent polypeptides occurs in two contexts and define CTU(A) as a component of a quality control system that marks proteins for destruction while they are being synthesized. PMID:23583076

  11. Mcl-1 Ubiquitination: Unique Regulation of an Essential Survival Protein

    Directory of Open Access Journals (Sweden)

    Barbara Mojsa

    2014-05-01

    Full Text Available Mcl-1 is an anti-apoptotic protein of the Bcl-2 family that is essential for the survival of multiple cell lineages and that is highly amplified in human cancer. Under physiological conditions, Mcl-1 expression is tightly regulated at multiple levels, involving transcriptional, post-transcriptional and post-translational processes. Ubiquitination of Mcl-1, that targets it for proteasomal degradation, allows for rapid elimination of the protein and triggering of cell death, in response to various cellular events. In the last decade, a number of studies have elucidated different pathways controlling Mcl-1 ubiquitination and degradation. Four different E3 ubiquitin-ligases (e.g., Mule, SCFβ-TrCP, SCFFbw7 and Trim17 and one deubiquitinase (e.g., USP9X, that respectively mediate and oppose Mcl-1 ubiquitination, have been formerly identified. The interaction between Mule and Mcl-1 can be modulated by other Bcl-2 family proteins, while recognition of Mcl-1 by the other E3 ubiquitin-ligases and deubiquitinase is influenced by phosphorylation of specific residues in Mcl-1. The protein kinases and E3 ubiquitin-ligases that are involved in the regulation of Mcl-1 stability vary depending on the cellular context, highlighting the complexity and pivotal role of Mcl-1 regulation. In this review, we attempt to recapitulate progress in understanding Mcl-1 regulation by the ubiquitin-proteasome system.

  12. Enhanced detection of ubiquitin isopeptides using reductive methylation.

    Science.gov (United States)

    Chicooree, Navin; Connolly, Yvonne; Tan, Chong-Teik; Malliri, Angeliki; Li, Yaoyong; Smith, Duncan L; Griffiths, John R

    2013-03-01

    Identification of ubiquitination (Ub) sites is of great interest due to the critical roles that the modification plays in cellular regulation. Current methods using mass spectrometry rely upon tryptic isopeptide diglycine tag generation followed by database searching. We present a novel approach to ubiquitin detection based upon the dimethyl labeling of isopeptide N-termini glycines. Ubiquitinated proteins were digested with trypsin and the resulting peptide mixture was derivatized using formaldehyde-D2 solution and sodium cyanoborohydride. The dimethylated peptide mixtures were next separated by liquid chromatography and analyzed on a quadrupole-TOF based mass spectrometer. Diagnostic b2' and a1' ions released from the isopeptide N-terminus upon collision-induced dissociation (CID) were used to spectrally improve the identification of ubiquitinated isopeptides. Proof of principle was established by application to a ubiquitinated protein tryptic digest spiked into a six-protein mix digest background. Extracted ion chromatograms of the a1' and b2' diagnostic product ions from the diglycine tag resulted in a significant reduction in signal complexity and demonstrated a selectivity towards the identification of diglycine branched isopeptides. The method was further shown to be capable of identifying diglycine isopeptides resulting from in-gel tryptic digests of ubiquitin enriched material from a His-Ub transfected cell line. We envisage that these ions may be utilized in global ubiquitination studies with post-acquisition MS/MS (or MSe) data interrogation on high resolution hybrid mass spectrometers. ᅟ PMID:23361369

  13. Endotoxin-stimulated Rat Hepatic Stellate Cells Induce Autophagy in Hepatocytes as a Survival Mechanism.

    Science.gov (United States)

    Dangi, Anil; Huang, Chao; Tandon, Ashish; Stolz, Donna; Wu, Tong; Gandhi, Chandrashekhar R

    2016-01-01

    Bacterial lipopolysaccharide (LPS)-stimulated hepatic stellate cells (HSCs) produce many cytokines including IFNβ, TNFα, and IL6, strongly inhibit DNA synthesis, but induce apoptosis of a small number of hepatocytes. In vivo administration of LPS (up to 10 mg/mL) causes modest inflammation and weight loss in rats but not mortality. We determined whether LPS-stimulated HSCs instigate mechanisms of hepatocyte survival. Rats received 10 mg/kg LPS (i.p.) and determinations were made at 6 h. In vitro, HSCs were treated with 100 ng/mL LPS till 24 h. The medium was transferred to hepatocytes, and determinations were made at 0-12 h. Controls were HSC-conditioned medium or medium-containing LPS. LPS treatment of rats caused autophagy in hepatocytes, a physiological process for clearance of undesirable material including injured or damaged organelles. This was accompanied by activation of c-Jun NH2 terminal kinase (JNK) and apoptosis of ~4-5% of hepatocytes. In vitro, LPS-conditioned HSC medium (LPS/HSC) induced autophagy in hepatocytes but apoptosis of only ~10% of hepatocytes. While LPS/HSC stimulated activation of JNK (associated with cell death), it also activated NFkB and ERK1/2 (associated with cell survival). LPS-stimulated HSCs produced IFNβ, and LPS/HSC-induced autophagy in hepatocytes and their apoptosis were significantly inhibited by anti-IFNβ antibody. Blockade of autophagy, on the other hand, strongly augmented hepatocyte apoptosis. While LPS-stimulated HSCs cause apoptosis of a subpopulation of hepatocytes by producing IFNβ, they also induce cell survival mechanisms, which may be of critical importance in resistance to liver injury during endotoxemia.

  14. Skeletal muscle homeostasis in Duchenne muscular dystrophy: modulating autophagy as a promising therapeutic strategy

    Directory of Open Access Journals (Sweden)

    Clara eDe Palma

    2014-07-01

    Full Text Available Muscular dystrophies are a group of genetic and heterogeneous neuromuscular disorders characterised by the primary wasting of skeletal muscle. In Duchenne muscular dystrophy (DMD, the most severe form of these diseases, the mutations in the dystrophin gene lead to muscle weakness and wasting, exhaustion of muscular regenerative capacity and chronic local inflammation leading to substitution of myofibres by connective and adipose tissue. DMD patients suffer of continuous and progressive skeletal muscle damage followed by complete paralysis and death, usually by respiratory and/or cardiac failure. No cure is yet available, but several therapeutic approaches aiming at reversing the ongoing degeneration have been investigated in preclinical and clinical settings. The autophagy is an important proteolytic system of the cell and has a crucial role in the removal of proteins, aggregates and organelles. Autophagy is constantly active in skeletal muscle and its role in tissue homeostasis is complex: at high levels it can be detrimental and contribute to muscle wasting; at low levels it can cause weakness and muscle degeneration, due to the unchecked accumulation of damaged proteins and organelles. The causal relationship between DMD pathogenesis and dysfunctional autophagy has been recently investigated. At molecular levels, the Akt axis is one of the key disregulated pathways, although the molecular events are not completely understood.The aim of this review is to describe and discuss the clinical relevance of the recent advances dissecting autophagy and its signalling pathway in DMD. The picture might pave the way for the development of interventions that are able to boost muscle growth and/or prevent muscle wasting.

  15. Autophagy in the light of sphingolipid metabolism

    DEFF Research Database (Denmark)

    Harvald, Eva Bang; Olsen, Anne Sofie Braun; Færgeman, Nils J.

    2015-01-01

    , has over the past decade been recognized as an essential part of metabolism. Autophagy not only rids the cell of excessive or damaged organelles, misfolded proteins, and invading microorganisms, it also provides nutrients to maintain crucial cellular functions. Besides serving as essential structural......Maintenance of cellular homeostasis requires tight and coordinated control of numerous metabolic pathways, which are governed by interconnected networks of signaling pathways and energy-sensing regulators. Autophagy, a lysosomal degradation pathway by which the cell self-digests its own components...

  16. Autophagy and proteins involved in vesicular trafficking.

    Science.gov (United States)

    Amaya, Celina; Fader, Claudio Marcelo; Colombo, María Isabel

    2015-11-14

    Autophagy is an intracellular degradation system that, as a basic mechanism it delivers cytoplasmic components to the lysosomes in order to maintain adequate energy levels and cellular homeostasis. This complex cellular process is activated by low cellular nutrient levels and other stress situations such as low ATP levels, the accumulation of damaged proteins or organelles, or pathogen invasion. Autophagy as a multistep process involves vesicular transport events leading to tethering and fusion of autophagic vesicles with several intracellular compartments. This review summarizes our current understanding of the autophagic pathway with emphasis in the trafficking machinery (i.e. Rabs GTPases and SNAP receptors (SNAREs)) involved in specific steps of the pathway.

  17. Water Evaporation and Conformational Changes from Partially Solvated Ubiquitin

    Directory of Open Access Journals (Sweden)

    Saravana Prakash Thirumuruganandham

    2010-01-01

    Full Text Available Using molecular dynamics simulation, we study the evaporation of water molecules off partially solvated ubiquitin. The evaporation and cooling rates are determined for a molecule at the initial temperature of 300 K. The cooling rate is found to be around 3 K/ns, and decreases with water temperature in the course of the evaporation. The conformation changes are monitored by studying a variety of intermediate partially solvated ubiquitin structures. We find that ubiquitin shrinks with decreasing hydration shell and exposes more of its hydrophilic surface area to the surrounding.

  18. LC3B is indispensable for selective autophagy of p62 but not basal autophagy

    Energy Technology Data Exchange (ETDEWEB)

    Maruyama, Yoko [Protein Metabolism Project, Tokyo Metropolitan Institute of Medical Science, Tokyo 156-8506 (Japan); Department of Pediatrics, School of Medicine, Keio University, Tokyo 160-8582 (Japan); Sou, Yu-Shin; Kageyama, Shun [Protein Metabolism Project, Tokyo Metropolitan Institute of Medical Science, Tokyo 156-8506 (Japan); Takahashi, Takao [Department of Pediatrics, School of Medicine, Keio University, Tokyo 160-8582 (Japan); Ueno, Takashi [Division of Proteomics and Biomolecular Science, Center for Biomedical Research Resources, Juntendo University Graduate School of Medicine, Tokyo 113-8421 (Japan); Tanaka, Keiji [Laboratory of Protein Metabolism, Tokyo Metropolitan Institute of Medical Science, Tokyo 156-8506 (Japan); Komatsu, Masaaki, E-mail: komatsu-ms@igakuken.or.jp [Protein Metabolism Project, Tokyo Metropolitan Institute of Medical Science, Tokyo 156-8506 (Japan); Department of Biochemistry, School of Medicine, Niigata University, Niigata 951-8510 (Japan); Ichimura, Yoshinobu, E-mail: ichimura-ys@igakuken.or.jp [Protein Metabolism Project, Tokyo Metropolitan Institute of Medical Science, Tokyo 156-8506 (Japan)

    2014-03-28

    Highlights: • Knockdown of LC3 or GABARAP families did not affect the basal autophagy. • LC3B has a higher affinity for the autophagy-specific substrate, p62, than GABARAPs. • siRNA-mediated knockdown of LC3B, but not that of GABARAPs, resulted in significant accumulation of p62. - Abstract: Autophagy is a unique intracellular protein degradation system accompanied by autophagosome formation. Besides its important role through bulk degradation in supplying nutrients, this system has an ability to degrade certain proteins, organelles, and invading bacteria selectively to maintain cellular homeostasis. In yeasts, Atg8p plays key roles in both autophagosome formation and selective autophagy based on its membrane fusion property and interaction with autophagy adaptors/specific substrates. In contrast to the single Atg8p in yeast, mammals have 6 homologs of Atg8p comprising LC3 and GABARAP families. However, it is not clear these two families have different or similar functions. The aim of this study was to determine the separate roles of LC3 and GABARAP families in basal/constitutive and/or selective autophagy. While the combined knockdown of LC3 and GABARAP families caused a defect in long-lived protein degradation through lysosomes, knockdown of each had no effect on the degradation. Meanwhile, knockdown of LC3B but not GABARAPs resulted in significant accumulation of p62/Sqstm1, one of the selective substrate for autophagy. Our results suggest that while mammalian Atg8 homologs are functionally redundant with regard to autophagosome formation, selective autophagy is regulated by specific Atg8 homologs.

  19. Extracts of Feijoa Inhibit Toll-Like Receptor 2 Signaling and Activate Autophagy Implicating a Role in Dietary Control of IBD.

    Directory of Open Access Journals (Sweden)

    Noha Ahmed Nasef

    Full Text Available Inflammatory bowel disease (IBD is a heterogeneous chronic inflammatory disease affecting the gut with limited treatment success for its sufferers. This suggests the need for better understanding of the different subtypes of the disease as well as nutritional interventions to compliment current treatments. In this study we assess the ability of a hydrophilic feijoa fraction (F3 to modulate autophagy a process known to regulate inflammation, via TLR2 using IBD cell lines.Mouse embryonic fibroblasts (MEF deleted for ATG5, and two intestinal epithelial cells HCT15 and HCT116, were used to test the anti-inflammatory effect of F3 after stimulating the cells with a TLR2 specific ligand PAM3CSK4.F3 was able to reduce TLR2 specific inflammation and stimulate autophagy in MEFs and HCT15 cells but not in HCT116 cells. The anti-inflammatory effect was reduced in the MEF cells deleted for ATG5. In addition, the activation of autophagy by F3 was enhanced by PAM3CSK4.F3 of feijoa can interact with cells via a TLR2 specific mechanism and reduce Nuclear factor kappa B (NF-κB activation in part due to stimulation of autophagy. These results suggest that there is potential benefit in using feijoa extracts as part of dietary interventions to manage IBD in patients.

  20. 细胞自噬与肿瘤的研究进展%Research progress of autophagy and tumor

    Institute of Scientific and Technical Information of China (English)

    徐俊杰; 王志莲; 郝敏

    2016-01-01

    细胞自噬是细胞在某些刺激因素下,细胞内膜性结构包绕损伤的蛋白或细胞器,并通过溶酶体融合降解损伤的蛋白质或细胞器的过程;生理状况下的自噬,对于维持细胞稳态有积极作用;肿瘤形成以后,细胞自噬反为癌细胞提供更多的营养,起到促进肿瘤发展的功能。自噬主要有三种方式,即分子伴侣介导的自噬、微自噬和巨自噬,均通过两个泛素化途径实现。细胞自噬可以通过影响 DNA 损伤修复、细胞应激反应、肿瘤基因表达、凋亡及病毒感染反应等诸多过程参与细胞癌变,也可能在肿瘤治疗过程中降低了对于化疗和放疗的敏感性,但是在肿瘤进展的不同时期,不同肿瘤之间自噬活性的变化不一。自噬过程中的重要中间产物或关键酶可能成为抗肿瘤药物的靶标,拓展了肿瘤防治思路。%Cell autophagy is one of the important process of the material flow in the cell in the process of some of the damaged proteins and organelles by double membrane structure of autophagy vesicles package, then, to the lysosome degradation and recycling. Autophagy has a positive role for the maintenance of cell homeostasis. After tumor formation, cell autophagy provides more nutrition for cancer cells, which plays a role in promoting tumor. There are three main ways of autophagy, that are, the molecular chaperone mediated autophagy, microautophagy and macroautophagy, which are achieved by the two ubiquitin pathways. Autophagy may be involved in the cellular carcinogenesis by affecting the DNA damage repair, cell stress response, tumor gene expression, apoptosis, and viral infection. But in different stages of the tumor, autophagy may also reduce the sensitivity of chemotherapy and radiotherapy in the treatment of cancer, the changes of the activity of the different tumors are different from each other. Important intermediate products or key enzymes in the process of

  1. Group IVA phospholipase A(2) deficiency prevents CCl4-induced hepatic cell death through the enhancement of autophagy.

    Science.gov (United States)

    Ishihara, Keiichi; Kanai, Shiho; Tanaka, Kikuko; Kawashita, Eri; Akiba, Satoshi

    2016-02-26

    Group IVA phospholipase A2 (IVA-PLA2), which generates arachidonate, plays a role in inflammation. IVA-PLA2-deficiency reduced hepatotoxicity and hepatocyte cell death in mice that received a single dose of carbon tetrachloride (CCl4) without any inhibitory effects on CCl4-induced lipid peroxidation. An immunoblot analysis of extracts from wild-type mouse- and IVA-PLA2 KO mouse-derived primary hepatocytes that transiently expressed microtubule-associated protein 1 light chain 3B (LC3) revealed a higher amount of LC3-II, a typical index of autophagosome formation, in IVA-PLA2-deficient cells, suggesting the enhancement of constitutive autophagy. IVA-PLA2 may promote CCl4-induced cell death through the suppression of constitutive autophagy in hepatocytes.

  2. Autophagy as a Potential Target for Sarcopenia.

    Science.gov (United States)

    Fan, Jingjing; Kou, Xianjuan; Jia, Shaohui; Yang, Xiaoqi; Yang, Yi; Chen, Ning

    2016-07-01

    Sarcopenia is an aging-related disease with a significant reduction in mass and strength of skeletal muscle due to the imbalance between protein synthesis and protein degradation. The loss of skeletal muscle is an inevitable event during aging process, which can result in the significant impact on the quality of life, and also can increase the risk for other aging-associated diseases in the elderly. However, the underlying molecular mechanism of aging-related skeletal muscle loss is still poorly understood. Autophagy is a degradation pathway for the clearance of dysfunctional organelles and damaged macromolecules during aging process. Appropriate induction or accurate regulation of autophagic process and improved quality control of mitochondria through autophagy or other strategies are required for the maintenance of skeletal muscle mass. In this article, we have summarized the current understanding of autophagic pathways in sarcopenia, and discussed the functional status of autophagy and autophagy-associated quality control of mitochondria in the pathogenesis of sarcopenia. Moreover, this article will provide some theoretical references for the exploration of scientific and optimal intervention strategies such as exercise and caloric restriction for the prevention and treatment of sarcopenia through the regulation of autophagic pathways. PMID:26580995

  3. Impairment of autophagy: from hereditary disorder to drug intoxication.

    Science.gov (United States)

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

    2013-09-15

    At first, the molecular mechanism of autophagy was unveiled in a unicellular organism Saccharomyces cerevisiae (budding yeast), followed by the discovery that the basic mechanism of autophagy is conserved in multicellular organisms including mammals. Although autophagy was considered to be a non-selective bulk protein degradation system to recycle amino acids during periods of nutrient starvation, it is also believed to be an essential mechanism for the selective elimination of proteins/organelles that are damaged under pathological conditions. Research advances made using autophagy-deficient animals have revealed that impairments of autophagy often underlie the pathogenesis of hereditary disorders such as Danon, Parkinson's, Alzheimer's, and Huntington's diseases, and amyotrophic lateral sclerosis. On the other hand, there are many reports that drugs and toxicants, including arsenic, cadmium, paraquat, methamphetamine, and ethanol, induce autophagy during the development of their toxicity on many organs including heart, brain, lung, kidney, and liver. Although the question as to whether autophagic machinery is involved in the execution of cell death or not remains controversial, the current view of the role of autophagy during cell/tissue injury is that it is an important, often essential, cytoprotective reaction; disturbances in cytoprotective autophagy aggravate cell/tissue injuries. The purpose of this review is to provide (1) a gross summarization of autophagy processes, which are becoming more important in the field of toxicology, and (2) examples of important studies reporting the involvement of perturbations in autophagy in cell/tissue injuries caused by acute as well as chronic intoxication. PMID:23851159

  4. Impairment of autophagy: From hereditary disorder to drug intoxication

    International Nuclear Information System (INIS)

    At first, the molecular mechanism of autophagy was unveiled in a unicellular organism Saccharomyces cerevisiae (budding yeast), followed by the discovery that the basic mechanism of autophagy is conserved in multicellular organisms including mammals. Although autophagy was considered to be a non-selective bulk protein degradation system to recycle amino acids during periods of nutrient starvation, it is also believed to be an essential mechanism for the selective elimination of proteins/organelles that are damaged under pathological conditions. Research advances made using autophagy-deficient animals have revealed that impairments of autophagy often underlie the pathogenesis of hereditary disorders such as Danon, Parkinson's, Alzheimer's, and Huntington's diseases, and amyotrophic lateral sclerosis. On the other hand, there are many reports that drugs and toxicants, including arsenic, cadmium, paraquat, methamphetamine, and ethanol, induce autophagy during the development of their toxicity on many organs including heart, brain, lung, kidney, and liver. Although the question as to whether autophagic machinery is involved in the execution of cell death or not remains controversial, the current view of the role of autophagy during cell/tissue injury is that it is an important, often essential, cytoprotective reaction; disturbances in cytoprotective autophagy aggravate cell/tissue injuries. The purpose of this review is to provide (1) a gross summarization of autophagy processes, which are becoming more important in the field of toxicology, and (2) examples of important studies reporting the involvement of perturbations in autophagy in cell/tissue injuries caused by acute as well as chronic intoxication

  5. Liver Autophagy in Anorexia Nervosa and Acute Liver Injury

    Directory of Open Access Journals (Sweden)

    Marouane Kheloufi

    2014-01-01

    Full Text Available Autophagy, a lysosomal catabolic pathway for long-lived proteins and damaged organelles, is crucial for cell homeostasis, and survival under stressful conditions. During starvation, autophagy is induced in numerous organisms ranging from yeast to mammals, and promotes survival by supplying nutrients and energy. In the early neonatal period, when transplacental nutrients supply is interrupted, starvation-induced autophagy is crucial for neonates’ survival. In adult animals, autophagy provides amino acids and participates in glucose metabolism following starvation. In patients with anorexia nervosa, autophagy appears initially protective, allowing cells to copes with nutrient deprivation. However, when starvation is critically prolonged and when body mass index reaches 13 kg/m2 or lower, acute liver insufficiency occurs with features of autophagic cell death, which can be observed by electron microscopy analysis of liver biopsy samples. In acetaminophen overdose, a classic cause of severe liver injury, autophagy is induced as a protective mechanism. Pharmacological enhancement of autophagy protects against acetaminophen-induced necrosis. Autophagy is also activated as a rescue mechanism in response to Efavirenz-induced mitochondrial dysfunction. However, Efavirenz overdose blocks autophagy leading to liver cell death. In conclusion, in acute liver injury, autophagy appears as a protective mechanism that can be however blocked or overwhelmed.

  6. Autophagy modulates articular cartilage vesicle formation in primary articular chondrocytes.

    Science.gov (United States)

    Rosenthal, Ann K; Gohr, Claudia M; Mitton-Fitzgerald, Elizabeth; Grewal, Rupinder; Ninomiya, James; Coyne, Carolyn B; Jackson, William T

    2015-05-22

    Chondrocyte-derived extracellular organelles known as articular cartilage vesicles (ACVs) participate in non-classical protein secretion, intercellular communication, and pathologic calcification. Factors affecting ACV formation and release remain poorly characterized; although in some cell types, the generation of extracellular vesicles is associated with up-regulation of autophagy. We sought to determine the role of autophagy in ACV production by primary articular chondrocytes. Using an innovative dynamic model with a light scatter nanoparticle counting apparatus, we determined the effects of autophagy modulators on ACV number and content in conditioned medium from normal adult porcine and human osteoarthritic chondrocytes. Healthy articular chondrocytes release ACVs into conditioned medium and show significant levels of ongoing autophagy. Rapamycin, which promotes autophagy, increased ACV numbers in a dose- and time-dependent manner associated with increased levels of autophagy markers and autophagosome formation. These effects were suppressed by pharmacologic autophagy inhibitors and short interfering RNA for ATG5. Caspase-3 inhibition and a Rho/ROCK inhibitor prevented rapamycin-induced increases in ACV number. Osteoarthritic chondrocytes, which are deficient in autophagy, did not increase ACV number in response to rapamycin. SMER28, which induces autophagy via an mTOR-independent mechanism, also increased ACV number. ACVs induced under all conditions had similar ecto-enzyme specific activities and types of RNA, and all ACVs contained LC3, an autophagosome-resident protein. These findings identify autophagy as a critical participant in ACV formation, and augment our understanding of ACVs in cartilage disease and repair.

  7. Crosstalk of clock gene expression and autophagy in aging

    Science.gov (United States)

    Kalfalah, Faiza; Janke, Linda; Schiavi, Alfonso; Tigges, Julia; Ix, Alexander; Ventura, Natascia; Boege, Fritz; Reinke, Hans

    2016-01-01

    Autophagy and the circadian clock counteract tissue degeneration and support longevity in many organisms. Accumulating evidence indicates that aging compromises both the circadian clock and autophagy but the mechanisms involved are unknown. Here we show that the expression levels of transcriptional repressor components of the circadian oscillator, most prominently the human Period homologue PER2, are strongly reduced in primary dermal fibroblasts from aged humans, while raising the expression of PER2 in the same cells partially restores diminished autophagy levels. The link between clock gene expression and autophagy is corroborated by the finding that the circadian clock drives cell-autonomous, rhythmic autophagy levels in immortalized murine fibroblasts, and that siRNA-mediated downregulation of PER2 decreases autophagy levels while leaving core clock oscillations intact. Moreover, the Period homologue lin-42 regulates autophagy and life span in the nematode Caenorhabditis elegans, suggesting an evolutionarily conserved role for Period proteins in autophagy control and aging. Taken together, this study identifies circadian clock proteins as set-point regulators of autophagy and puts forward a model, in which age-related changes of clock gene expression promote declining autophagy levels. PMID:27574892

  8. A role for PCNA ubiquitination in immunoglobulin hypermutation.

    Directory of Open Access Journals (Sweden)

    Hiroshi Arakawa

    2006-11-01

    Full Text Available Proliferating cell nuclear antigen (PCNA is a DNA polymerase cofactor and regulator of replication-linked functions. Upon DNA damage, yeast and vertebrate PCNA is modified at the conserved lysine K164 by ubiquitin, which mediates error-prone replication across lesions via translesion polymerases. We investigated the role of PCNA ubiquitination in variants of the DT40 B cell line that are mutant in K164 of PCNA or in Rad18, which is involved in PCNA ubiquitination. Remarkably, the PCNA(K164R mutation not only renders cells sensitive to DNA-damaging agents, but also strongly reduces activation induced deaminase-dependent single-nucleotide substitutions in the immunoglobulin light-chain locus. This is the first evidence, to our knowledge, that vertebrates exploit the PCNA-ubiquitin pathway for immunoglobulin hypermutation, most likely through the recruitment of error-prone DNA polymerases.

  9. Substrate specificity of the ubiquitin and Ubl proteases

    Science.gov (United States)

    Ronau, Judith A; Beckmann, John F; Hochstrasser, Mark

    2016-01-01

    Conjugation and deconjugation of ubiquitin and ubiquitin-like proteins (Ubls) to cellular proteins are highly regulated processes integral to cellular homeostasis. Most often, the C-termini of these small polypeptides are attached to lysine side chains of target proteins by an amide (isopeptide) linkage. Deubiquitinating enzymes (DUBs) and Ubl-specific proteases (ULPs) comprise a diverse group of proteases that recognize and remove ubiquitin and Ubls from their substrates. How DUBs and ULPs distinguish among different modifiers, or different polymeric forms of these modifiers, remains poorly understood. The specificity of ubiquitin/Ubl-deconjugating enzymes for particular substrates depends on multiple factors, ranging from the topography of specific substrate features, as in different polyubiquitin chain types, to structural elements unique to each enzyme. Here we summarize recent structural and biochemical studies that provide insights into mechanisms of substrate specificity among various DUBs and ULPs. We also discuss the unexpected specificities of non-eukaryotic proteases in these families. PMID:27012468

  10. Autophagy-related prognostic signature for breast cancer.

    Science.gov (United States)

    Gu, Yunyan; Li, Pengfei; Peng, Fuduan; Zhang, Mengmeng; Zhang, Yuanyuan; Liang, Haihai; Zhao, Wenyuan; Qi, Lishuang; Wang, Hongwei; Wang, Chenguang; Guo, Zheng

    2016-03-01

    Autophagy is a process that degrades intracellular constituents, such as long-lived or damaged proteins and organelles, to buffer metabolic stress under starvation conditions. Deregulation of autophagy is involved in the progression of cancer. However, the predictive value of autophagy for breast cancer prognosis remains unclear. First, based on gene expression profiling, we found that autophagy genes were implicated in breast cancer. Then, using the Cox proportional hazard regression model, we detected autophagy prognostic signature for breast cancer in a training dataset. We identified a set of eight autophagy genes (BCL2, BIRC5, EIF4EBP1, ERO1L, FOS, GAPDH, ITPR1 and VEGFA) that were significantly associated with overall survival in breast cancer. The eight autophagy genes were assigned as a autophagy-related prognostic signature for breast cancer. Based on the autophagy-related signature, the training dataset GSE21653 could be classified into high-risk and low-risk subgroups with significantly different survival times (HR = 2.72, 95% CI = (1.91, 3.87); P = 1.37 × 10(-5)). Inactivation of autophagy was associated with shortened survival of breast cancer patients. The prognostic value of the autophagy-related signature was confirmed in the testing dataset GSE3494 (HR = 2.12, 95% CI = (1.48, 3.03); P = 1.65 × 10(-3)) and GSE7390 (HR = 1.76, 95% CI = (1.22, 2.54); P = 9.95 × 10(-4)). Further analysis revealed that the prognostic value of the autophagy signature was independent of known clinical prognostic factors, including age, tumor size, grade, estrogen receptor status, progesterone receptor status, ERBB2 status, lymph node status and TP53 mutation status. Finally, we demonstrated that the autophagy signature could also predict distant metastasis-free survival for breast cancer.

  11. Induction of cytoprotective autophagy in PC-12 cells by cadmium

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Qiwen [College of Veterinary Medicine, Yangzhou University, Yangzhou 225009 (China); Jiangsu Co-innovation Center for Prevention and Control of Important Animal Infectious Diseases and Zoonoses, Yangzhou 225009 (China); Bijie Pilot Area Research Institute of Bijie University, Bijie 551700 (China); Zhu, Jiaqiao; Zhang, Kangbao; Jiang, Chenyang; Wang, Yi; Yuan, Yan; Bian, Jianchun; Liu, Xuezhong; Gu, Jianhong [College of Veterinary Medicine, Yangzhou University, Yangzhou 225009 (China); Jiangsu Co-innovation Center for Prevention and Control of Important Animal Infectious Diseases and Zoonoses, Yangzhou 225009 (China); Liu, Zongping, E-mail: liuzongping@yzu.edu.cn [College of Veterinary Medicine, Yangzhou University, Yangzhou 225009 (China); Jiangsu Co-innovation Center for Prevention and Control of Important Animal Infectious Diseases and Zoonoses, Yangzhou 225009 (China)

    2013-08-16

    Highlights: •Cadmium can promote early upregulation of autophagy in PC-12 cells. •Autophagy precedes apoptosis in cadmium-treated PC-12 cells. •Cadmium-induced autophagy is cytoprotective in PC-12 cells. •Class III PI3K/beclin-1/Bcl-2 signaling pathway plays a positive role in cadmium-triggered autophagy. -- Abstract: Laboratory data have demonstrated that cadmium (Cd) may induce neuronal apoptosis. However, little is known about the role of autophagy in neurons. In this study, cell viability decreased in a dose- and time-dependent manner after treatment with Cd in PC-12 cells. As cells were exposed to Cd, the levels of LC3-II proteins became elevated, specific punctate distribution of endogenous LC3-II increased, and numerous autophagosomes appeared, which suggest that Cd induced a high level of autophagy. In the late stages of autophagy, an increase in the apoptosis ratio was observed. Likewise, pre-treatment with chloroquine (an autophagic inhibitor) and rapamycin (an autophagic inducer) resulted in an increased and decreased percentage of apoptosis in contrast to other Cd-treated groups, respectively. The results indicate that autophagy delayed apoptosis in Cd-treated PC-12 cells. Furthermore, co-treatment of cells with chloroquine reduced autophagy and cell activity. However, rapamycin had an opposite effect on autophagy and cell activity. Moreover, class III PI3 K/beclin-1/Bcl-2 signaling pathways served a function in Cd-induced autophagy. The findings suggest that Cd can induce cytoprotective autophagy by activating class III PI3 K/beclin-1/Bcl-2 signaling pathways. In sum, this study strongly suggests that autophagy may serve a positive function in the reduction of Cd-induced cytotoxicity.

  12. Multiple roles of the cytoskeleton in autophagy.

    Science.gov (United States)

    Monastyrska, Iryna; Rieter, Ester; Klionsky, Daniel J; Reggiori, Fulvio

    2009-08-01

    Autophagy is involved in a wide range of physiological processes including cellular remodeling during development, immuno-protection against heterologous invaders and elimination of aberrant or obsolete cellular structures. This conserved degradation pathway also plays a key role in maintaining intracellular nutritional homeostasis and during starvation, for example, it is involved in the recycling of unnecessary cellular components to compensate for the limitation of nutrients. Autophagy is characterized by specific membrane rearrangements that culminate with the formation of large cytosolic double-membrane vesicles called autophagosomes. Autophagosomes sequester cytoplasmic material that is destined for degradation. Once completed, these vesicles dock and fuse with endosomes and/or lysosomes to deliver their contents into the hydrolytically active lumen of the latter organelle where, together with their cargoes, they are broken down into their basic components. Specific structures destined for degradation via autophagy are in many cases selectively targeted and sequestered into autophagosomes. A number of factors required for autophagy have been identified, but numerous questions about the molecular mechanism of this pathway remain unanswered. For instance, it is unclear how membranes are recruited and assembled into autophagosomes. In addition, once completed, these vesicles are transported to cellular locations where endosomes and lysosomes are concentrated. The mechanism employed for this directed movement is not well understood. The cellular cytoskeleton is a large, highly dynamic cellular scaffold that has a crucial role in multiple processes, several of which involve membrane rearrangements and vesicle-mediated events. Relatively little is known about the roles of the cytoskeleton network in autophagy. Nevertheless, some recent studies have revealed the importance of cytoskeletal elements such as actin microfilaments and microtubules in specific aspects of

  13. The thiazole derivative CPTH6 impairs autophagy.

    Science.gov (United States)

    Ragazzoni, Y; Desideri, M; Gabellini, C; De Luca, T; Carradori, S; Secci, D; Nescatelli, R; Candiloro, A; Condello, M; Meschini, S; Del Bufalo, D; Trisciuoglio, D

    2013-01-01

    We have previously demonstrated that the thiazole derivative 3-methylcyclopentylidene-[4-(4'-chlorophenyl)thiazol-2-yl]hydrazone (CPTH6) induces apoptosis and cell cycle arrest in human leukemia cells. The aim of this study was to evaluate whether CPTH6 is able to affect autophagy. By using several human tumor cell lines with different origins we demonstrated that CPTH6 treatment induced, in a dose-dependent manner, a significant increase in autophagic features, as imaged by electron microscopy, immunoblotting analysis of membrane-bound form of microtubule-associated protein 1 light chain 3 (LC3B-II) levels and by appearance of typical LC3B-II-associated autophagosomal puncta. To gain insights into the molecular mechanisms of elevated markers of autophagy induced by CPTH6 treatment, we silenced the expression of several proteins acting at different steps of autophagy. We found that the effect of CPTH6 on autophagy developed through a noncanonical mechanism that did not require beclin-1-dependent nucleation, but involved Atg-7-mediated elongation of autophagosomal membranes. Strikingly, a combined treatment of CPTH6 with late-stage autophagy inhibitors, such as chloroquine and bafilomycin A1, demonstrates that under basal condition CPTH6 reduces autophagosome turnover through an impairment of their degradation pathway, rather than enhancing autophagosome formation, as confirmed by immunofluorescence experiments. According to these results, CPTH6-induced enhancement of autophagy substrate p62 and NBR1 protein levels confirms a blockage of autophagic cargo degradation. In addition, CPTH6 inhibited autophagosome maturation and compounds having high structural similarities with CPTH6 produced similar effects on the autophagic pathway. Finally, the evidence that CPTH6 treatment decreased α-tubulin acetylation and failed to increase autophagic markers in cells in which acetyltransferase ATAT1 expression was silenced indicates a possible role of α-tubulin acetylation in

  14. The role of hypoxia in intestinal inflammation.

    Science.gov (United States)

    Shah, Yatrik M

    2016-12-01

    Inflammatory bowel disease (IBD) is a chronic relapsing inflammatory disease of the intestine. IBD is a multifactorial disorder, and IBD-associated genes are critical in innate immune response, inflammatory response, autophagy, and epithelial barrier integrity. Moreover, epithelial oxygen tension plays a critical role in intestinal inflammation and resolution in IBD. The intestines have a dynamic and rapid fluctuation in cellular oxygen tension, which is dysregulated in IBD. Intestinal epithelial cells have a steep oxygen gradient where the tips of the villi are hypoxic and the oxygenation increases at the base of the villi. IBD results in heightened hypoxia throughout the mucosa. Hypoxia signals through a well-conserved family of transcription factors, where hypoxia-inducible factor (HIF)-1α and HIF-2α are essential in maintaining intestinal homeostasis. In inflamed mucosa, HIF-1α increases barrier protective genes, elicits protective innate immune responses, and activates an antimicrobial response through the increase in β-defensins. HIF-2α is essential in maintaining an epithelial-elicited inflammatory response and the regenerative and proliferative capacity of the intestine following an acute injury. HIF-1α activation in colitis leads to a protective response, whereas chronic activation of HIF-2α increases the pro-inflammatory response, intestinal injury, and cancer. In this mini-review, we detail the role of HIF-1α and HIF-2α in intestinal inflammation and injury and therapeutic implications of targeting HIF signaling in IBD. PMID:26812949

  15. The possible role of the ubiquitin proteasome system in the development of atherosclerosis in diabetes

    Directory of Open Access Journals (Sweden)

    sasso Ferdinando

    2007-10-01

    Full Text Available Abstract We have reviewed the impact of the ubiquitin proteasome system (UPS on atherosclerosis progression of diabetic patients. A puzzle of many pieces of evidence suggests that UPS, in addition to its role in the removal of damaged proteins, is involved in a number of biological processes including inflammation, proliferation and apoptosis, all of which constitute important characteristics of atherosclerosis. From what can be gathered from the very few studies on the UPS in diabetic cardiovascular diseases published so far, the system seems to be functionally active to a different extent in the initiation, progression, and complication stage of atherosclerosis in the diabetic people. Further evidence for this theory, however, has to be given, for instance by specifically targeted antagonism of the UPS. Nonetheless, this hypothesis may help us understand why diverse therapeutic interventions, which have in common the ability to reduce ubiquitin-proteasome activity, can impede or delay the onset of diabetes and cardiovascular diseases (CVD. People with type 2 diabetes are disproportionately affected by CVD, compared with those without diabetes 1. The prevalence, incidence, and mortality from all forms of CVD (myocardial infarction, cerebro-vascular disease and congestive heart failure are strikingly increased in persons with diabetes compared with those withoutdiabetes 2. Furthermore, diabetic patients have not benefited by the advances in the management of obesity, dyslipidemia, and hypertension that have resulted in a decrease in mortality for coronary heart disease (CHD patients without diabetes 3. Nevertheless, these risk factors do not fully explain the excess risk for CHD associated with diabetes 45. Thus, the determinants of progression of atherosclerosis in persons with diabetes must be elucidated. Beyond the major risk factors, several studies have demonstrated that such factors, strictly related to diabetes, as insulin

  16. Ubiquitin Metabolism Affects Cellular Response to Volatile Anesthetics in Yeast

    OpenAIRE

    Wolfe, Darren; Reiner, Thomas; Keeley, Jessica L.; Pizzini, Mark; Keil, Ralph L.

    1999-01-01

    To investigate the mechanism of action of volatile anesthetics, we are studying mutants of the yeast Saccharomyces cerevisiae that have altered sensitivity to isoflurane, a widely used clinical anesthetic. Several lines of evidence from these studies implicate a role for ubiquitin metabolism in cellular response to volatile anesthetics: (i) mutations in the ZZZ1 gene render cells resistant to isoflurane, and the ZZZ1 gene is identical to BUL1 (binds ubiquitin ligase), which appears to be invo...

  17. Reconstitution of CHIP E3 Ubiquitin Ligase Activity

    OpenAIRE

    Ren, Hong Yu; Patterson, Cam; Cyr, Douglas M.; Rosser, Meredith F. N.

    2011-01-01

    CHIP, the carboxyl-terminus of Hsp70 interacting protein, is both an E3 ubiquitin ligase and an Hsp70 co-chaperone and is implicated in the degradation of cytosolic quality control and numerous disease substrates. CHIP has been shown to monitor the folding status of the CFTR protein, and we have successfully reconstituted this activity using a recombinant CFTR fragment consisting of the cytosolic NBD1 and R domains. We have found that efficient ubiquitination of substrates requires chaperone ...

  18. High-mobility group box protein 1 promotes the survival of myeloid-derived suppressor cells by inducing autophagy.

    Science.gov (United States)

    Parker, Katherine H; Horn, Lucas A; Ostrand-Rosenberg, Suzanne

    2016-09-01

    Myeloid-derived suppressor cells are immune-suppressive cells that are elevated in most individuals with cancer, where their accumulation and suppressive activity are driven by inflammation. As myeloid-derived suppressor cells inhibit anti-tumor immunity and promote tumor progression, we are determining how their viability is regulated. Previous studies have established that the damage-associated molecular pattern molecule high-mobility group box protein 1 drives myeloid-derived suppressor cell accumulation and suppressive potency and is ubiquitously present in the tumor microenvironment. As high-mobility group box protein 1 also facilitates tumor cell survival by inducing autophagy, we sought to determine if high-mobility group box protein 1 regulates myeloid-derived suppressor cell survival through induction of autophagy. Inhibition of autophagy increased the quantity of apoptotic myeloid-derived suppressor cells, demonstrating that autophagy extends the survival and increases the viability of myeloid-derived suppressor cells. Inhibition of high-mobility group box protein 1 similarly increased the level of apoptotic myeloid-derived suppressor cells and reduced myeloid-derived suppressor cell autophagy, demonstrating that in addition to inducing the accumulation of myeloid-derived suppressor cells, high-mobility group box protein 1 sustains myeloid-derived suppressor cell viability. Circulating myeloid-derived suppressor cells have a default autophagic phenotype, and tumor-infiltrating myeloid-derived suppressor cells are more autophagic, consistent with the concept that inflammatory and hypoxic conditions within the microenvironment of solid tumors contribute to tumor progression by enhancing immune-suppressive myeloid-derived suppressor cells. Overall, these results demonstrate that in addition to previously recognized protumor effects, high-mobility group box protein 1 contributes to tumor progression by increasing myeloid-derived suppressor cell viability by

  19. Detection of ubiquitinated huntingtin species in intracellular aggregates

    Directory of Open Access Journals (Sweden)

    Katrin eJuenemann

    2015-01-01

    Full Text Available Protein conformation diseases, including polyglutamine diseases, result from the accumulation and aggregation of misfolded proteins. Huntington’s disease is one of nine diseases caused by an expanded polyglutamine repeat within the affected protein and is hallmarked by intracellular inclusion bodies composed of aggregated N-terminal huntingtin fragments and other sequestered proteins. Fluorescence microscopy and filter trap assay are conventional methods to study protein aggregates, but cannot be used to analyze the presence and levels of post-translational modifications of aggregated huntingtin such as ubiquitination. Ubiquitination of proteins can be a signal for degradation and intracellular localization, but also affects protein activity and protein-protein interactions. The function of ubiquitination relies on its mono- and polymeric isoforms attached to protein substrates. Studying the ubiquitination pattern of aggregated huntingtin fragments offers an important possibility to understand huntingtin degradation and aggregation processes within the cell. For the identification of aggregated huntingtin and its ubiquitinated species, solubilization of the cellular aggregates is mandatory. Here we describe methods to identify post-translational modifications such as ubiquitination of aggregated mutant huntingtin. This approach is specifically described for use with mammalian cell culture and is suitable to study other disease-related proteins prone to aggregate.

  20. Ancient origin of animal U-box ubiquitin ligases

    Directory of Open Access Journals (Sweden)

    Marín Ignacio

    2010-10-01

    Full Text Available Abstract Background The patterns of emergence and diversification of the families of ubiquitin ligases provide insights about the evolution of the eukaryotic ubiquitination system. U-box ubiquitin ligases (UULs are proteins characterized by containing a peculiar protein domain known as U box. In this study, the origin of the animal UUL genes is described. Results Phylogenetic and structural data indicate that six of the seven main UUL-encoding genes found in humans (UBE4A, UBE4B, UIP5, PRP19, CHIP and CYC4 were already present in the ancestor of all current metazoans and the seventh (WDSUB1 is found in placozoans, cnidarians and bilaterians. The fact that only 4 - 5 genes orthologous to the human ones are present in the choanoflagellate Monosiga brevicollis suggests that several animal-specific cooptions of the U box to generate new genes occurred. Significantly, Monosiga contains five additional UUL genes that are not present in animals. One of them is also present in distantly-related protozoans. Along animal evolution, losses of UUL-encoding genes are rare, except in nematodes, which lack three of them. These general patterns are highly congruent with those found for other two families (RBR, HECT of ubiquitin ligases. Conclusions Finding that the patterns of emergence, diversification and loss of three unrelated families of ubiquitin ligases (RBR, HECT and U-box are parallel indicates that there are underlying, linage-specific evolutionary forces shaping the complexity of the animal ubiquitin system.

  1. In silico elucidation of the recognition dynamics of ubiquitin.

    Directory of Open Access Journals (Sweden)

    Dong Long

    2011-04-01

    Full Text Available Elucidation of the mechanism of biomacromolecular recognition events has been a topic of intense interest over the past century. The inherent dynamic nature of both protein and ligand molecules along with the continuous reshaping of the energy landscape during the binding process renders it difficult to characterize this process at atomic detail. Here, we investigate the recognition dynamics of ubiquitin via microsecond all-atom molecular dynamics simulation providing both thermodynamic and kinetic information. The high-level of consistency found with respect to experimental NMR data lends support to the accuracy of the in silico representation of the conformational substates and their interconversions of free ubiquitin. Using an energy-based reweighting approach, the statistical distribution of conformational states of ubiquitin is monitored as a function of the distance between ubiquitin and its binding partner Hrs-UIM. It is found that extensive and dense sampling of conformational space afforded by the µs MD trajectory is essential for the elucidation of the binding mechanism as is Boltzmann sampling, overcoming inherent limitations of sparsely sampled empirical ensembles. The results reveal a population redistribution mechanism that takes effect when the ligand is at intermediate range of 1-2 nm from ubiquitin. This mechanism, which may be depicted as a superposition of the conformational selection and induced fit mechanisms, also applies to other binding partners of ubiquitin, such as the GGA3 GAT domain.

  2. Post-translational control of IL-1β via the human papillomavirus type 16 E6 oncoprotein: a novel mechanism of innate immune escape mediated by the E3-ubiquitin ligase E6-AP and p53.

    Directory of Open Access Journals (Sweden)

    Martina Niebler

    Full Text Available Infections with high-risk human papillomaviruses (HPVs are causally involved in the development of anogenital cancer. HPVs apparently evade the innate immune response of their host cells by dysregulating immunomodulatory factors such as cytokines and chemokines, thereby creating a microenvironment that favors malignancy. One central key player in the immune surveillance interactome is interleukin-1 beta (IL-1β which not only mediates inflammation, but also links innate and adaptive immunity. Because of its pleiotropic physiological effects, IL-1β production is tightly controlled on transcriptional, post-translational and secretory levels. Here, we describe a novel mechanism how the high-risk HPV16 E6 oncoprotein abrogates IL-1β processing and secretion in a NALP3 inflammasome-independent manner. We analyzed IL-1β regulation in immortalized keratinocytes that harbor the HPV16 E6 and/or E7 oncogenes as well as HPV-positive cervical tumor cells. While in primary and in E7-immortalized human keratinocytes the secretion of IL-1β was highly inducible upon inflammasome activation, E6-positive cells did not respond. Western blot analyses revealed a strong reduction of basal intracellular levels of pro-IL-1β that was independent of dysregulation of the NALP3 inflammasome, autophagy or lysosomal activity. Instead, we demonstrate that pro-IL-1β is degraded in a proteasome-dependent manner in E6-positive cells which is mediated via the ubiquitin ligase E6-AP and p53. Conversely, in E6- and E6/E7-immortalized cells pro-IL-1β levels were restored by siRNA knock-down of E6-AP and simultaneous recovery of functional p53. In the context of HPV-induced carcinogenesis, these data suggest a novel post-translational mechanism of pro-IL-1β regulation which ultimately inhibits the secretion of IL-1β in virus-infected keratinocytes. The clinical relevance of our results was further confirmed in HPV-positive tissue samples, where a gradual decrease of IL-1

  3. Autophagy inhibitor Lys05 has single-agent antitumor activity and reproduces the phenotype of a genetic autophagy deficiency.

    Science.gov (United States)

    McAfee, Quentin; Zhang, Zhihui; Samanta, Arabinda; Levi, Samuel M; Ma, Xiao-Hong; Piao, Shengfu; Lynch, John P; Uehara, Takeshi; Sepulveda, Antonia R; Davis, Lisa E; Winkler, Jeffrey D; Amaravadi, Ravi K

    2012-05-22

    Autophagy is a lysosome-dependent degradative process that protects cancer cells from multiple stresses. In preclinical models, autophagy inhibition with chloroquine (CQ) derivatives augments the efficacy of many anticancer therapies, but CQ has limited activity as a single agent. Clinical trials are underway combining anticancer agents with hydroxychloroquine (HCQ), but concentrations of HCQ required to inhibit autophagy are not consistently achievable in the clinic. We report the synthesis and characterization of bisaminoquinoline autophagy inhibitors that potently inhibit autophagy and impair tumor growth in vivo. The structural motifs that are necessary for improved autophagy inhibition compared with CQ include the presence of two aminoquinoline rings and a triamine linker and C-7 chlorine. The lead compound, Lys01, is a 10-fold more potent autophagy inhibitor than HCQ. Compared with HCQ, Lys05, a water-soluble salt of Lys01, more potently accumulates within and deacidifies the lysosome, resulting in impaired autophagy and tumor growth. At the highest dose administered, some mice develop Paneth cell dysfunction that resembles the intestinal phenotype of mice and humans with genetic defects in the autophagy gene ATG16L1, providing in vivo evidence that Lys05 targets autophagy. Unlike HCQ, significant single-agent antitumor activity is observed without toxicity in mice treated with lower doses of Lys05, establishing the therapeutic potential of this compound in cancer. PMID:22566612

  4. Ubiquitin Ligase TRIM62 Regulates CARD9-Mediated Anti-fungal Immunity and Intestinal Inflammation

    NARCIS (Netherlands)

    Cao, Zhifang; Conway, Kara L.; Heath, Robert J.; Rush, Jason S.; Leshchiner, Elizaveta S.; Ramirez-Ortiz, Zaida G.; Nedelsky, Natalia B.; Huang, Hailiang; Ng, Aylwin; Gardet, Agnes; Cheng, Shih-Chin; Shamji, Alykhan F.; Rioux, John D.; Wijmenga, Cisca; Netea, Mihai G.; Means, Terry K.; Daly, Mark J.; Xavier, Ramnik J.

    2015-01-01

    CARD9 is a central component of anti-fungal innate immune signaling via C-type lectin receptors, and several immune-related disorders are associated with CARD9 alterations. Here, we used a rare CARD9 variant that confers protection against inflammatory bowel disease as an entry point to investigatin

  5. Nanomaterial-modulated autophagy: underlying mechanisms and functional consequences.

    Science.gov (United States)

    Zheng, Wei; Wei, Min; Li, Song; Le, Weidong

    2016-06-01

    Autophagy is an essential lysosome-dependent process that controls the quality of the cytoplasm and maintains cellular homeostasis, and dysfunction of this protein degradation system is correlated with various disorders. A growing body of evidence suggests that nanomaterials (NMs) have autophagy-modulating effects, thus predicting a valuable and promising application potential of NMs in the diagnosis and treatment of autophagy-related diseases. NMs exhibit unique physical, chemical and biofunctional properties, which may endow NMs with capabilities to modulate autophagy via various mechanisms. The present review highlights the impacts of various NMs on autophagy and their functional consequences. The possible underlying mechanisms for NM-modulated autophagy are also discussed. PMID:27193191

  6. Research Progression of Cellular Autophagy in Liver System Diseases

    Institute of Scientific and Technical Information of China (English)

    Liu Chunyun; Gong Xiangwen; Xiao Xinfa; Yuan Xiangying

    2013-01-01

    Autophagy is a basic biological phenomenon widely existed in eukaryotic cells and an important mechanism for cells to adjust to the surrounding environment, prevent invasion of pathogenic micro-organisms and maintain homeostasis, whose activity changes evidently in multiple liver system diseases, suggesting that there is close association between autophagy and the generation and development of liver system diseases. It is also reported that autophagy develops and exerts an important function in many liver-related diseases, such as hepatic carcinoma, non-alcoholic fatty liver disease, alcoholic liver disease, viral liver disease and acute liver injury. Therefore, this study aimed to summarize the relationship between autophagy and multiple liver diseases, hoping to explore the effect of autophagy in liver system diseases and further study the regulative effect of autophagy so as to provide new thoughts for their treatment.

  7. Autophagy modulation as a target for anticancer drug discovery

    Institute of Scientific and Technical Information of China (English)

    Xin LI; Huai-long XU; Yong-xi LIU; Na AN; Si ZHAO; Jin-ku BAO

    2013-01-01

    Autophagy,an evolutionarily conserved catabolic process involving the engulfment and degradation of non-essential or abnormal cellular organelles and proteins,is crucial for homeostatic maintenance in living cells.This highly regulated,multi-step process has been implicated in diverse diseases including cancer.Autophagy can function as either a promoter or a suppressor of cancer,which makes it a promising and challenging therapeutic target.Herein,we overview the regulatory mechanisms and dual roles of autophagy in cancer.We also describe some of the representative agents that exert their anticancer effects by regulating autophagy.Additionally,some emerging strategies aimed at modulating autophagy are discussed as having the potential for future anticancer drug discovery.In summary,these findings will provide valuable information to better utilize autophagy in the future development of anticancer therapeutics that meet clinical requirements.

  8. Forms, Crosstalks, and the Role of Phospholipid Biosynthesis in Autophagy

    Directory of Open Access Journals (Sweden)

    Leanne Pereira

    2012-01-01

    Full Text Available Autophagy is a highly conserved cellular process occurring during periods of stress to ensure a cell's survival by recycling cytosolic constituents and making products that can be used in energy generation and other essential processes. Three major forms of autophagy exist according to the specific mechanism through which cytoplasmic material is transported to a lysosome. Chaperone-mediated autophagy is a highly selective form of autophagy that delivers specific proteins for lysosomal degradation. Microautophagy is a less selective form of autophagy that occurs through lysosomal membrane invaginations, forming tubes and directly engulfing cytoplasm. Finally, macroautophagy involves formation of new membrane bilayers (autophagosomes that engulf cytosolic material and deliver it to lysosomes. This review provides new insights on the crosstalks between different forms of autophagy and the significance of bilayer-forming phospholipid synthesis in autophagosomal membrane formation.

  9. Epigenetic modifications as regulatory elements of autophagy in cancer.

    Science.gov (United States)

    Sui, Xinbing; Zhu, Jing; Zhou, Jichun; Wang, Xian; Li, Da; Han, Weidong; Fang, Yong; Pan, Hongming

    2015-05-01

    Epigenetic modifications have been considered as hallmarks of cancer and play an important role in tumor initiation and development. Epigenetic mechanisms, including DNA methylation, histone modifications, and microRNAs, may regulate cell cycle and apoptosis, as well as macroautophagy (hereafter referred to as autophagy). Autophagy, as a crucial cellular homeostatic mechanism, performs a dual role, having pro-survival or pro-death properties. A variety of signaling pathways including epigenetic control have been implicated in the upregulation or downregulation of autophagy. However, the role of epigenetic regulation in autophagy is still less well acknowledged. Recent studies have linked epigenetic control to the autophagic process. Some epigenetic modifiers are also involved in the regulation of autophagy and potentiate the efficacy of traditional therapeutics. Thus, understanding the novel functions of epigenetic control in autophagy may allow us to develop potential therapeutic approaches for cancer treatment.

  10. Roles of autophagy in elimination of intracellular bacterial pathogens

    Directory of Open Access Journals (Sweden)

    Eun-Kyeong eJo

    2013-05-01

    Full Text Available As a fundamental intracellular catabolic process, autophagy is important and required for the elimination of protein aggregates and damaged cytosolic organelles during a variety of stress conditions. Autophagy is now being recognized as an essential component of innate immunity; i.e., the recognition, selective targeting, and elimination of microbes. Because of its crucial roles in the innate immune system, therapeutic targeting of bacteria by means of autophagy activation may prove a useful strategy to combat intracellular infections. However, important questions remain, including which molecules are critical in bacterial targeting by autophagy, and which mechanisms are involved in autophagic clearance of intracellular microbes. In this review, we discuss the roles of antibacterial autophagy in intracellular bacterial infections (Mycobacteria, Salmonella, Shigella, Listeria, and Legionella and present recent evidence in support of molecular mechanisms driving autophagy to target bacteria and eliminate invading pathogens.

  11. Protein Camouflage: Supramolecular Anion Recognition by Ubiquitin.

    Science.gov (United States)

    Mallon, Madeleine; Dutt, Som; Schrader, Thomas; Crowley, Peter B

    2016-04-15

    Progress in the field of bio-supramolecular chemistry, the bottom-up assembly of protein-ligand systems, relies on a detailed knowledge of molecular recognition. To address this issue, we have characterised complex formation between human ubiquitin (HUb) and four supramolecular anions. The ligands were: pyrenetetrasulfonic acid (4PSA), p-sulfonato-calix[4]arene (SCLX4), bisphosphate tweezers (CLR01) and meso-tetrakis (4-sulfonatophenyl)porphyrin (TPPS), which vary in net charge, size, shape and hydrophobicity. All four ligands induced significant changes in the HSQC spectrum of HUb. Chemical shift perturbations and line-broadening effects were used to identify binding sites and to quantify affinities. Supporting data were obtained from docking simulations. It was found that these weakly interacting ligands bind to extensive surface patches on HUb. A comparison of the data suggests some general indicators for the protein-binding specificity of supramolecular anions. Differences in binding were observed between the cavity-containing and planar ligands. The former had a preference for the arginine-rich, flexible C terminus of HUb. PMID:26818656

  12. Exercise, Inflammation and Aging

    OpenAIRE

    Jeffrey A Woods; Wilund, Kenneth R.; Martin, Stephen A.; Kistler, Brandon M.

    2011-01-01

    Aging results in chronic low grade inflammation that is associated with increased risk for disease, poor physical functioning and mortality. Strategies that reduce age-related inflammation may improve the quality of life in older adults. Regular exercise is recommended for older people for a variety of reasons including increasing muscle mass and reducing risk for chronic diseases of the heart and metabolic systems. Only recently has exercise been examined in the context of inflammation. This...

  13. Characterization of the Arabidopsis thaliana E3 ubiquitin-ligase AtSINAL7 and identification of the ubiquitination sites.

    Directory of Open Access Journals (Sweden)

    Diego A Peralta

    Full Text Available Protein ubiquitination leading to degradation by the proteasome is an important mechanism in regulating key cellular functions. Protein ubiquitination is carried out by a three step process involving ubiquitin (Ub activation by a E1 enzyme, the transfer of Ub to a protein E2, finally an ubiquitin ligase E3 catalyzes the transfer of the Ub peptide to an acceptor protein. The E3 component is responsible for the specific recognition of the target, making the unveiling of E3 components essential to understand the mechanisms regulating fundamental cell processes through the protein degradation pathways. The Arabidopsis thaliana seven in absentia-like 7 (AtSINAL7 gene encodes for a protein with characteristics from a C3HC4-type E3 ubiquitin ligase. We demonstrate here that AtSINAL7 protein is indeed an E3 protein ligase based on the self-ubiquitination in vitro assay. This activity is dependent of the presence of a Lys residue in position 124. We also found that higher AtSINAL7 transcript levels are present in tissues undergoing active cell division during floral development. An interesting observation is the circadian expression pattern of AtSINAL7 mRNA in floral buds. Furthermore, UV-B irradiation induces the expression of this transcript indicating that AtSINAL7 may be involved in a wide range of different cell processes.

  14. Rimmed vacuoles with beta-amyloid and ubiquitinated filamentous deposits in the muscles of patients with long-standing denervation (postpoliomyelitis muscular atrophy): similarities with inclusion body myositis.

    Science.gov (United States)

    Semino-Mora, C; Dalakas, M C

    1998-10-01

    In the chronically denervated muscles of patients with prior paralytic poliomyelitis, there are secondary myopathic features, including endomysial inflammation and rare vacuolated fibers. To assess the frequency and characteristics of the vacuoles and their similarities with those seen in inclusion body myositis (IBM), we examined 58 muscle biopsy specimens from patients with prior paralytic poliomyelitis for (1) the presence of rimmed vacuoles; (2) acid-phosphatase reactivity; (3) Congo-red-positive amyloid deposits; (4) electron microscopy, searching for tubulofilaments; and (5) immunoelectron microscopy, using antibodies against beta-amyloid and ubiquitin. We found vacuolated muscle fibers in 18 of 58 (31%) biopsies, with a mean frequency of 2.06 +/- 0.42 fibers per specimen. The vacuoles contained acid phosphatase-positive material in 6 of the 18 (33.30%) specimens and stained positive for Congo red in five (27.80%). By immunoelectron microscopy, the vacuoles contained 5.17 +/- 0.13 nm fibrils and 14.9 +/- 0.31 nm filaments that immunoreacted with antibodies to beta-amyloid and ubiquitin in a pattern identical to the one seen in IBM. We conclude that vacuolated muscle fibers containing filamentous inclusions positive for amyloid and ubiquitin are not unique to IBM and the other vacuolar myopathies but can also occur in a chronic neurogenic condition, such as postpoliomyelitis. The chronicity of the underlying disease, rather than the cause, may lead to vacuolar formation, amyloid deposition, and accumulation of ubiquitinated filaments.

  15. Autophagy-Associated Protein SmATG12 Is Required for Fruiting-Body Formation in the Filamentous Ascomycete Sordaria macrospora.

    Directory of Open Access Journals (Sweden)

    Antonia Werner

    Full Text Available In filamentous fungi, autophagy functions as a catabolic mechanism to overcome starvation and to control diverse developmental processes under normal nutritional conditions. Autophagy involves the formation of double-membrane vesicles, termed autophagosomes that engulf cellular components and bring about their degradation via fusion with vacuoles. Two ubiquitin-like (UBL conjugation systems are essential for the expansion of the autophagosomal membrane: the UBL protein ATG8 is conjugated to the lipid phosphatidylethanolamine and the UBL protein ATG12 is coupled to ATG5. We recently showed that in the homothallic ascomycete Sordaria macrospora autophagy-related genes encoding components of the conjugation systems are required for fruiting-body development and/or are essential for viability. In the present work, we cloned and characterized the S. macrospora (Smatg12 gene. Two-hybrid analysis revealed that SmATG12 can interact with SmATG7 and SmATG3. To examine its role in S. macrospora, we replaced the open reading frame of Smatg12 with a hygromycin resistance cassette and generated a homokaryotic ΔSmatg12 knockout strain, which displayed slower vegetative growth under nutrient starvation conditions and was unable to form fruiting bodies. In the hyphae of S. macrospora EGFP-labeled SmATG12 was detected in the cytoplasm and as punctate structures presumed to be phagophores or phagophore assembly sites. Delivery of EGFP-labelled SmATG8 to the vacuole was entirely dependent on SmATG12.

  16. Hsp90 regulates processing of NF-κB2 p100 involving protection of NF-κB-inducing kinase (NIK) from autophagy-mediated degradation

    Institute of Scientific and Technical Information of China (English)

    Guoliang Qing; Pengrong Yan; Zhaoxia Qu; Hudan Liu; Gutian Xiao

    2007-01-01

    NF-κB-inducing kinase (NIK) is required for NF-κB activation based on the processing of NF-κB2 p100. Here we report a novel mechanism of NIK regulation involving the chaperone 90 kDa heat shock protein (Hsp90) and autophagy.Functional inhibition of lisp90 by the anti-tumor agent geldanamycin (GA) efficiently disrupts its interaction with NIK,resulting in NIK degradation and subsequent blockage of p100 processing. Surprisingly, GA-induced NIK degradation is mediated by autophagy, but largely independent of the ubiquitin-proteasome system. Hsp90 seems to be specifically involved in the folding/stabilization of NIK protein, because GA inhibition does not affect NIK mRNA transcription and translation. Furthermore, Hsp90 is not required for NIK-mediated recruitment of the α subunit of IκB kinase to p100, a key step in induction of p100 processing. These findings define an alternative mechanism for Hsp90 client degradation and identify a novel function of autophagy in NF-κB regulation. These findings also suggest a new therapeutic strategy for diseases associated with p100 processing.

  17. Autophagy-Associated Protein SmATG12 Is Required for Fruiting-Body Formation in the Filamentous Ascomycete Sordaria macrospora.

    Science.gov (United States)

    Werner, Antonia; Herzog, Britta; Frey, Stefan; Pöggeler, Stefanie

    2016-01-01

    In filamentous fungi, autophagy functions as a catabolic mechanism to overcome starvation and to control diverse developmental processes under normal nutritional conditions. Autophagy involves the formation of double-membrane vesicles, termed autophagosomes that engulf cellular components and bring about their degradation via fusion with vacuoles. Two ubiquitin-like (UBL) conjugation systems are essential for the expansion of the autophagosomal membrane: the UBL protein ATG8 is conjugated to the lipid phosphatidylethanolamine and the UBL protein ATG12 is coupled to ATG5. We recently showed that in the homothallic ascomycete Sordaria macrospora autophagy-related genes encoding components of the conjugation systems are required for fruiting-body development and/or are essential for viability. In the present work, we cloned and characterized the S. macrospora (Sm)atg12 gene. Two-hybrid analysis revealed that SmATG12 can interact with SmATG7 and SmATG3. To examine its role in S. macrospora, we replaced the open reading frame of Smatg12 with a hygromycin resistance cassette and generated a homokaryotic ΔSmatg12 knockout strain, which displayed slower vegetative growth under nutrient starvation conditions and was unable to form fruiting bodies. In the hyphae of S. macrospora EGFP-labeled SmATG12 was detected in the cytoplasm and as punctate structures presumed to be phagophores or phagophore assembly sites. Delivery of EGFP-labelled SmATG8 to the vacuole was entirely dependent on SmATG12. PMID:27309377

  18. Autophagy and mitophagy in the myocardium: therapeutic potential and concerns

    OpenAIRE

    Jimenez, Rebecca E; Kubli, Dieter A.; Gustafsson, Åsa B.

    2014-01-01

    The autophagic-lysosomal degradation pathway is critical for cardiac homeostasis, and defects in this pathway are associated with development of cardiomyopathy. Autophagy is responsible for the normal turnover of organelles and long-lived proteins. Autophagy is also rapidly up-regulated in response to stress, where it rapidly clears dysfunctional organelles and cytotoxic protein aggregates in the cell. Autophagy is also important in clearing dysfunctional mitochondria before they can cause ha...

  19. Application and interpretation of current autophagy inhibitors and activators

    Institute of Scientific and Technical Information of China (English)

    Ya-ping YANG; Li-fang HU; Hui-fen ZHENG; Cheng-jie MAO; Wei-dong HU; Kang-ping XIONG; Fen WANG

    2013-01-01

    Aut ophagy is the major intracellular degradation system,by which cytoplasmic materials are delivered to and degraded in the lysosome.As a quality control mechanism for cytoplasmic proteins and organelles,autophagy plays important roles in a variety of human diseases,including neurodegenerative diseases,cancer,cardiovascular disease,diabetes and infectious and inflammatory diseases.The discovery of ATG genes and the dissection of the signaling pathways involved in regulating autophagy have greatly enriched our knowledge on the occurrence and development of this lysosomal degradation pathway.In addition to its role in degradation,autophagy may also promote a type of programmed cell death that is different from apoptosis,termed type II programmed cell death.Owing to the dual roles of autophagy in cell death and the specificity of diseases,the exact mechanisms of autophagy in various diseases require more investigation.The application of autophagy inhibitors and activators will help us understand the regulation of autophagy in human diseases,and provide insight into the use of autophagy-targeted drugs.In this review,we summarize the latest research on autophagy inhibitors and activators and discuss the possibility of their application in human disease therapy.

  20. The Mucosal Immune System and Its Regulation by Autophagy.

    Science.gov (United States)

    Kabat, Agnieszka M; Pott, Johanna; Maloy, Kevin J

    2016-01-01

    The gastrointestinal tract presents a unique challenge to the mucosal immune system, which has to constantly monitor the vast surface for the presence of pathogens, while at the same time maintaining tolerance to beneficial or innocuous antigens. In the intestinal mucosa, specialized innate and adaptive immune components participate in directing appropriate immune responses toward these diverse challenges. Recent studies provide compelling evidence that the process of autophagy influences several aspects of mucosal immune responses. Initially described as a "self-eating" survival pathway that enables nutrient recycling during starvation, autophagy has now been connected to multiple cellular responses, including several aspects of immunity. Initial links between autophagy and host immunity came from the observations that autophagy can target intracellular bacteria for degradation. However, subsequent studies indicated that autophagy plays a much broader role in immune responses, as it can impact antigen processing, thymic selection, lymphocyte homeostasis, and the regulation of immunoglobulin and cytokine secretion. In this review, we provide a comprehensive overview of mucosal immune cells and discuss how autophagy influences many aspects of their physiology and function. We focus on cell type-specific roles of autophagy in the gut, with a particular emphasis on the effects of autophagy on the intestinal T cell compartment. We also provide a perspective on how manipulation of autophagy may potentially be used to treat mucosal inflammatory disorders. PMID:27446072

  1. Autophagy is essential for cardiac morphogenesis during vertebrate development.

    Science.gov (United States)

    Lee, Eunmyong; Koo, Yeon; Ng, Aylwin; Wei, Yongjie; Luby-Phelps, Kate; Juraszek, Amy; Xavier, Ramnik J; Cleaver, Ondine; Levine, Beth; Amatruda, James F

    2014-04-01

    Genetic analyses indicate that autophagy, an evolutionarily conserved lysosomal degradation pathway, is essential for eukaryotic differentiation and development. However, little is known about whether autophagy contributes to morphogenesis during embryogenesis. To address this question, we examined the role of autophagy in the early development of zebrafish, a model organism for studying vertebrate tissue and organ morphogenesis. Using zebrafish that transgenically express the fluorescent autophagy reporter protein, GFP-LC3, we found that autophagy is active in multiple tissues, including the heart, during the embryonic period. Inhibition of autophagy by morpholino knockdown of essential autophagy genes (including atg5, atg7, and becn1) resulted in defects in morphogenesis, increased numbers of dead cells, abnormal heart structure, and reduced organismal survival. Further analyses of cardiac development in autophagy-deficient zebrafish revealed defects in cardiac looping, abnormal chamber morphology, aberrant valve development, and ectopic expression of critical transcription factors including foxn4, tbx5, and tbx2. Consistent with these results, Atg5-deficient mice displayed abnormal Tbx2 expression and defects in valve development and chamber septation. Thus, autophagy plays an essential, conserved role in cardiac morphogenesis during vertebrate development.

  2. Autophagy and bacterial infection: an evolving arms race.

    Science.gov (United States)

    Choy, Augustine; Roy, Craig R

    2013-09-01

    Autophagy is an important membrane transport pathway that is conserved among eukaryotic cells. Although first described as an intracellular catabolic pathway used to break down self-components, autophagy has been found to play an important role in the elimination of intracellular pathogens. A variety of host mechanisms exist for recognizing and targeting intracellular bacteria to autophagosomes. Several intracellular bacteria have evolved ways to manipulate, inhibit, or avoid autophagy in order to survive in the cell. Thus, the autophagy pathway can be viewed as an evolutionarily conserved host response to infection.

  3. Autophagy: A boon or bane in oral cancer.

    Science.gov (United States)

    Adhauliya, Namrata; Kalappanavar, Anupama N; Ali, I M; Annigeri, Rajeshwari G

    2016-10-01

    Autophagy is a catabolic process involving cellular recycling and is believed to play a distinct role in cell survival especially when exposed to stressors, rendering it comparable to the elixir sustaining life. It plays a significant role in various conditions like cancers, neuropathies, heart diseases, auto-immune diseases, etc. Its role in tumorigenesis and cancer therapeutics is worth exploring. Autophagy is believed to help in survival and longevity of cancer cells by buffering metabolic stress. Inhibition of autophagy in an environment of nutrient deprivation leads to cell death. Autophagy is also seen to facilitate metastasizing tumor cells in surviving the conditions of metabolic deprivation and in recovery when conditions turn favorable. Many current cancer therapies tend to inflict metabolic stress, thus autophagy inhibitors may be useful in cancer treatment. As per the adage, "excess of anything is bad", the autophagy promoters can also be exploited as beneficial tools in the fight against cancer. Another method for tumor-cell elimination can be by inducing autophagic cell death through over-stimulation. Oral cancers are becoming a leading cause of deaths worldwide. Much remains to be explored about the role autophagy plays in progression of head and neck cancers, so as to harness it in the therapeutics of these cancers. Research on autophagy is still in its infancy. There are knowledge gaps in understanding this complex process. But there is no doubt that understanding exact mechanism behind autophagy will open up new avenues in cancer therapeutics and even prevention. PMID:27688114

  4. Significant role of PB1 and UBA domains in multimerization of Joka2, a selective autophagy cargo receptor from tobacco

    Directory of Open Access Journals (Sweden)

    Katarzyna eZientara-Rytter

    2014-01-01

    Full Text Available Tobacco Joka2 protein is a hybrid homolog of two mammalian selective autophagy cargo receptors, p62 and NBR1. These proteins can directly interact with the members of ATG8 family and the polyubiquitinated cargoes designed for degradation. Function of the selective autophagy cargo receptors relies on their ability to form protein aggregates. It has been shown that the N-terminal PB1 domain of p62 is involved in formation of aggregates, while the UBA domains of p62 and NBR1 have been associated mainly with cargo binding. Here we focus on roles of PB1 and UBA domains in localization and aggregation of Joka2 in plant cells. We show that Joka2 can homodimerize not only through its N-terminal PB1-PB1 interactions but also via interaction between N-terminal PB1 and C-terminal UBA domains. We also demonstrate that Joka2 co-localizes with recombinant ubiquitin and sequestrates it into aggregates and that C-terminal part (containing UBA domains is sufficient for this effect. Our results indicate that Joka2 accumulates in cytoplasmic aggregates and suggest that in addition to these multimeric forms it also exists in the nucleus and cytoplasm in a monomeric form.

  5. Autophagy Regulatory Network - a systems-level bioinformatics resource for studying the mechanism and regulation of autophagy.

    Science.gov (United States)

    Türei, Dénes; Földvári-Nagy, László; Fazekas, Dávid; Módos, Dezső; Kubisch, János; Kadlecsik, Tamás; Demeter, Amanda; Lenti, Katalin; Csermely, Péter; Vellai, Tibor; Korcsmáros, Tamás

    2015-01-01

    Autophagy is a complex cellular process having multiple roles, depending on tissue, physiological, or pathological conditions. Major post-translational regulators of autophagy are well known, however, they have not yet been collected comprehensively. The precise and context-dependent regulation of autophagy necessitates additional regulators, including transcriptional and post-transcriptional components that are listed in various datasets. Prompted by the lack of systems-level autophagy-related information, we manually collected the literature and integrated external resources to gain a high coverage autophagy database. We developed an online resource, Autophagy Regulatory Network (ARN; http://autophagy-regulation.org), to provide an integrated and systems-level database for autophagy research. ARN contains manually curated, imported, and predicted interactions of autophagy components (1,485 proteins with 4,013 interactions) in humans. We listed 413 transcription factors and 386 miRNAs that could regulate autophagy components or their protein regulators. We also connected the above-mentioned autophagy components and regulators with signaling pathways from the SignaLink 2 resource. The user-friendly website of ARN allows researchers without computational background to search, browse, and download the database. The database can be downloaded in SQL, CSV, BioPAX, SBML, PSI-MI, and in a Cytoscape CYS file formats. ARN has the potential to facilitate the experimental validation of novel autophagy components and regulators. In addition, ARN helps the investigation of transcription factors, miRNAs and signaling pathways implicated in the control of the autophagic pathway. The list of such known and predicted regulators could be important in pharmacological attempts against cancer and neurodegenerative diseases.

  6. Inflammation and coagulation

    NARCIS (Netherlands)

    M. Levi; T. van der Poll

    2010-01-01

    In the pathogenesis of sepsis, inflammation and coagulation play a pivotal role. Increasing evidence points to an extensive cross-talk between these two systems, whereby inflammation leads to activation of coagulation, and coagulation also considerably affects inflammatory activity. Molecular pathwa

  7. Activation of RARα induces autophagy in SKBR3 breast cancer cells and depletion of key autophagy genes enhances ATRA toxicity.

    Science.gov (United States)

    Brigger, D; Schläfli, A M; Garattini, E; Tschan, M P

    2015-01-01

    All-trans retinoic acid (ATRA), a pan-retinoic acid receptor (RAR) agonist, is, along with other retinoids, a promising therapeutic agent for the treatment of a variety of solid tumors. On the one hand, preclinical studies have shown promising anticancer effects of ATRA in breast cancer; on the other hand, resistances occurred. Autophagy is a cellular recycling process that allows the degradation of bulk cellular contents. Tumor cells may take advantage of autophagy to cope with stress caused by anticancer drugs. We therefore wondered if autophagy is activated by ATRA in mammary tumor cells and if modulation of autophagy might be a potential novel treatment strategy. Indeed, ATRA induces autophagic flux in ATRA-sensitive but not in ATRA-resistant human breast cancer cells. Moreover, using different RAR agonists as well as RARα-knockdown breast cancer cells, we demonstrate that autophagy is dependent on RARα activation. Interestingly, inhibition of autophagy in breast cancer cells by either genetic or pharmacological approaches resulted in significantly increased apoptosis under ATRA treatment and attenuated epithelial differentiation. In summary, our findings demonstrate that ATRA-induced autophagy is mediated by RARα in breast cancer cells. Furthermore, inhibition of autophagy results in enhanced apoptosis. This points to a potential novel treatment strategy for a selected group of breast cancer patients where ATRA and autophagy inhibitors are applied simultaneously. PMID:26313912

  8. Triggers of airway inflammation.

    Science.gov (United States)

    Kerrebijn, K F

    1986-01-01

    Most asthmatics have hyperresponsive airways. This makes them more sensitive than non-asthmatics to bronchoconstricting environmental exposures which, in their turn, may enhance responsiveness. Airway inflammation is considered to be a key determinant of airway hyperresponsiveness: the fact that chronic airway inflammation in cystic fibrosis does not lead to airway hyperresponsiveness of any importance indicates, however, that the role of airway inflammation is complex and incompletely elucidated. The main inducers of airway inflammation are viral infections, antigens, occupational stimuli and pollutants. Although exercise, airway cooling and hyper- or hypotonic aerosols are potent stimuli of bronchoconstriction, it is questionable if airway inflammation is involved in their mode of action. Each of the above-mentioned stimuli is discussed, with emphasis laid on the relation of symptoms to mechanisms. PMID:3533597

  9. Selective inhibition of PI3K/Akt/mTOR signaling pathway regulates autophagy of macrophage and vulnerability of atherosclerotic plaque.

    Directory of Open Access Journals (Sweden)

    Chungang Zhai

    Full Text Available Macrophage infiltration contributes to the instability of atherosclerotic plaques. In the present study, we investigated whether selective inhibition of PI3K/Akt/mTOR signaling pathway can enhance the stability of atherosclerotic plaques by activation of macrophage autophagy. In vitro study, selective inhibitors or siRNA of PI3K/Akt/mTOR pathways were used to treat the rabbit's peritoneal primary macrophage cells. Inflammation related cytokines secreted by macrophages were measured. Ultrastructure changes of macrophages were examined by transmission electron microscope. mRNA or protein expression levels of autophagy related gene Beclin 1, protein 1 light chain 3 II dots (LC3-II or Atg5-Atg12 conjugation were assayed by quantitative RT-PCR or Western blot. In vivo study, vulnerable plaque models were established in 40 New Zealand White rabbits and then drugs or siRNA were given for 8 weeks to inhibit the PI3K/Akt/mTOR signaling pathway. Intravascular ultrasound (IVUS was performed to observe the plaque imaging. The ultrastructure of the abdominal aortic atherosclerosis lesions were analyzed with histopathology. RT-PCR or Western blot methods were used to measure the expression levels of corresponding autophagy related molecules. We found that macrophage autophagy was induced in the presence of Akt inhibitor, mTOR inhibitor and mTOR-siRNA in vitro study, while PI3K inhibitor had the opposite role. In vivo study, we found that macrophage autophagy increased significantly and the rabbits had lower plaque rupture incidence, lower plaque burden and decreased vulnerability index in the inhibitors or siRNA treated groups. We made a conclusion that selective inhibition of the Akt/mTOR signal pathway can reduce macrophages and stabilize the vulnerable atherosclerotic plaques by promoting macrophage autophagy.

  10. Autophagy sensitivity of neuroendocrine lung tumor cells

    OpenAIRE

    HONG, SEUNG-KEUN; Kim, Jin-Hwan; Starenki, Dmytro; Park, Jong-In

    2013-01-01

    Neuroendocrine (NE) phenotypes characterize a spectrum of lung tumors, including low-grade typical and intermediate-grade atypical carcinoid, high-grade large-cell NE carcinoma and small cell lung carcinoma. Currently, no effective treatments are available to cure NE lung tumors, demanding identification of biological features specific to these tumors. Here, we report that autophagy has an important role for NE lung tumor cell proliferation and survival. We found that the expression levels of...

  11. The Role of Autophagy in Lupus Nephritis

    OpenAIRE

    Linlin Wang; Helen Ka Wai Law

    2015-01-01

    Systemic lupus erythematosus (SLE) is a multifactorial autoimmune disease characterized by the generation of immune responses to self-antigens. Lupus nephritis is one of the most common and severe complications in SLE patients. Though the pathogenesis of lupus nephritis has been studied extensively, unresolved questions are still left and new therapeutic methods are needed for disease control. Autophagy is a conserved catabolic process through which cytoplasmic constituents can be degraded in...

  12. Autophagy in Mycobacterium tuberculosis and HIV infections

    OpenAIRE

    Espert, Lucile; Beaumelle, Bruno; Vergne, Isabelle

    2015-01-01

    Human Immunodeficiency Virus (HIV) and Mycobacterium tuberculosis (M.tb) are among the most lethal human pathogens worldwide, each being responsible for around 1.5 million deaths annually. Moreover, synergy between acquired immune deficiency syndrome (AIDS) and tuberculosis (TB) has turned HIV/M.tb co-infection into a major public health threat in developing countries. In the past decade, autophagy, a lysosomal catabolic process, has emerged as a major host immune defense mechanism against in...

  13. UHRF2, another E3 ubiquitin ligase for p53

    Energy Technology Data Exchange (ETDEWEB)

    Bai, Lu; Wang, Xiaohui; Jin, Fangmin; Yang, Yan; Qian, Guanhua [Department of Cell Biology and Medical Genetics, Chongqing Medical University, Chongqing (China); Duan, Changzhu, E-mail: duanchzhu@cqmu.edu.cn [Key Laboratory of Clinical Laboratory Diagnostics of Ministry of Education, Faculty of Laboratory Medicine, Chongqing Medical University, Chongqing (China); Department of Cell Biology and Medical Genetics, Chongqing Medical University, Chongqing (China)

    2012-09-07

    Highlights: Black-Right-Pointing-Pointer UHRF2 associates with p53 in vivo and in vitro. Black-Right-Pointing-Pointer UHRF2 interacts with p53 through its SRA/YDG domain. Black-Right-Pointing-Pointer UHRF2 ubiquitinates p53 in vivo and in vitro. -- Abstract: UHRF2, ubiquitin-like with PHD and ring finger domains 2, is a nuclear E3 ubiquitin ligase, which is involved in cell cycle and epigenetic regulation. UHRF2 interacts with multiple cell cycle proteins, including cyclins (A2, B1, D1, and E1), CDK2, and pRb; moreover, UHRF2 could ubiquitinate cyclin D1 and cyclin E1. Also, UHRF2 has been shown to be implicated in epigenetic regulation by associating with DNMTs, G9a, HDAC1, H3K9me2/3 and hemi-methylated DNA. We found that UHRF2 associates with tumor suppressor protein p53, and p53 is ubiquitinated by UHRF2 in vivo and in vitro. Given that both UHRF2 and p53 are involved in cell cycle regulation, this study may suggest a novel signaling pathway on cell proliferation.

  14. Roles of protein ubiquitination and degradation kinetics in biological oscillations.

    Directory of Open Access Journals (Sweden)

    Lida Xu

    Full Text Available Protein ubiquitination and degradation play important roles in many biological functions and are associated with many human diseases. It is well known that for biochemical oscillations to occur, proper degradation rates of the participating proteins are needed. In most mathematical models of biochemical reactions, linear degradation kinetics has been used. However, the degradation kinetics in real systems may be nonlinear, and how nonlinear degradation kinetics affects biological oscillations are not well understood. In this study, we first develop a biochemical reaction model of protein ubiquitination and degradation and calculate the degradation rate against the concentration of the free substrate. We show that the protein degradation kinetics mainly follows the Michaelis-Menten formulation with a time delay caused by ubiquitination and deubiquitination. We then study analytically how the Michaelis-Menten degradation kinetics affects the instabilities that lead to oscillations using three generic oscillation models: 1 a positive feedback mediated oscillator; 2 a positive-plus-negative feedback mediated oscillator; and 3 a negative feedback mediated oscillator. In all three cases, nonlinear degradation kinetics promotes oscillations, especially for the negative feedback mediated oscillator, resulting in much larger oscillation amplitudes and slower frequencies than those observed with linear kinetics. However, the time delay due to protein ubiquitination and deubiquitination generally suppresses oscillations, reducing the amplitude and increasing the frequency of the oscillations. These theoretical analyses provide mechanistic insights into the effects of specific proteins in the ubiquitination-proteasome system on biological oscillations.

  15. The Challenge of Producing Ubiquitinated Proteins for Structural Studies

    Directory of Open Access Journals (Sweden)

    Serena Faggiano

    2014-06-01

    Full Text Available Protein ubiquitination is an important post-translational modification involved in several essential signalling pathways. It has different effects on the target protein substrate, i.e., it can trigger the degradation of the protein in the proteasome, change the interactions of the modified protein with its partners, or affect its localization and activity. In order to understand the molecular mechanisms underlying the consequences of protein ubiquitination, scientists have to face the challenging task of producing ubiquitinated proteins for structural characterization with X-ray crystallography and/or nuclear magnetic resonance (NMR spectroscopy. These techniques require milligrams of homogeneous samples of high purity. The strategies proposed so far for the production of ubiquitinated proteins can be divided into two groups, i.e., chemical (or non-enzymatic and enzymatic methodologies. In this review, we summarize the still very sparse examples available in the literature that describe successful production of ubiquitinated proteins amenable for biochemical and structural studies, and discuss advantages and disadvantages of the techniques proposed. We also give a perspective of the direction in which the field might evolve.

  16. Regulation of autophagy by nucleoporin Tpr.

    Science.gov (United States)

    Funasaka, Tatsuyoshi; Tsuka, Eriko; Wong, Richard W

    2012-01-01

    The nuclear pore complex (NPC) consists of a conserved set of ~30 different proteins, termed nucleoporins, and serves as a gateway for the exchange of materials between the cytoplasm and nucleus. Tpr (translocated promoter region) is a component of NPC that presumably localizes at intranuclear filaments. Here, we show that Tpr knockdown caused a severe reduction in the number of nuclear pores. Furthermore, our electron microscopy studies indicated a significant reduction in the number of inner nuclear filaments. In addition, Tpr siRNA treatment impaired cell growth and proliferation compared to control siRNA-treated cells. In Tpr-depleted cells, the levels of p53 and p21 proteins were enhanced. Surprisingly, Tpr depletion increased p53 nuclear accumulation and facilitated autophagy. Our study demonstrates for the first time that Tpr plays a role in autophagy through controlling HSP70 and HSF1 mRNA export, p53 trafficking with karyopherin CRM1, and potentially through direct transcriptional regulation of autophagy factors.

  17. Autophagy and ethanol-induced liver injury

    Institute of Scientific and Technical Information of China (English)

    Terrence M Donohue Jr

    2009-01-01

    The majority of ethanol metabolism occurs in the liver. Consequently, this organ sustains the greatest damage from ethanol abuse. Ethanol consumption disturbs the delicate balance of protein homeostasis in the liver, causing intracellular protein accumulation due to a disruption of hepatic protein catabolism.Evidence indicates that ethanol or its metabolism impairs trafficking events in the liver, including the process of macroautophagy, which is the engulfment and degradation of cytoplasmic constituents by the lysosomal system. Autophagy is an essential, ongoing cellular process that is highly regulated by nutrients,endocrine factors and signaling pathways. A great number of the genes and gene products that govern the autophagic response have been characterized and the major metabolic and signaling pathways that activate or suppress autophagy have been identified. This review describes the process of autophagy, its regulation and the possible mechanisms by which ethanol disrupts the process of autophagic degradation. The implications of autophagic suppression are discussed in relation to the pathogenesis of alcohol-induced liver injury.

  18. Dynamics of ubiquitin-mediated signalling: insights from mathematical modelling and experimental studies.

    Science.gov (United States)

    Nguyen, Lan K

    2016-05-01

    Post-translational modification of cellular proteins by ubiquitin is a pivotal regulatory event that controls not only protein degradation, but also a variety of non-proteolytic functions. Ubiquitination is involved in a broad array of physiological processes, and its dysregulation has been associated with many human diseases, including neuronal disorders and cancers. Ubiquitin-mediated signalling has thus come to the forefront of biomedical research. It is increasingly apparent that ubiquitination is a highly complex and dynamic process, evidenced by a myriad of ways of ubiquitin chain formation, tightly regulatory mechanisms involving E3 ligases and deubiquitinating enzymes and extensive crosstalk with other post-translational modifications. To unravel the complexity of ubiquitination and understand the dynamic properties of ubiquitin-mediated signalling are challenging, but critical topics in ubiquitin research, which will undoubtedly benefit our effort in developing strategies that could target ubiquitin signalling for therapeutics. Computational modelling and model-based approaches are emerging as promising tools that help tackle the complexity and provide useful frameworks for quantitative and dynamical analysis of ubiquitin signalling. In this article, I will discuss recent advances in our understanding of the dynamic behaviour of ubiquitination from both theoretical and experimental studies, and aspects of ubiquitin signalling that may have major dynamical consequences. It is expected the discussed issues will be of relevant interest to both the ubiquitin and systems biology fields.

  19. Autophagy-associated atrophy and metabolic remodeling of the mouse diaphragm after short-term intermittent hypoxia.

    Directory of Open Access Journals (Sweden)

    Christian Giordano

    Full Text Available Short-term intermittent hypoxia (IH is common in patients with acute respiratory disorders. Although prolonged exposure to hypoxia induces atrophy and increased fatigability of skeletal muscle, the response to short-term IH is less well known. We hypothesized that the diaphragm and limb muscles would adapt differently to short-term IH given that hypoxia stimulates ventilation and triggers a superimposed exercise stimulus in the diaphragm.We determined the structural, metabolic, and contractile properties of the mouse diaphragm after 4 days of IH (8 hours per day, 30 episodes per hour to a FiO2 nadir=6%, and compared responses in the diaphragm to a commonly studied reference limb muscle, the tibialis anterior. Outcome measures included muscle fiber size, assays of muscle proteolysis (calpain, ubiquitin-proteasome, and autophagy pathways, markers of oxidative stress and mitochondrial function, quantification of intramyocellular lipid and lipid metabolism genes, type I myosin heavy chain (MyHC expression, and in vitro contractile properties.After 4 days of IH, the diaphragm alone demonstrated significant atrophy (30% decrease of myofiber size together with increased LC3B-II protein (2.4-fold and mRNA markers of the autophagy pathway (LC3B, Gabarapl1, Bnip3, whereas active calpain and E3 ubiquitin ligases (MuRF1, atrogin-1 were unaffected in both muscles. Succinate dehydrogenase activity was significantly reduced by IH in both muscles. However, only the diaphragm exhibited increased intramyocellular lipid droplets (2.5-fold after IH, along with upregulation of genes linked to activated lipid metabolism. In addition, although the diaphragm showed evidence for acute fatigue immediately following IH, it underwent an adaptive fiber type switch toward slow type I MyHC-expressing fibers, associated with greater intrinsic endurance of the muscle during repetitive stimulation in vitro.Short-term IH induces preferential atrophy in the mouse diaphragm

  20. The Role of Ubiquitine Proteasome Pathway in Carcinogenesis

    Directory of Open Access Journals (Sweden)

    N.Ceren Sumer Turanligil

    2010-02-01

    Full Text Available Ubiquitin works as a marker protein which targets misfolded or injured proteins to cellular degradation. It brings the abnormal proteins to a subcellular organelle named proteasome and it maintains the degradation of proteins in limited lenghts of peptides by leaving the process withuout being changed. Mistakes in ubiquitin-dependent proteolysis in various steps of carcinogenesis is known. In this review, we dealed with the effects of ubiquitin-proteasome pathway (UPP on carcinogenesis via intercellular signaling molecules like Ras, transcription factors like NF-kB, cytokines like TNF-alfa Tumor necrosis factor, protooncogenes like p53 and MDM2(murine double minute 2, components of cell cycle and DNA repair proteins like BRCA1. We also focused on the relationship of UPP on antigen presentation which is active in immune response and its place in the aetiology of colon cancer to provide a specific example. [Archives Medical Review Journal 2010; 19(1.000: 36-55

  1. Autophagy: Friend or Foe in Breast Cancer Development, Progression, and Treatment

    Directory of Open Access Journals (Sweden)

    Damian E. Berardi

    2011-01-01

    Although autophagy inhibition, combined with anticancer agents, could be therapeutically beneficial in some cases, autophagy induction by itself could lead to cell death in some apoptosis-resistant cancers, indicating that autophagy induction may also be used as a therapy. This paper summarizes the most important findings described in the literature about autophagy and also discusses the importance of this process in clinical settings.

  2. Biochemical Analysis of Autophagy in Algae and Plants by Monitoring the Electrophoretic Mobility of ATG8.

    Science.gov (United States)

    Pérez-Pérez, María Esther; Andrés-Garrido, Ascensión; Crespo, José L

    2016-01-01

    Identification of specific autophagy markers has been fundamental to investigate autophagy as catabolic process. Among them, the ATG8 protein turned out to be one of the most widely used and specific molecular markers of autophagy both in higher and lower eukaryotes. Here, we describe how ATG8 can be used to monitor autophagy in Chlamydomonas and Arabidopsis by western blot analysis. PMID:27424752

  3. 自噬蛋白NDP52与细菌感染的研究进展%Research Advances in Autophagy Protein NDP52 and Bacterial Infection

    Institute of Scientific and Technical Information of China (English)

    王芬芬; 张吉翔

    2012-01-01

    自噬(autophagy)是哺乳动物清除入侵细菌的主要途径,可保卫宿主细胞免受细菌的损伤.核点蛋白52(nuclear dot protein 52,NDP52)——核点家族成员之一,是除p62/SQSTM1和NBR1 等之外最新发现的自噬关键蛋白.它连接自噬体表面的微管相关蛋白1轻链3(microtubule associated protein l light chain 3,LC3),将披上“泛素大衣”的病原菌(如沙门氏菌和化脓性链球菌)递送至自噬体内加以清除.这一发现有助于人们深入了解自噬抵抗病原微生物感染的具体分子机制,为预防和治疗细菌感染提供了新靶点.%Autophagy is an important pathway to eliminate intracellular bacteria in mammals and defend the host cell from bacteria damage. In addition to p62/SQSTMl and NBR1, NDP52 (nuclear dot protein 52)—one of the members of nuclear dots, is a novel autophagy related protein has been found. NDP52 is an adaptor protein that binds to both ubiquitinated bacteria (such as Salmonella and Streptococcus pyogenes) and LC3 (microtubule associated protein 1 light chain 3), and then delivers these invaders into autophagosomes. This finding helps further understand the specific molecular mechanism of autophagy resisting pathogens infection, and provides a new target for the prevention and treatment of bacteria infection.

  4. In silico analysis of ubiquitin/ubiquitin-like modifiers and their conjugating enzymes in Entamoeba species.

    Science.gov (United States)

    Arya, Shweta; Sharma, Gaurav; Gupta, Preeti; Tiwari, Swati

    2012-07-01

    Covalent modification of proteins by ubiquitin (Ub) and ubiquitin-like modifiers (Ubls) regulates many cellular functions in eukaryotes. These modifications are likely to be associated with pathogenesis, growth, and development of many protozoan parasites but molecular details about this pathway are unavailable for most protozoa. This study presents an analysis of the Ub pathway in three members of the Entamoeba species. Using bioinformatics tools we have identified all Ub and Ubl genes along with their corresponding activating, conjugating, and ligating enzymes (E1, E2s, and E3s) in three Entamoeba species, Entamoeba histolytica, Entamoeba dispar, and Entamoeba invadens. Phylogenetic trees were established for the identified E2s and RING finger E3s using maximum-likelihood method to infer the relationship among these proteins. In silico co-domain analysis of RING finger E3s implicates these proteins in a variety of functions. Several known and putative regulatory motifs were identified in the upstream regions of RING finger domain containing E3 genes. All E2 and E3 genes were analyzed in genomic context in E. histolytica and E. dispar. Most E2s and E3s were in syntenic positions in the two genomes. Association of these genes with transposable elements (TEs) was compared between E. histolytica and E. dispar. A closer association was found between RING finger E3s with TEs in E. histolytica. In summary, our analyses suggests that the complexity of the Ub pathway in Entamoeba species is close to that observed in higher eukaryotes. This study provides important data for further understanding the role of Ub pathway in the biology of these organisms.

  5. Autophagy Is Associated with Pathogenesis of Haemophilus parasuis

    Science.gov (United States)

    Zhang, Yaning; Li, Yufeng; Yuan, Wentao; Xia, Yuting; Shen, Yijuan

    2016-01-01

    Haemophilus parasuis (H. parasuis) is a common commensal Gram-negative extracellular bacterium in the upper respiratory tract of swine, which can cause Glässer's disease in stress conditions. Research on the pathogenicity of H. parasuis has mainly focused on immune evasion and bacterial virulence factors, while few studies have examined the interactions of H. parasuis and its host. Autophagy is associated with the replication and proliferation of many pathogenic bacteria, but whether it plays a role during infection by H. parasuis is unknown. In this study, an adenovirus construct expressing GFP, RFP, and LC3 was used to infect H. parasuis. Western blotting, laser confocal microscopy, and electron microscopy showed that Hps5 infection induced obvious autophagy in PK-15 cells. In cells infected with strains of H. parasuis differing in invasiveness, the levels of autophagy were positively correlated with the presence of alive bacteria in PK-15 cells. In addition, autophagy inhibited the invasion of Hps5 in PK-15 cells. Autophagy related genes Beclin, Atg5 and Atg7 were silenced with RNA interference, the results showed that autophagy induced by H. parasuis infection is a classical pathway. Our observations demonstrate that H. parasuis can induce autophagy and that the levels of autophagy are associated with the presence of alive bacteria in cells, which opened novel avenues to further our understanding of H. parasuis-host interplay and pathogenesis. PMID:27703447

  6. Autophagy: A double-edged sword in Alzheimer's disease

    Indian Academy of Sciences (India)

    Ying-Tsen Tung; Bo-Jeng Wang; Ming-Kuan Hu; Wen-Ming Hsu; Hsinyu Lee; Wei-Pang Huang; Yung-Feng Liao

    2012-03-01

    Autophagy is a major protein degradation pathway that is essential for stress-induced and constitutive protein turnover. Accumulated evidence has demonstrated that amyloid- (A) protein can be generated in autophagic vacuoles, promoting its extracellular deposition in neuritic plaques as the pathological hallmark of Alzheimer’s disease (AD). The molecular machinery for A generation, including APP, APP-C99 and -/-secretases, are all enriched in autophagic vacuoles. The induction of autophagy can be vividly observed in the brain at early stages of sporadic AD and in an AD transgenic mouse model. Accumulated evidence has also demonstrated a neuroprotective role of autophagy in mediating the degradation of aggregated proteins that are causative of various neurodegenerative diseases. Autophagy is thus widely regarded as an intracellular hub for the removal of the detrimental A peptides and Tau aggregates. Nonetheless, compelling data also reveal an unfavorable function of autophagy in facilitating the production of intracellular A. The two faces of autophagy on the homeostasis of A place it in a very unique and intriguing position in ADpathogenesis. This article briefly summarizes seminal discoveries that are shedding new light on the critical and unique roles of autophagy in AD and potential therapeutic approaches against autophagy-elicited AD.

  7. Autophagy: A Potential Link between Obesity and Insulin Resistance

    NARCIS (Netherlands)

    P. Codogno; A.J. Meijer

    2010-01-01

    Dysregulation of autophagy contributes to aging and to diseases such as neurodegeneration, cardiomyopathy, and cancer. The paper by Yang et al. (2010) in this issue of Cell Metabolism indicates that defective autophagy may also underlie impaired insulin sensitivity in obesity and that upregulating a

  8. Altered autophagy in human adipose tissues in obesity

    Science.gov (United States)

    Context: Autophagy is a housekeeping mechanism, involved in metabolic regulation and stress response, shown recently to regulate lipid droplets biogenesis/breakdown and adipose tissue phenotype. Objective: We hypothesized that in human obesity autophagy may be altered in adipose tissue in a fat d...

  9. Autophagy in ageing and ageing-associated diseases

    Institute of Scientific and Technical Information of China (English)

    Li-qiang HE; Jia-hong LU; Zhen-yu YUE

    2013-01-01

    Autophagy is a cell self-digestion process via lysosomes that clears "cellular waste",including aberrantly modified proteins or protein aggregates and damaged organelles.Therefore,autophagy is considered a protein and organelle quality control mechanism that maintains normal cellular homeostasis.Dysfunctional autophagy has been observed in ageing tissues and several ageing-associated diseases.Lifespan of model organisms such as yeast,worms,flies,and mice can be extended through promoting autophagy,either by genetic manipulations such as over-expression of Sirtuin 1,or by administrations of rapamycin,resveratrol or spermidine.The evidence supports that autophagy may play an important role in delaying ageing or extending lifespan.In this review,we summarize the current knowledge about autophagy and its regulation,outline recent developments ie the genetic and pharmacological manipulations of autophagy that affects the lifespan,and discuss the role of autophagy in the ageing-related diseases.ow in Center for Neurodegenerative and Neuroimmunologic Diseases,Department of Neurology,University of Medicine and Dentistry of New Jersey-Robert Wood Johnson Medical School,Piscataway,NJ 08854,USA

  10. Polymorphisms in autophagy genes and susceptibility to tuberculosis.

    NARCIS (Netherlands)

    Songane, M.; Kleinnijenhuis, J.; Alisjahbana, B.; Sahiratmadja, E.; Parwati, I.; Oosting, M.; Plantinga, T.S.; Joosten, L.A.B.; Netea, M.G.; Ottenhoff, T.H.; Vosse, E. van de; Crevel, R. van

    2012-01-01

    Recent data suggest that autophagy is important for intracellular killing of Mycobacterium tuberculosis, and polymorphisms in the autophagy gene IRGM have been linked with susceptibility to tuberculosis (TB) among African-Americans, and with TB caused by particular M. tuberculosis genotypes in Ghana

  11. The ubiquitin ligase HERC3 attenuates NF-κB-dependent transcription independently of its enzymatic activity by delivering the RelA subunit for degradation.

    Science.gov (United States)

    Hochrainer, Karin; Pejanovic, Nadja; Olaseun, Victoria A; Zhang, Sheng; Iadecola, Costantino; Anrather, Josef

    2015-11-16

    Activation of NF-κB-dependent transcription represents an important hallmark of inflammation. While the acute inflammatory response is per se beneficial, it can become deleterious if its spatial and temporal profile is not tightly controlled. Classically, NF-κB activity is limited by cytoplasmic retention of the NF-κB dimer through binding to inhibitory IκB proteins. However, increasing evidence suggests that NF-κB activity can also be efficiently contained by direct ubiquitination of NF-κB subunits. Here, we identify the HECT-domain ubiquitin ligase HERC3 as novel negative regulator of NF-κB activity. We find that HERC3 restricts NF-κB nuclear import and DNA binding without affecting IκBα degradation. Instead HERC3 indirectly binds to the NF-κB RelA subunit after liberation from IκBα inhibitor leading to its ubiquitination and protein destabilization. Remarkably, the regulation of RelA activity by HERC3 is independent of its inherent ubiquitin ligase activity. Rather, we show that HERC3 and RelA are part of a multi-protein complex containing the proteasome as well as the ubiquitin-like protein ubiquilin-1 (UBQLN1). We present evidence that HERC3 and UBQLN1 provide a link between NF-κB RelA and the 26S proteasome, thereby facilitating RelA protein degradation. Our findings establish HERC3 as novel candidate regulating the inflammatory response initiated by NF-κB. PMID:26476452

  12. The ubiquitin conjugating enzyme UbcH7, controls cell migration

    Science.gov (United States)

    Post translational modification by ubiquitination can target proteins for degradation, allow the interaction of proteins to form complexes or direct relocalization of proteins to different subcellular compartments. As such, ubiquitin controls a variety of essential cellular processes. Previously we ...

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

    NARCIS (Netherlands)

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

    2014-01-01

    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,

  14. Omega-3 free fatty acids suppress macrophage inflammasome activation by inhibiting NF-κB activation and enhancing autophagy.

    Directory of Open Access Journals (Sweden)

    Yolanda Williams-Bey

    Full Text Available The omega-3 (ω3 fatty acid docosahexaenoic acid (DHA can suppress inflammation, specifically IL-1β production through poorly understood molecular mechanisms. Here, we show that DHA reduces macrophage IL-1β production by limiting inflammasome activation. Exposure to DHA reduced IL-1β production by ligands that stimulate the NLRP3, AIM2, and NAIP5/NLRC4 inflammasomes. The inhibition required Free Fatty Acid Receptor (FFAR 4 (also known as GPR120, a G-protein coupled receptor (GPR known to bind DHA. The exposure of cells to DHA recruited the adapter protein β-arrestin1/2 to FFAR4, but not to a related lipid receptor. DHA treatment reduced the initial inflammasome priming step by suppressing the nuclear translocation of NF-κB. DHA also reduced IL-1β levels by enhancing autophagy in the cells. As a consequence macrophages derived from mice lacking the essential autophagy protein ATG7 were partially resistant to suppressive effects of DHA. Thus, DHA suppresses inflammasome activation by two distinct mechanisms, inhibiting the initial priming step and by augmenting autophagy, which limits inflammasome activity.

  15. The Ubiquitin-like modifier FAT10 in tolerance induction

    OpenAIRE

    Bürger, Stefanie

    2013-01-01

    HLA-F adjacent transcript 10 (FAT10) is a ubiquitin-like modifier which is predominantly expressed in lymphoid tissue like thymus, lymph node and spleen. The expression of FAT10 is inducible in various cell types with pro-inflammatory cytokines tumor necrosis factor-alpha and interferon-gamma. FAT10 is able to target proteins for proteasomal degradation, but unlike ubiquitin, it is degraded along with its substrates. Only a few biological functions of FAT10 have been described, for instance, ...

  16. FAT10, a Ubiquitin-Independent Signal for Proteasomal Degradation

    OpenAIRE

    Hipp, Mark Steffen; Kalveram, Birte; Raasi, Shahri; Groettrup, Marcus; Schmidtke, Gunter

    2005-01-01

    FAT10 is a small ubiquitin-like modifier that is encoded in the major histocompatibility complex and is synergistically inducible by tumor necrosis factor alpha and gamma interferon. It is composed of two ubiquitin-like domains and possesses a free C-terminal diglycine motif that is required for the formation of FAT10 conjugates. Here we show that unconjugated FAT10 and a FAT10 conjugate were rapidly degraded by the proteasome at a similar rate. Fusion of FAT10 to the N terminus of very long-...

  17. Autophagy in Plants--What's New on the Menu?

    Science.gov (United States)

    Michaeli, Simon; Galili, Gad; Genschik, Pascal; Fernie, Alisdair R; Avin-Wittenberg, Tamar

    2016-02-01

    Autophagy is a major cellular degradation pathway in eukaryotes. Recent studies have revealed the importance of autophagy in many aspects of plant life, including seedling establishment, plant development, stress resistance, metabolism, and reproduction. This is manifested by the dual ability of autophagy to execute bulk degradation under severe environmental conditions, while simultaneously to be highly selective in targeting specific compartments and protein complexes to regulate key cellular processes, even during favorable growth conditions. Delivery of cellular components to the vacuole enables their recycling, affecting the plant metabolome, especially under stress. Recent research in Arabidopsis has further unveiled fundamental mechanistic aspects in autophagy which may have relevance in non-plant systems. We review the most recent discoveries concerning autophagy in plants, touching upon all these aspects.

  18. Targeting autophagy to sensitive glioma to temozolomide treatment.

    Science.gov (United States)

    Yan, Yuanliang; Xu, Zhijie; Dai, Shuang; Qian, Long; Sun, Lunquan; Gong, Zhicheng

    2016-02-02

    Temozolomide (TMZ), an alkylating agent, is widely used for treating primary and recurrent high-grade gliomas. However, the efficacy of TMZ is often limited by the development of resistance. Recently, studies have found that TMZ treatment could induce autophagy, which contributes to therapy resistance in glioma. To enhance the benefit of TMZ in the treatment of glioblastomas, effective combination strategies are needed to sensitize glioblastoma cells to TMZ. In this regard, as autophagy could promote cell survival or autophagic cell death, modulating autophagy using a pharmacological inhibitor, such as chloroquine, or an inducer, such as rapamycin, has received considerably more attention. To understand the effectiveness of regulating autophagy in glioblastoma treatment, this review summarizes reports on glioblastoma treatments with TMZ and autophagic modulators from in vitro and in vivo studies, as well as clinical trials. Additionally, we discuss the possibility of using autophagy regulatory compounds that can sensitive TMZ treatment as a chemotherapy for glioma treatment.

  19. Opening new doors in autophagy research: Patrice Codogno.

    Science.gov (United States)

    Codogno, Patrice; Klionsky, Daniel J

    2016-06-01

    Patrice Codogno ( Fig. 1 ), one of the associate editors of Autophagy since it was established, is well known in the autophagy field, and has played a particularly important role in France, serving as the first president of Club Francophone de l'AuTophaGie (CFATG). Patrice's research career spans from the predominantly biochemical analyses that were commonly used in the 1980s to the molecular studies that are the primary focus of many labs currently studying autophagy today. Anyone who has met Patrice knows that he is modest, which means his contributions to autophagy and to promoting the careers of scientists globally, are underappreciated. In addition, there is a fun-loving side to Patrice that is often hidden to the casual observer, and it is time to share some of his personality and thoughts with the rest of the autophagy community. PMID:27158743

  20. Autophagy process is associated with anti-neoplastic function

    Institute of Scientific and Technical Information of China (English)

    Chong Wang; Yachen Wang; Michael A. McNutt; Wei-Guo Zhu

    2011-01-01

    Autophagy is a highly conserved process of cellular degradation, which is present in yeast, plants, and mammals.Under normal physiological conditions, autophagy acts to maintain cellular homeostasis and regulate the turnover of organelles.In response to cellular stresses, autophagy prevents the accumulation of impaired proteins and organelles, which serves to inhibit carcinogenesis.On this basis,it is widely accepted that most tumor suppressors, such as beclin 1 associated proteins, forkhead box class O (FoxO)family proteins, multiple mammalian target of Rapamycin (mTOR) inactivators, and nuclear p53 play a role in indu cing autophagy.Here, we focus on how the process of autophagy is associated with anti-neoplastic function.

  1. Modulation of Autophagy-Like Processes by Tumor Viruses

    Directory of Open Access Journals (Sweden)

    Karl Munger

    2012-06-01

    Full Text Available Autophagy is an intracellular degradation pathway for long-lived proteins and organelles. This process is activated above basal levels upon cell intrinsic or environmental stress and dysregulation of autophagy has been linked to various human diseases, including those caused by viral infection. Many viruses have evolved strategies to directly interfere with autophagy, presumably to facilitate their replication or to escape immune detection. However, in some cases, modulation of autophagy appears to be a consequence of the virus disturbing the cell’s metabolic signaling networks. Here, we summarize recent advances in research at the interface of autophagy and viral infection, paying special attention to strategies that human tumor viruses have evolved.

  2. Characterization of early autophagy signaling by quantitative phosphoproteomics

    DEFF Research Database (Denmark)

    Rigbolt, Kristoffer Tg; Zarei, Mostafa; Sprenger, Adrian;

    2014-01-01

    Under conditions of nutrient shortage autophagy is the primary cellular mechanism ensuring availability of substrates for continuous biosynthesis. Subjecting cells to starvation or rapamycin efficiently induces autophagy by inhibiting the MTOR signaling pathway triggering increased autophagic flux....... To elucidate the regulation of early signaling events upon autophagy induction, we applied quantitative phosphoproteomics characterizing the temporal phosphorylation dynamics after starvation and rapamycin treatment. We obtained a comprehensive atlas of phosphorylation kinetics within the first 30 min upon...... induction of autophagy with both treatments affecting widely different cellular processes. The identification of dynamic phosphorylation already after 2 min demonstrates that the earliest events in autophagy signaling occur rapidly after induction. The data was subjected to extensive bioinformatics analysis...

  3. Host Cell Autophagy in Immune Response to Zoonotic Infections

    Directory of Open Access Journals (Sweden)

    Panagiotis Skendros

    2012-01-01

    Full Text Available Autophagy is a fundamental homeostatic process in which cytoplasmic targets are sequestered within double-membraned autophagosomes and subsequently delivered to lysosomes for degradation. Accumulating evidence supports the pivotal role of autophagy in host defense against intracellular pathogens implicating both innate and adaptive immunity. Many of these pathogens cause common zoonotic infections worldwide. The induction of the autophagic machinery by innate immune receptors signaling, such as TLRs, NOD1/2, and p62/SQSTM1 in antigen-presenting cells results in inhibition of survival and elimination of invading pathogens. Furthermore, Th1 cytokines induce the autophagic process, whereas autophagy also contributes to antigen processing and MHC class II presentation, linking innate to adaptive immunity. However, several pathogens have developed strategies to avoid autophagy or exploit autophagic machinery to their advantage. This paper focuses on the role of host cell autophagy in the regulation of immune response against intracellular pathogens, emphasizing on selected bacterial and protozoan zoonoses.

  4. Autophagy in Plants--What's New on the Menu?

    Science.gov (United States)

    Michaeli, Simon; Galili, Gad; Genschik, Pascal; Fernie, Alisdair R; Avin-Wittenberg, Tamar

    2016-02-01

    Autophagy is a major cellular degradation pathway in eukaryotes. Recent studies have revealed the importance of autophagy in many aspects of plant life, including seedling establishment, plant development, stress resistance, metabolism, and reproduction. This is manifested by the dual ability of autophagy to execute bulk degradation under severe environmental conditions, while simultaneously to be highly selective in targeting specific compartments and protein complexes to regulate key cellular processes, even during favorable growth conditions. Delivery of cellular components to the vacuole enables their recycling, affecting the plant metabolome, especially under stress. Recent research in Arabidopsis has further unveiled fundamental mechanistic aspects in autophagy which may have relevance in non-plant systems. We review the most recent discoveries concerning autophagy in plants, touching upon all these aspects. PMID:26598298

  5. Role of autophagy in the regulation of epithelial cell junctions.

    Science.gov (United States)

    Nighot, Prashant; Ma, Thomas

    2016-01-01

    Autophagy is a cell survival mechanism by which bulk cytoplasmic material, including soluble macromolecules and organelles, is targeted for lysosomal degradation. The role of autophagy in diverse cellular processes such as metabolic stress, neurodegeneration, cancer, aging, immunity, and inflammatory diseases is being increasingly recognized. Epithelial cell junctions play an integral role in the cell homeostasis via physical binding, regulating paracellular pathways, integrating extracellular cues into intracellular signaling, and cell-cell communication. Recent data indicates that cell junction composition is very dynamic. The junctional protein complexes are actively regulated in response to various intra- and extra-cellular clues by intracellular trafficking and degradation pathways. This review discusses the recent and emerging information on how autophagy regulates various epithelial cell junctions. The knowledge of autophagy regulation of epithelial junctions will provide further rationale for targeting autophagy in a wide variety of human disease conditions. PMID:27583189

  6. Noncanonical Autophagy Is Required for Type I Interferon Secretion in Response to DNA-Immune Complexes

    Science.gov (United States)

    Riggs, Jeffrey M.; Tian, Jane; Mehta, Payal; Clarke, Lorraine; Sasai, Miwa; Latz, Eicke; Brinkmann, Melanie M.; Iwasaki, Akiko; Coyle, Anthony J.; Kolbeck, Roland

    2013-01-01

    SUMMARY Toll-like receptor-9 (TLR9) is largely responsible for discriminating self from pathogenic DNA. However, association of host DNA with autoantibodies activates TLR9, inducing the pathogenic secretion of type I interferons (IFNs) from plasmacytoid dendritic cells (pDCs). Here, we found that in response to DNA-containing immune complexes (DNA-IC), but not to soluble ligands, IFN-α production depended upon the convergence of the phagocytic and autophagic pathways, a process called microtubule-associated protein 1A/1B-light chain 3 (LC3)-associated phagocytosis (LAP). LAP was required for TLR9 trafficking into a specialized interferon signaling compartment by a mechanism that involved autophagy-related proteins, but not the conventional autophagic preinitiation complex, or adaptor protein-3 (AP-3). Our findings unveil a new role for nonconventional autophagy in inflammation and provide one mechanism by which anti-DNA autoantibodies, such as those found in several autoimmune disorders, bypass the controls that normally restrict the apportionment of pathogenic DNA and TLR9 to the interferon signaling compartment. PMID:23219390

  7. Estrogens, inflammation and cognition.

    Science.gov (United States)

    Au, April; Feher, Anita; McPhee, Lucy; Jessa, Ailya; Oh, Soojin; Einstein, Gillian

    2016-01-01

    The effects of estrogens are pleiotropic, affecting multiple bodily systems. Changes from the body's natural fluctuating levels of estrogens, through surgical removal of the ovaries, natural menopause, or the administration of exogenous estrogens to menopausal women have been independently linked to an altered immune profile, and changes to cognitive processes. Here, we propose that inflammation may mediate the relationship between low levels of estrogens and cognitive decline. In order to determine what is known about this connection, we review the literature on the cognitive effects of decreased estrogens due to oophorectomy or natural menopause, decreased estrogens' role on inflammation--both peripherally and in the brain--and the relationship between inflammation and cognition. While this review demonstrates that much is unknown about the intersection between estrogens, cognition, inflammation, we propose that there is an important interaction between these literatures.

  8. Extralysosomal turnover of cellular proteins: Targeting substrates in the ubiquitin, ATP-dependent degradation system

    Energy Technology Data Exchange (ETDEWEB)

    Marriott, D.

    1988-01-01

    Calmodulin derived from a cloned chicken gene can be ubiquitinated and degraded by an in vitro reticulocyte lysate system. The chemical reactivity and the surface accessibility of the {epsilon}-amino group on lysine 115 in the calmodulin polypeptide chain were studied by trace labeling with acetic anhydride and with a ubiquitin derivative containing an azido group at the C-terminal glycine residue. Fractionation of reticulocyte lysate proteins separated the activity which degrades the calmodulin moiety of ubiquitin-calmodulin conjugates from that which acts on the isopeptide linkage. Neither of these two activities act on a synthetic isopeptide, which mimics the junction of ubiquitin-calmodulin, indicating the importance of the folding of ubiquitin for recognition. Based on recent findings that the ubiquitin moieties linked to {beta}galactosidase exist as a single multiubiquitin chain, studies were carried out to determine the structure of the ubiquitin-ubiquitin linkage. Ubiquitin was in vivo labeled with ({sup 3}H) and conjugated to {beta}galactosidase. Individual conjugates were isolated and subjected to peptide mapping by trypsin digestion, and tryptic fragments were analyzed of HPLC. The results indicated that the ubiquitin-ubiquitin linkage involves lysine residue 48 in the ubiquitin sequence.

  9. UBPY-mediated epidermal growth factor receptor (EGFR) de-ubiquitination promotes EGFR degradation

    NARCIS (Netherlands)

    Alwan, H.A.J.; Leeuwen, J.E.M. van

    2007-01-01

    Whereas poly-ubiquitination targets protein substrates for proteasomal degradation, mono-ubiquitination is known to regulate protein trafficking in the endosomal system and to target cargo proteins for lysosomal degradation. The role of the de-ubiquitinating enzymes AMSH and UBPY in endosomal traffi

  10. Selectivity of E2-E3 interactions in the human ubiquitin system

    NARCIS (Netherlands)

    van Wijk, S.J.L.

    2010-01-01

    Highly selective interactions between ubiquitin-conjugating enzymes and RING-type E3 ligases are crucial for the adequate and efficient action of ubiquitin and ubiquitin-like pathways. Within these cascades, E2 enzymes provide a connecting link between activation and the final covalent conjugation,

  11. Orbital inflammation: Corticosteroids first.

    Science.gov (United States)

    Dagi Glass, Lora R; Freitag, Suzanne K

    2016-01-01

    Orbital inflammation is common, and may affect all ages and both genders. By combining a thorough history and physical examination, targeted ancillary laboratory testing and imaging, a presumptive diagnosis can often be made. Nearly all orbital inflammatory pathology can be empirically treated with corticosteroids, thus obviating the need for histopathologic diagnosis prior to initiation of therapy. In addition, corticosteroids may be effective in treating concurrent systemic disease. Unless orbital inflammation responds atypically or incompletely, patients can be spared biopsy.

  12. Genetic animal models for evaluating the role of autophagy in etiopathogenesis of Parkinson disease.

    Science.gov (United States)

    Lachenmayer, M Lenard; Yue, Zhenyu

    2012-12-01

    Parkinson disease (PD) is the most common neurodegenerative movement disorder and is characterized pathologically by the formation of ubiquitin and SNCA/α-synuclein-containing inclusions (Lewy bodies), dystrophic midbrain dopaminergic (DAergic) terminals, and degeneration of midbrain DAergic neurons. The vast majority of PD occurs sporadically, while approximately 5% of all PD cases are inherited. Genetic mutations of a few genes have been identified as causes of familiar PD, i.e., mutations in SNCA, PARK2/parkin, UCHL1, PARK7/DJ1, PINK1 and LRRK2, leading to DAergic cell death, but variable pathological changes. The evidence supports the hypothesis that several pathogenic mechanisms are likely involved at initial stages of the disease, and eventually they merge to cause parkinsonism. The current challenge facing PD research is to unravel the components in these pathways that contribute to the pathogenesis of PD. Accumulating evidence has implicated dysfunctional autophagy, a regulated lysosomal pathway with a capacity for clearing protein aggregates and cellular organelles, as one of the pathogenic systems contributing to the development of idiopathic PD. PMID:22931754

  13. A Ubl/ubiquitin switch in the activation of Parkin.

    Science.gov (United States)

    Sauvé, Véronique; Lilov, Asparouh; Seirafi, Marjan; Vranas, Marta; Rasool, Shafqat; Kozlov, Guennadi; Sprules, Tara; Wang, Jimin; Trempe, Jean-François; Gehring, Kalle

    2015-10-14

    Mutations in Parkin and PINK1 cause an inherited early-onset form of Parkinson's disease. The two proteins function together in a mitochondrial quality control pathway whereby PINK1 accumulates on damaged mitochondria and activates Parkin to induce mitophagy. How PINK1 kinase activity releases the auto-inhibited ubiquitin ligase activity of Parkin remains unclear. Here, we identify a binding switch between phospho-ubiquitin (pUb) and the ubiquitin-like domain (Ubl) of Parkin as a key element. By mutagenesis and SAXS, we show that pUb binds to RING1 of Parkin at a site formed by His302 and Arg305. pUb binding promotes disengagement of the Ubl from RING1 and subsequent Parkin phosphorylation. A crystal structure of Parkin Δ86-130 at 2.54 Å resolution allowed the design of mutations that specifically release the Ubl domain from RING1. These mutations mimic pUb binding and promote Parkin phosphorylation. Measurements of the E2 ubiquitin-conjugating enzyme UbcH7 binding to Parkin and Parkin E3 ligase activity suggest that Parkin phosphorylation regulates E3 ligase activity downstream of pUb binding. PMID:26254305

  14. Strategies to Detect Endogenous Ubiquitination of a Target Mammalian Protein.

    Science.gov (United States)

    Sigismund, Sara; Polo, Simona

    2016-01-01

    Different biochemical techniques are well established to investigate target's ubiquitination in mammals without overexpressing a tagged version of ubiquitin (Ub). The simplest and more direct approach is to immunoprecipitate (IP) your target protein from cell lysate (stimulated and/or properly treated), followed by western blot analysis utilizing specific antibodies against Ub (see Subheading 3.1). This approach requires a good antibody against the target working in IP; alternatively, one could express a tagged version of the protein, possibly at the endogenous level. Another approach consists in IP ubiquitinated proteins from total cell lysate followed by detection with the antibody against the protein of interest. This second method relies on the availability of specific and very efficient antibodies against Ub (see Subheading 3.2). A more quantitative approach is the DELFIA assay (Perkin Elmer), an ELISA-based assay, which allows comparing more samples and conditions (see Subheading 3.3). Cross-validation with more than one approach is usually recommended in order to prove that your protein is modified by ubiquitin.Here we will use the EGFR as model system but protocols can be easily modified according to the protein of interest. PMID:27613032

  15. Mechanism for the selective conjugation of ubiquitin to phytochrome

    Energy Technology Data Exchange (ETDEWEB)

    Vierstra, R.D.

    1990-01-01

    The goal of this project is to understand at the molecular level how phytochrome functions and how intracellular proteins are degraded. Phytochrome is marked for degradation by covalent attachment of ubiquitin. Ubiquitin-phytochrome conjugates (UbP) were characterized with respect to formation kinetics, subcellular localization and site of ubiquitin attachment. UbP appears to be a general phenomenon during phytochrome degradation in a variety of species. UbP was isolated from oat seedlings and characterized. Residues 747-830 of phytochrome have been identified as a possible attachment site for ubiquitin. By placing the gene for etiolated phytochrome in tobacco we have created a transgenic system for over expressing phytochrome. The effects of this over expression are described, and it appears that tobacco degrades this foreign protein through formation of UbP. We have created a series of site-directed mutants of the oat phytochrome gene, and are in the process of characterizing them to determine sequence requirements for ubiquination. 8 refs., 1 fig. (MHB)

  16. How the ubiquitin proteasome system regulates the regulators of transcription.

    Science.gov (United States)

    Ee, Gary; Lehming, Norbert

    2012-01-01

    The ubiquitin proteasome system plays an important role in transcription. Monoubiquitination of activators is believed to aid their function, while the 26S proteasomal degradation of repressors is believed to restrict their function. What remains controversial is the question of whether the degradation of activators aids or restricts their function.

  17. Autophagy protects monocytes from Wolbachia heat shock protein 60-induced apoptosis and senescence.

    Directory of Open Access Journals (Sweden)

    Vijayan Kamalakannan

    2015-04-01

    Full Text Available Monocyte dysfunction by filarial antigens has been a major mechanism underlying immune evasion following hyporesponsiveness during patent lymphatic filariasis. Recent studies have initiated a paradigm shift to comprehend the immunological interactions of Wolbachia and its antigens in inflammation, apoptosis, lymphocyte anergy, etc. Here we showed that recombinant Wolbachia heat shock protein 60 (rWmhsp60 interacts with TLR-4 and induces apoptosis in monocytes of endemic normal but not in chronic patients. Higher levels of reactive oxygen species (ROS induced after TLR-4 stimulation resulted in loss of mitochondrial membrane potential and caspase cascade activation, which are the plausible reason for apoptosis. Furthermore, release in ROS owing to TLR-4 signaling resulted in the activation of NF-κB p65 nuclear translocation which leads to inflammation and apoptosis via TNF receptor pathway following the increase in IL-6 and TNF-α level. Here for the first time, we report that in addition to apoptosis, rWmhsp60 antigen in filarial pathogenesis also induces molecular senescence in monocytes. Targeting TLR-4, therefore, presents a promising candidate for treating rWmhsp60-induced apoptosis and senescence. Strikingly, induction of autophagy by rapamycin detains TLR-4 in late endosomes and subverts TLR-4-rWmhsp60 interaction, thus protecting TLR-4-mediated apoptosis and senescence. Furthermore, rapamycin-induced monocytes were unresponsive to rWmhsp60, and activated lymphocytes following PHA stimulation. This study demonstrates that autophagy mediates the degradation of TLR-4 signaling and protects monocytes from rWmhsp60 induced apoptosis and senescence.

  18. Autophagy protects monocytes from Wolbachia heat shock protein 60-induced apoptosis and senescence.

    Science.gov (United States)

    Kamalakannan, Vijayan; Shiny, Abijit; Babu, Subash; Narayanan, Rangarajan Badri

    2015-04-01

    Monocyte dysfunction by filarial antigens has been a major mechanism underlying immune evasion following hyporesponsiveness during patent lymphatic filariasis. Recent studies have initiated a paradigm shift to comprehend the immunological interactions of Wolbachia and its antigens in inflammation, apoptosis, lymphocyte anergy, etc. Here we showed that recombinant Wolbachia heat shock protein 60 (rWmhsp60) interacts with TLR-4 and induces apoptosis in monocytes of endemic normal but not in chronic patients. Higher levels of reactive oxygen species (ROS) induced after TLR-4 stimulation resulted in loss of mitochondrial membrane potential and caspase cascade activation, which are the plausible reason for apoptosis. Furthermore, release in ROS owing to TLR-4 signaling resulted in the activation of NF-κB p65 nuclear translocation which leads to inflammation and apoptosis via TNF receptor pathway following the increase in IL-6 and TNF-α level. Here for the first time, we report that in addition to apoptosis, rWmhsp60 antigen in filarial pathogenesis also induces molecular senescence in monocytes. Targeting TLR-4, therefore, presents a promising candidate for treating rWmhsp60-induced apoptosis and senescence. Strikingly, induction of autophagy by rapamycin detains TLR-4 in late endosomes and subverts TLR-4-rWmhsp60 interaction, thus protecting TLR-4-mediated apoptosis and senescence. Furthermore, rapamycin-induced monocytes were unresponsive to rWmhsp60, and activated lymphocytes following PHA stimulation. This study demonstrates that autophagy mediates the degradation of TLR-4 signaling and protects monocytes from rWmhsp60 induced apoptosis and senescence. PMID:25849993

  19. Canonical autophagy does not contribute to cellular radioresistance

    International Nuclear Information System (INIS)

    Background: (Pre)clinical studies indicate that autophagy inhibition increases response to anti-cancer therapies. Although promising, due to contradicting reports, it remains unclear if radiation therapy changes autophagy activity and if autophagy inhibition changes the cellular intrinsic radiosensitivity. Discrepancies may result from different assays and models through off-target effects and influencing other signaling routes. In this study, we directly compared the effects of genetic and pharmacological inhibition of autophagy after irradiation in human cancer cell lines. Materials and methods: Changes in autophagy activity after ionizing radiation (IR) were assessed by flux analysis in eight cell lines. Clonogenic survival, DNA damage (COMET-assay) and H2AX phosphorylation were assessed after chloroquine or 3-methyladenine pretreatment and after ATG7 or LC3b knockdown. Results: IR failed to induce autophagy and chloroquine failed to change intrinsic radiosensitivity of cells. Interestingly, 3-methyladenine and ATG7- or LC3b-deficiency sensitized cancer cells to irradiation. Surprisingly, the radiosensitizing effect of 3-methyladenine was also observed in ATG7 and LC3b deficient cells and was associated with attenuated γ-H2AX formation and DNA damage repair. Conclusion: Our data demonstrate that the anti-tumor effects of chloroquine are independent of changes in intrinsic radioresistance. Furthermore, ATG7 and LC3b support radioresistance independent of canonical autophagy that involves lysosomal degradation

  20. Transcriptional regulation of mammalian autophagy at a glance.

    Science.gov (United States)

    Füllgrabe, Jens; Ghislat, Ghita; Cho, Dong-Hyung; Rubinsztein, David C

    2016-08-15

    Macroautophagy, hereafter referred to as autophagy, is a catabolic process that results in the lysosomal degradation of cytoplasmic contents ranging from abnormal proteins to damaged cell organelles. It is activated  under diverse conditions, including nutrient deprivation and hypoxia. During autophagy, members of the core autophagy-related (ATG) family of proteins mediate membrane rearrangements, which lead to the engulfment and degradation of cytoplasmic cargo. Recently, the nuclear regulation of autophagy, especially by transcription factors and histone modifiers, has gained increased attention. These factors are not only involved in rapid responses to autophagic stimuli, but also regulate the long-term outcome of autophagy. Now there are more than 20 transcription factors that have been shown to be linked to the autophagic process. However, their interplay and timing appear enigmatic as several have been individually shown to act as major regulators of autophagy. This Cell Science at a Glance article and the accompanying poster highlights the main cellular regulators of transcription involved in mammalian autophagy and their target genes. PMID:27528206

  1. Targeting autophagy in cancer management – strategies and developments

    Directory of Open Access Journals (Sweden)

    Ozpolat B

    2015-09-01

    Full Text Available Bulent Ozpolat,1 Doris M Benbrook2 1Department of Experimental Therapeutics, The University of Texas – Houston, MD Anderson Cancer Center, Houston, TX, 2Department of Obstetrics and Gynecology, University of Oklahoma HSC, Oklahoma City, OK, USA Abstract: Autophagy is a highly regulated catabolic process involving lysosomal degradation of intracellular components, damaged organelles, misfolded proteins, and toxic aggregates, reducing oxidative stress and protecting cells from damage. The process is also induced in response to various conditions, including nutrient deprivation, metabolic stress, hypoxia, anticancer therapeutics, and radiation therapy to adapt cellular conditions for survival. Autophagy can function as a tumor suppressor mechanism in normal cells and dysregulation of this process (ie, monoallelic Beclin-1 deletion may lead to malignant transformation and carcinogenesis. In tumors, autophagy is thought to promote tumor growth and progression by helping cells to adapt and survive in metabolically-challenged and harsh tumor microenvironments (ie, hypoxia and acidity. Recent in vitro and in vivo studies in preclinical models suggested that modulation of autophagy can be used as a therapeutic modality to enhance the efficacy of conventional therapies, including chemo and radiation therapy. Currently, more than 30 clinical trials are investigating the effects of autophagy inhibition in combination with cytotoxic chemotherapies and targeted agents in various cancers. In this review, we will discuss the role, molecular mechanism, and regulation of autophagy, while targeting this process as a novel therapeutic modality, in various cancers. Keywords: autophagy inhibition, chemotherapy, tumor microenvironment

  2. The Regulation of Autophagy by Influenza A Virus

    Directory of Open Access Journals (Sweden)

    Rong Zhang

    2014-01-01

    Full Text Available Influenza A virus is a dreadful pathogen of animals and humans, causing widespread infection and severe morbidity and mortality. It is essential to characterize the influenza A virus-host interaction and develop efficient counter measures against the viral infection. Autophagy is known as a catabolic process for the recycling of the cytoplasmic macromolecules. Recently, it has been shown that autophagy is a critical mechanism underlying the interaction between influenza A virus and its host. Autophagy can be induced by the infection with influenza A virus, which is considered as a necessary process for the viral proliferation, including the accumulation of viral elements during the replication of influenza A virus. On the other hand, influenza A virus can inhibit the autophagic formation via interaction with the autophagy-related genes (Atg and signaling pathways. In addition, autophagy is involved in the influenza virus-regulated cell deaths, leading to significant changes in host apoptosis. Interestingly, the high pathogenic strains of influenza A virus, such as H5N1, stimulate autophagic cell death and appear to interplay with the autophagy in distinct ways as compared with low pathogenic strains. This review discusses the regulation of autophagy, an influenza A virus driven process.

  3. Autophagy and Transporter-Based Multi-Drug Resistance

    Directory of Open Access Journals (Sweden)

    Zhe-Sheng Chen

    2012-08-01

    Full Text Available All the therapeutic strategies for treating cancers aim at killing the cancer cells via apoptosis (programmed cell death type I. Defective apoptosis endow tumor cells with survival. The cell can respond to such defects with autophagy. Autophagy is a cellular process by which cytoplasmic material is either degraded to maintain homeostasis or recycled for energy and nutrients in starvation. A plethora of evidence has shown that the role of autophagy in tumors is complex. A lot of effort is needed to underline the functional status of autophagy in tumor progression and treatment, and elucidate how to tweak autophagy to treat cancer. Furthermore, during the treatment of cancer, the limitation for the cure rate and survival is the phenomenon of multi drug resistance (MDR. The development of MDR is an intricate process that could be regulated by drug transporters, enzymes, anti-apoptotic genes or DNA repair mechanisms. Reports have shown that autophagy has a dual role in MDR. Furthermore, it has been reported that activation of a death pathway may overcome MDR, thus pointing the importance of other death pathways to regulate tumor cell progression and growth. Therefore, in this review we will discuss the role of autophagy in MDR tumors and a possible link amongst these phenomena.

  4. Laser stimulation can activate autophagy in HeLa cells

    Science.gov (United States)

    Wang, Yisen; Lan, Bei; He, Hao; Hu, Minglie; Cao, Youjia; Wang, Chingyue

    2014-10-01

    For decades, lasers have been a daily tool in most biological research for fluorescent excitation by confocal or multiphoton microscopy. More than 20 years ago, cell photodamage caused by intense laser stimulation was noticed by generating reactive oxygen species, which was then thought as the main damage effect by photons. In this study, we show that laser stimulation can induce autophagy, an important cell lysosomal pathway responding to immune stimulation and starvation, without any biochemical treatment. Two different types of laser stimulations are found to be capable of activating autophagy: continuous scanning by continuous-wave visible lasers and a short-time flash of femtosecond laser irradiation. The autophagy generation is independent from wavelength, power, and scanning duration of the visible lasers. In contrast, the power of femtosecond laser is very critical to autophagy because the multiphoton excited Ca2+ dominates autophagy signaling. In general, we show here the different mechanisms of autophagy generation by such laser stimulation, which correspond to confocal microscopy and cell surgery, respectively. Those results can help further understanding of photodamage and autophagy signaling.

  5. TOR-dependent post-transcriptional regulation of autophagy.

    Science.gov (United States)

    Hu, Guowu; McQuiston, Travis; Bernard, Amélie; Park, Yoon-Dong; Qiu, Jin; Vural, Ali; Zhang, Nannan; Waterman, Scott R; Blewett, Nathan H; Myers, Timothy G; Maraia, Richard J; Kehrl, John H; Uzel, Gulbu; Klionsky, Daniel J; Williamson, Peter R

    2015-01-01

    Regulation of autophagy is required to maintain cellular equilibrium and prevent disease. While extensive study of post-translational mechanisms has yielded important insights into autophagy induction, less is known about post-transcriptional mechanisms that could potentiate homeostatic control. In our study, we showed that the RNA-binding protein, Dhh1 in Saccharomyces cerevisiae and Vad1 in the pathogenic yeast Cryptococcus neoformans is involved in recruitment and degradation of key autophagy mRNAs. In addition, phosphorylation of the decapping protein Dcp2 by the target of rapamycin (TOR), facilitates decapping and degradation of autophagy-related mRNAs, resulting in repression of autophagy under nutrient-replete conditions. The post-transcriptional regulatory process is conserved in both mouse and human cells and plays a role in autophagy-related modulation of the inflammasome product IL1B. These results were then applied to provide mechanistic insight into autoimmunity of a patient with a PIK3CD/p110δ gain-of-function mutation. These results thus identify an important new post-transcriptional mechanism of autophagy regulation that is highly conserved between yeast and mammals.

  6. Nanomaterials and Autophagy: New Insights in Cancer Treatment

    Energy Technology Data Exchange (ETDEWEB)

    Panzarini, Elisa; Inguscio, Valentina; Tenuzzo, Bernardetta Anna; Carata, Elisabetta; Dini, Luciana, E-mail: luciana.dini@unisalento.it [Department of Biological and Environmental Science and Technology (Di.S.Te.B.A.), University of Salento, Lecce 73100 (Italy)

    2013-03-21

    Autophagy represents a cell’s response to stress. It is an evolutionarily conserved process with diversified roles. Indeed, it controls intracellular homeostasis by degradation and/or recycling intracellular metabolic material, supplies energy, provides nutrients, eliminates cytotoxic materials and damaged proteins and organelles. Moreover, autophagy is involved in several diseases. Recent evidences support a relationship between several classes of nanomaterials and autophagy perturbation, both induction and blockade, in many biological models. In fact, the autophagic mechanism represents a common cellular response to nanomaterials. On the other hand, the dynamic nature of autophagy in cancer biology is an intriguing approach for cancer therapeutics, since during tumour development and therapy, autophagy has been reported to trigger both an early cell survival and a late cell death. The use of nanomaterials in cancer treatment to deliver chemotherapeutic drugs and target tumours is well known. Recently, autophagy modulation mediated by nanomaterials has become an appealing notion in nanomedicine therapeutics, since it can be exploited as adjuvant in chemotherapy or in the development of cancer vaccines or as a potential anti-cancer agent. Herein, we summarize the effects of nanomaterials on autophagic processes in cancer, also considering the therapeutic outcome of synergism between nanomaterials and autophagy to improve existing cancer therapies.

  7. Laser stimulation can activate autophagy in HeLa cells

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Yisen; Hu, Minglie; Wang, Chingyue [Ultrafast Laser Laboratory, Key Laboratory of Optoelectronic Information Technology (Ministry of Education), College of Precision Instrument and Optoelectronics Engineering, Tianjin University, Tianjin (China); Lan, Bei; Cao, Youjia [Key Laboratory of Microbial Functional Genomics of Ministry of Education, College of Life Sciences, Nankai University, Tianjin (China); He, Hao, E-mail: haohe@tju.edu.cn [Ultrafast Laser Laboratory, Key Laboratory of Optoelectronic Information Technology (Ministry of Education), College of Precision Instrument and Optoelectronics Engineering, Tianjin University, Tianjin (China); Med-X Research Institute, School of Biomedical Engineering, Shanghai Jiao Tong University, Shanghai (China)

    2014-10-27

    For decades, lasers have been a daily tool in most biological research for fluorescent excitation by confocal or multiphoton microscopy. More than 20 years ago, cell photodamage caused by intense laser stimulation was noticed by generating reactive oxygen species, which was then thought as the main damage effect by photons. In this study, we show that laser stimulation can induce autophagy, an important cell lysosomal pathway responding to immune stimulation and starvation, without any biochemical treatment. Two different types of laser stimulations are found to be capable of activating autophagy: continuous scanning by continuous-wave visible lasers and a short-time flash of femtosecond laser irradiation. The autophagy generation is independent from wavelength, power, and scanning duration of the visible lasers. In contrast, the power of femtosecond laser is very critical to autophagy because the multiphoton excited Ca{sup 2+} dominates autophagy signaling. In general, we show here the different mechanisms of autophagy generation by such laser stimulation, which correspond to confocal microscopy and cell surgery, respectively. Those results can help further understanding of photodamage and autophagy signaling.

  8. YY1-MIR372-SQSTM1 regulatory axis in autophagy.

    Science.gov (United States)

    Feng, Lifeng; Ma, Yanning; Sun, Jie; Shen, Qi; Liu, Leiming; Lu, Haiqi; Wang, Faliang; Yue, Yongfang; Li, Jiaqiu; Zhang, Shenjie; Lin, Xiaoying; Chu, Jue; Han, Weidong; Wang, Xian; Jin, Hongchuan

    2014-08-01

    Autophagy is a self-proteolytic process that degrades intracellular material to enable cellular survival under unfavorable conditions. However, how autophagy is activated in human carcinogenesis remains largely unknown. Herein we report an epigenetic regulation of autophagy in human cancer cells. YY1 (YY1 transcription factor) is a well-known epigenetic regulator and is upregulated in many cancers. We found that YY1 knockdown inhibited cell viability and autophagy flux through downregulating SQSTM1 (sequestosome 1). YY1 regulated SQSTM1 expression through the epigenetic modulation of the transcription of MIR372 (microRNA 372) which was found to target SQSTM1 directly. During nutrient starvation, YY1 was stimulated to promote SQSTM1 expression and subsequent autophagy activation by suppressing MIR372 expression. Similar to YY1 depletion, MIR372 overexpression blocked autophagy activation and inhibited in vivo tumor growth. SQSTM1 upregulation and competent autophagy flux thus contributed to the oncogenic function of YY1. YY1-promoted SQSTM1 upregulation might be a useful histological marker for cancer detection and a potential target for drug development.

  9. Piperlongumine induces autophagy by targeting p38 signaling.

    Science.gov (United States)

    Wang, Y; Wang, J-W; Xiao, X; Shan, Y; Xue, B; Jiang, G; He, Q; Chen, J; Xu, H-G; Zhao, R-X; Werle, K D; Cui, R; Liang, J; Li, Y-L; Xu, Z-X

    2013-01-01

    Piperlongumine (PL), a natural product isolated from the plant species Piper longum L., can selectively induce apoptotic cell death in cancer cells by targeting the stress response to reactive oxygen species (ROS). Here we show that PL induces cell death in the presence of benzyloxycarbonylvalyl-alanyl-aspartic acid (O-methyl)-fluoro-methylketone (zVAD-fmk), a pan-apoptotic inhibitor, and in the presence of necrostatin-1, a necrotic inhibitor. Instead PL-induced cell death can be suppressed by 3-methyladenine, an autophagy inhibitor, and substantially attenuated in cells lacking the autophagy-related 5 (Atg5) gene. We further show that PL enhances autophagy activity without blocking autophagy flux. Application of N-acetyl-cysteine, an antioxidant, markedly reduces PL-induced autophagy and cell death, suggesting an essential role for intracellular ROS in PL-induced autophagy. Furthermore, PL stimulates the activation of p38 protein kinase through ROS-induced stress response and p38 signaling is necessary for the action of PL as SB203580, a p38 inhibitor, or dominant-negative p38 can effectively reduce PL-mediated autophagy. Thus, we have characterized a new mechanism for PL-induced cell death through the ROS-p38 pathway. Our findings support the therapeutic potential of PL by triggering autophagic cell death. PMID:24091667

  10. Autophagy in the immune response to tuberculosis: clinical perspectives.

    LENUS (Irish Health Repository)

    Ní Cheallaigh, C

    2011-06-01

    A growing body of evidence points to autophagy as an essential component in the immune response to tuberculosis. Autophagy is a direct mechanism of killing intracellular Mycobacterium tuberculosis and also acts as a modulator of proinflammatory cytokine secretion. In addition, autophagy plays a key role in antigen processing and presentation. Autophagy is modulated by cytokines; it is stimulated by T helper type 1 (Th1) cytokines such as tumour necrosis factor (TNF)-α and interferon (IFN)-γ, and is inhibited by the Th2 cytokines interleukin (IL)-4 and IL-13 and the anti-inflammatory cytokine IL-10. Vitamin D, via cathelicidin, can also induce autophagy, as can Toll-like receptor (TLR)-mediated signals. Autophagy-promoting agents, administered either locally to the lungs or systemically, could have a clinical application as adjunctive treatment of drug-resistant and drug-sensitive tuberculosis. Moreover, vaccines which effectively induce autophagy could be more successful in preventing acquisition or reactivation of latent tuberculosis.

  11. The interplays between autophagy and apoptosis induced by enterovirus 71.

    Directory of Open Access Journals (Sweden)

    Xueyan Xi

    Full Text Available BACKGROUND: Enterovirus 71 (EV71 is the causative agent of human diseases with distinct severity, from mild hand, foot and mouth disease to severe neurological syndromes, such as encephalitis and meningitis. The lack of understanding of viral pathogenesis as well as lack of efficient vaccine and drugs against this virus impedes the control of EV71 infection. EV71 virus induces autophagy and apoptosis; however, the relationship between EV71-induced autophagy and apoptosis as well as the influence of autophagy and apoptosis on virus virulence remains unclear. METHODOLOGY/PRINCIPAL FINDINGS: In this study, it was observed that the Anhui strain of EV71 induced autophagy and apoptosis in human rhabdomyosarcoma (RD-A cells. Additionally, by either applying chemical inhibitors or knocking down single essential autophagic or apoptotic genes, inhibition of EV71 induced autophagy inhibited the apoptosis both at the autophagosome formation stage and autophagy execution stage. However, inhibition of autophagy at the stage of autophagosome and lysosome fusion promoted apoptosis. In reverse, the inhibition of EV71-induced apoptosis contributed to the conversion of microtubule-associated protein 1 light chain 3-I (LC3-I to LC3-II and degradation of sequestosome 1 (SQSTM1/P62. Furthermore, the inhibition of autophagy in the autophagsome formation stage or apoptosis decreased the release of EV71 viral particles. CONCLUSIONS/SIGNIFICANCE: In conclusion, the results of this study not only revealed novel aspect of the interplay between autophagy and apoptosis in EV71 infection, but also provided a new insight to control EV71 infection.

  12. Autophagy contributes to gefitinib-induced glioma cell growth inhibition

    Energy Technology Data Exchange (ETDEWEB)

    Chang, Cheng-Yi [Department of Surgery, Fong-Yuan Hospital, Taichung 420, Taiwan (China); Graduate Institute of Pharmaceutical Science and Technology, Central Taiwan University of Science and Technology, Taichung 406, Taiwan (China); Kuan, Yu-Hsiang [Department of Pharmacology, School of Medicine, Chung Shan Medical University, Taichung 402, Taiwan (China); Department of Pharmacy, Chung Shan Medical University Hospital, Taichung 402, Taiwan (China); Ou, Yen-Chuan; Li, Jian-Ri [Division of Urology, Taichung Veterans General Hospital, Taichung 407, Taiwan (China); Wu, Chih-Cheng [Department of Anesthesiology, Taichung Veterans General Hospital, Taichung 407, Taiwan (China); Department of Financial and Computational Mathematics, Providence University, Taichung 433, Taiwan (China); Pan, Pin-Ho [Department of Pediatrics, Tungs’ Taichung MetroHarbor Hospital, Taichung 435, Taiwan (China); Chen, Wen-Ying [Department of Veterinary Medicine, National Chung Hsing University, Taichung 402, Taiwan (China); Huang, Hsuan-Yi [Department of Surgery, Fong-Yuan Hospital, Taichung 420, Taiwan (China); Chen, Chun-Jung, E-mail: cjchen@vghtc.gov.tw [Department of Medical Research, Taichung Veterans General Hospital, Taichung 407, Taiwan (China); Institute of Biomedical Sciences, National Chung Hsing University, Taichung 402, Taiwan (China); Rong Hsing Research Center for Translational Medicine, National Chung Hsing University, Taichung 402, Taiwan (China); Center for General Education, Tunghai University, Taichung 407, Taiwan (China); Department of Nursing, HungKuang University, Taichung 433, Taiwan (China)

    2014-09-10

    Epidermal growth factor receptor tyrosine kinase inhibitors, including gefitinib, have been evaluated in patients with malignant gliomas. However, the molecular mechanisms involved in gefitinib-mediated anticancer effects against glioma are incompletely understood. In the present study, the cytostatic potential of gefitinib was demonstrated by the inhibition of glioma cell growth, long-term clonogenic survival, and xenograft tumor growth. The cytostatic consequences were accompanied by autophagy, as evidenced by monodansylcadaverine staining of acidic vesicle formation, conversion of microtubule-associated protein-1 light chain 3-II (LC3-II), degradation of p62, punctate pattern of GFP-LC3, and conversion of GFP-LC3 to cleaved-GFP. Autophagy inhibitor 3-methyladenosine and chloroquine and genetic silencing of LC3 or Beclin 1 attenuated gefitinib-induced growth inhibition. Gefitinib-induced autophagy was not accompanied by the disruption of the Akt/mammalian target of rapamycin signaling. Instead, the activation of liver kinase-B1/AMP-activated protein kinase (AMPK) signaling correlated well with the induction of autophagy and growth inhibition caused by gefitinib. Silencing of AMPK suppressed gefitinib-induced autophagy and growth inhibition. The crucial role of AMPK activation in inducing glioma autophagy and growth inhibition was further supported by the actions of AMP mimetic AICAR. Gefitinib was shown to be capable of reducing the proliferation of glioma cells, presumably by autophagic mechanisms involving AMPK activation. - Highlights: • Gefitinib causes cytotoxic and cytostatic effect on glioma. • Gefitinib induces autophagy. • Gefitinib causes cytostatic effect through autophagy. • Gefitinib induces autophagy involving AMPK.

  13. Autophagy contributes to gefitinib-induced glioma cell growth inhibition

    International Nuclear Information System (INIS)

    Epidermal growth factor receptor tyrosine kinase inhibitors, including gefitinib, have been evaluated in patients with malignant gliomas. However, the molecular mechanisms involved in gefitinib-mediated anticancer effects against glioma are incompletely understood. In the present study, the cytostatic potential of gefitinib was demonstrated by the inhibition of glioma cell growth, long-term clonogenic survival, and xenograft tumor growth. The cytostatic consequences were accompanied by autophagy, as evidenced by monodansylcadaverine staining of acidic vesicle formation, conversion of microtubule-associated protein-1 light chain 3-II (LC3-II), degradation of p62, punctate pattern of GFP-LC3, and conversion of GFP-LC3 to cleaved-GFP. Autophagy inhibitor 3-methyladenosine and chloroquine and genetic silencing of LC3 or Beclin 1 attenuated gefitinib-induced growth inhibition. Gefitinib-induced autophagy was not accompanied by the disruption of the Akt/mammalian target of rapamycin signaling. Instead, the activation of liver kinase-B1/AMP-activated protein kinase (AMPK) signaling correlated well with the induction of autophagy and growth inhibition caused by gefitinib. Silencing of AMPK suppressed gefitinib-induced autophagy and growth inhibition. The crucial role of AMPK activation in inducing glioma autophagy and growth inhibition was further supported by the actions of AMP mimetic AICAR. Gefitinib was shown to be capable of reducing the proliferation of glioma cells, presumably by autophagic mechanisms involving AMPK activation. - Highlights: • Gefitinib causes cytotoxic and cytostatic effect on glioma. • Gefitinib induces autophagy. • Gefitinib causes cytostatic effect through autophagy. • Gefitinib induces autophagy involving AMPK

  14. Inflammation and keratoconus.

    Science.gov (United States)

    McMonnies, Charles W

    2015-02-01

    Keratoconus (KC) has been traditionally classified as a noninflammatory disease. Barring loss of function, the other classic signs of inflammation (heat, redness, swelling, pain) are not usually obvious or even apparent in KC. This clinical perspective examines the evidence and implications of numerous inflammatory processes that have been recognized in the tears of KC patients as well as some inflammation relevant differences found in the KC cornea. The roles of inflammation in corneal trauma attributed to eye rubbing and/or contact lens wear are examined as is the significance of atopy, allergic disease, dry eye disease, degradative enzyme activity, wound healing, reduced anti-inflammatory capacity, and ultraviolet irradiation. It is possible that any comorbidity that is inflammatory in nature may add synergistically to other forms of KC-related inflammation and exacerbate its pathogenetic processes. For example, some features of inflammation in ocular rosacea and associated corneal thinning and distortion could have some possible relevance to KC. An analogy is drawn with osteoarthritis, which also involves significant inflammatory processes but, like KC, does not meet all the classic criteria for an inflammatory disease. Classifying KC as quasi-inflammatory (inflammatory-related) rather than a noninflammatory disease appears to be more appropriate and may help focus attention on the possibility of developing effective anti-inflammatory therapies for its management. PMID:25397925

  15. A Co-Translational Ubiquitination Pathway For Quality Control of Misfolded Proteins

    Science.gov (United States)

    Wang, Feng; Durfee, Larissa A.; Huibregtse, Jon M.

    2013-01-01

    Previous studies have indicated that 6–30% of all newly synthesized proteins are rapidly degraded by the ubiquitin-proteasome system, however the relationship of ubiquitination to translation for these proteins has been unclear. We report that co-translational ubiquitination (CTU) is a robust process, with ~12–15% of nascent polypeptides being ubiquitinated in human cells. CTU products contained primarily K48-linked polyubiquitin chains, consistent with a proteasomal targeting function. While nascent chains have been shown previously to be ubiquitinated within stalled complexes (CTUS), the majority of nascent chain ubiquitination occurred within active translation complexes (CTUA). CTUA was increased in response to agents that induce protein misfolding, while CTUS was increased in response to agents that lead to translational errors or stalling. These results indicate that ubiquitination of nascent polypeptides occurs in two contexts, and define CTUA as a component of a quality control system that marks proteins for destruction while they are being synthesized. PMID:23583076

  16. The protective roles of autophagy in ischemic preconditioning

    Institute of Scientific and Technical Information of China (English)

    Wen-jun YAN; Hai-long DONG; Li-ze XIONG

    2013-01-01

    Autophagy,a process for the degradation of protein aggregates and dysfunctional organelles,is required for cellular homeostasis and cell survival in response to stress and is implicated in endogenous protection.Ischemic preconditioning is a brief and nonlethal episode of ischemia,confers protection against subsequent ischemia-repenfusion through the up-regulation of endogenous protective mechanisms.Emerging evidence shows that autophagy is associated with the protective effect of ischemic preconditioning.This review summarizes recent progress in research on the functions and regulations of the autophagy pathway in preconditioning-induced protection and cellular survival.

  17. Alternative autophagy, brefeldin A and viral trafficking pathways

    Science.gov (United States)

    Grose, Charles; Klionsky, Daniel J.

    2016-01-01

    ABSTRACT Two topics that have attracted recent attention in the field of autophagy concern the source of the membrane that is used to form the autophagosome during macroautophagy and the role of noncanonical autophagic pathways. The 2 topics may converge when considering the intersection of autophagy with viral infection. We suggest that noncanonical autophagy, which is sensitive to treatment with brefeldin A, may converge with the infectious cycles of certain DNA and RNA viruses that utilize membrane from the ER and cis-Golgi. PMID:27439673

  18. Role of autophagy in acute myeloid leukemia therapy

    Institute of Scientific and Technical Information of China (English)

    Su-Ping Zhang; Yu-Na Niu; Na Yuan; Ai-Hong Zhang; Dan Chao; Qiu-Ping Xu; Li-Jun Wang

    2013-01-01

    Despite its dual role in determining cell fate in a wide array of solid cancer cell lines,autophagy has been robustly shown to suppress or kill acute myeloid leukemia cells via degradation of the oncogenic fusion protein that drives leukemogenesis.However,autophagy also induces the demise of acute leukemia cells that do not express the known fusion protein,though the molecular mechanism remains elusive.Nevertheless,since it can induce cooperation with apoptosis and differentiation in response to autophagic signals,autophagy can be manipulated for a better therapy on acute myeloid leukemia.

  19. Histone Deacetylase Inhibitors Increase p27Kip1 by Affecting Its Ubiquitin-Dependent Degradation through Skp2 Downregulation

    Directory of Open Access Journals (Sweden)

    Adriana Borriello

    2016-01-01

    Full Text Available Histone deacetylase inhibitors (HDACIs represent an intriguing class of pharmacologically active compounds. Currently, some HDACIs are FDA approved for cancer therapy and many others are in clinical trials, showing important clinical activities at well tolerated doses. HDACIs also interfere with the aging process and are involved in the control of inflammation and oxidative stress. In vitro, HDACIs induce different cellular responses including growth arrest, differentiation, and apoptosis. Here, we evaluated the effects of HDACIs on p27Kip1, a key cyclin-dependent kinase inhibitor (CKI. We observed that HDACI-dependent antiproliferative activity is associated with p27Kip1 accumulation due to a reduced protein degradation. p27Kip1 removal requires a preliminary ubiquitination step due to the Skp2-SCF E3 ligase complex. We demonstrated that HDACIs increase p27Kip1 stability through downregulation of Skp2 protein levels. Skp2 decline is only partially due to a reduced Skp2 gene expression. Conversely, the protein decrease is more profound and enduring compared to the changes of Skp2 transcript. This argues for HDACIs effects on Skp2 protein posttranslational modifications and/or on its removal. In summary, we demonstrate that HDACIs increase p27Kip1 by hampering its nuclear ubiquitination/degradation. The findings might be of relevance in the phenotypic effects of these compounds, including their anticancer and aging-modulating activities.

  20. High-mobility group box-1 in sterile inflammation.

    Science.gov (United States)

    Tsung, A; Tohme, S; Billiar, T R

    2014-11-01

    High-mobility group box 1 (HMGB1) was originally defined as a ubiquitous nuclear protein, but it was later determined that the protein has different roles both inside and outside of cells. Nuclear HMGB1 regulates chromatin structure and gene transcription, whereas cytosolic HMGB1 is involved in inflammasome activation and autophagy. Extracellular HMGB1 has drawn attention because it can bind to related cell signalling transduction receptors, such as the receptor for advanced glycation end products, Toll-like receptor (TLR)2, TLR4 and TLR9. It also participates in the development and progression of a variety of diseases. HMGB1 is actively secreted by stimulation of the innate immune system, and it is passively released by ischaemia or cell injury. This review focuses on the important role of HMGB1 in the pathogenesis of acute and chronic sterile inflammatory conditions. Strategies that target HMGB1 have been shown to significantly decrease inflammation in several disease models of sterile inflammation, and this may represent a promising clinical approach for treatment of certain conditions associated with sterile inflammation. PMID:24935761

  1. Astemizole-Histamine induces Beclin-1-independent autophagy by targeting p53-dependent crosstalk between autophagy and apoptosis.

    Science.gov (United States)

    Jakhar, Rekha; Paul, Souren; Bhardwaj, Monika; Kang, Sun Chul

    2016-03-01

    Apoptosis and autophagy are genetically regulated, evolutionarily conserved processes that can jointly seal cancer cell fates, and numerous death stimuli are capable of activating either pathway. Although crosstalk between apoptosis and autophagy is quite complex and sometimes contradictory, it remains a key factor determining the outcomes of death-related pathologies such as cancer. In the present study, exposure of MCF-7 breast cancer cells to HIS and the H1 receptor antagonist AST both alone and together with HIS (AST-HIS) led to generation of intracellular ROS, which induced massive cellular vacuolization through dilation of the ER and mitochondria. Consequently, apoptosis by Bax translocation, cytochrome c release, and caspase activation were triggered. In addition, AST-HIS caused ER stress-induced autophagy in MCF-7 cells, as evidenced by an increased LC3-II/LC3-I ratio, with surprisingly no changes in Beclin-1 expression. Non-canonical autophagy was induced via p53 phosphorylation, which increased p53-p62 interactions to enhance Beclin-1-independent autophagy as evidenced by immunocytochemistry and immunoprecipitation. In the absence of Beclin-1, enhanced autophagy further activated apoptosis through caspase induction. In conclusion, these findings indicate that AST-HIS-induced apoptosis and autophagy can be regulated by ROS-mediated signaling pathways. PMID:26739061

  2. Zymophagy: Selective Autophagy of Secretory Granules

    Directory of Open Access Journals (Sweden)

    Maria I. Vaccaro

    2012-01-01

    Full Text Available Timing is everything. That's especially true when it comes to the activation of enzymes created by the pancreas to break down food. Pancreatic enzymes are packed in secretory granules as precursor molecules called zymogens. In physiological conditions, those zymogens are activated only when they reach the gut, where they get to work releasing and distributing nutrients that we need to survive. If this process fails and the enzymes are prematurely activated within the pancreatic cell, before they are released from the gland, they break down the pancreas itself causing acute pancreatitis. This is a painful disease that ranges from a mild and autolimited process to a severe and lethal condition. Recently, we demonstrated that the pancreatic acinar cell is able to switch on a refined mechanism that could explain the autolimited form of the disease. This is a novel selective form of autophagy named zymophagy, a cellular process to specifically detect and degrade secretory granules containing activated enzymes before they can digest the organ. In this work, we revise the molecules and mechanisms that mediate zymophagy, a selective autophagy of secretory granules.

  3. Chaperone-mediated autophagy: roles in neuroprotection.

    Science.gov (United States)

    Cai, Zhibiao; Zeng, Weijun; Tao, Kai; E, Zhen; Wang, Bao; Yang, Qian

    2015-08-01

    Chaperone-mediated autophagy (CMA), one of the main pathways of lysosomal proteolysis, is characterized by the selective targeting and direct translocation into the lysosomal lumen of substrate proteins containing a targeting motif biochemically related to the pentapeptide KFERQ. Along with the other two lysosomal pathways, macro- and micro-autophagy, CMA is essential for maintaining cellular homeostasis and survival by selectively degrading misfolded, oxidized, or damaged cytosolic proteins. CMA plays an important role in pathologies such as cancer, kidney disorders, and neurodegenerative diseases. Neurons are post-mitotic and highly susceptible to dysfunction of cellular quality-control systems. Maintaining a balance between protein synthesis and degradation is critical for neuronal functions and homeostasis. Recent studies have revealed several new mechanisms by which CMA protects neurons through regulating factors critical for their viability and homeostasis. In the current review, we summarize recent advances in the understanding of the regulation and physiology of CMA with a specific focus on its possible roles in neuroprotection. PMID:26206599

  4. ER stress, autophagy, and RNA viruses

    Directory of Open Access Journals (Sweden)

    Jia-Rong eJheng

    2014-08-01

    Full Text Available Endoplasmic reticulum (ER stress is a general term for representing the pathway by which various stimuli affect ER functions. ER stress induces the evolutionarily conserved signaling pathways, called the unfolded protein response (UPR, which compromises the stimulus and then determines whether the cell survives or dies. In recent years, ongoing research has suggested that these pathways may be linked to the autophagic response, which plays a key role in the cell’s response to various stressors. Autophagy performs a self-digestion function, and its activation protects cells against certain pathogens. However, the link between the UPR and autophagy may be more complicated. These two systems may act dependently, or the induction of one system may interfere with the other. Experimental studies have found that different viruses modulate these mechanisms to allow them to escape the host immune response or, worse, to exploit the host’s defense to their advantage; thus, this topic is a critical area in antiviral research. In this review, we summarize the current knowledge about how RNA viruses, including influenza virus, poliovirus, coxsackievirus, enterovirus 71, Japanese encephalitis virus, hepatitis C virus, and dengue virus, regulate these processes. We also discuss recent discoveries and how these will produce novel strategies for antiviral treatment.

  5. Ubiquitin vinyl methyl ester binding orients the misaligned active site of the ubiquitin hydrolase UCHL1 into productive conformation

    Energy Technology Data Exchange (ETDEWEB)

    Boudreaux, David A.; Maiti, Tushar K.; Davies, Christopher W.; Das, Chittaranjan (Purdue)

    2010-07-06

    Ubiquitin carboxy-terminal hydrolase L1 (UCHL1) is a Parkinson disease-associated, putative cysteine protease found abundantly and selectively expressed in neurons. The crystal structure of apo UCHL1 showed that the active-site residues are not aligned in a canonical form, with the nucleophilic cysteine being 7.7 {angstrom} from the general base histidine, an arrangement consistent with an inactive form of the enzyme. Here we report the crystal structures of the wild type and two Parkinson disease-associated variants of the enzyme, S18Y and I93M, bound to a ubiquitin-based suicide substrate, ubiquitin vinyl methyl ester. These structures reveal that ubiquitin vinyl methyl ester binds primarily at two sites on the enzyme, with its carboxy terminus at the active site and with its amino-terminal {beta}-hairpin at the distal site - a surface-exposed hydrophobic crevice 17 {angstrom} away from the active site. Binding at the distal site initiates a cascade of side-chain movements in the enzyme that starts at a highly conserved, surface-exposed phenylalanine and is relayed to the active site resulting in the reorientation and proximal placement of the general base within 4 {angstrom} of the catalytic cysteine, an arrangement found in productive cysteine proteases. Mutation of the distal-site, surface-exposed phenylalanine to alanine reduces ubiquitin binding and severely impairs the catalytic activity of the enzyme. These results suggest that the activity of UCHL1 may be regulated by its own substrate.

  6. Ubiquitination independent of E1 and E2 enzymes by bacterial effectors.

    Science.gov (United States)

    Qiu, Jiazhang; Sheedlo, Michael J; Yu, Kaiwen; Tan, Yunhao; Nakayasu, Ernesto S; Das, Chittaranjan; Liu, Xiaoyun; Luo, Zhao-Qing

    2016-05-01

    Signalling by ubiquitination regulates virtually every cellular process in eukaryotes. Covalent attachment of ubiquitin to a substrate is catalysed by the E1, E2 and E3 three-enzyme cascade, which links the carboxy terminus of ubiquitin to the ε-amino group of, in most cases, a lysine of the substrate via an isopeptide bond. Given the essential roles of ubiquitination in the regulation of the immune system, it is not surprising that the ubiquitination network is a common target for diverse infectious agents. For example, many bacterial pathogens exploit ubiquitin signalling using virulence factors that function as E3 ligases, deubiquitinases or as enzymes that directly attack ubiquitin. The bacterial pathogen Legionella pneumophila utilizes approximately 300 effectors that modulate diverse host processes to create a permissive niche for its replication in phagocytes. Here we demonstrate that members of the SidE effector family of L. pneumophila ubiquitinate multiple Rab small GTPases associated with the endoplasmic reticulum. Moreover, we show that these proteins are capable of catalysing ubiquitination without the need for the E1 and E2 enzymes. A putative mono-ADP-ribosyltransferase motif critical for the ubiquitination activity is also essential for the role of the SidE family in intracellular bacterial replication in a protozoan host. The E1/E2-independent ubiquitination catalysed by these enzymes is energized by nicotinamide adenine dinucleotide, which activates ubiquitin by the formation of ADP-ribosylated ubiquitin. These results establish that ubiquitination can be catalysed by a single enzyme, the activity of which does not require ATP.

  7. mir-30d Regulates multiple genes in the autophagy pathway and impairs autophagy process in human cancer cells

    Energy Technology Data Exchange (ETDEWEB)

    Yang, Xiaojun [Ovarian Cancer Research Center and Department of Obstetrics and Gynecology, University of Pennsylvania, Philadelphia, PA 19104 (United States); Department of General Surgery, Gansu Provincial Hospital, Lanzhou, Gansu 710000 (China); Zhong, Xiaomin [Ovarian Cancer Research Center and Department of Obstetrics and Gynecology, University of Pennsylvania, Philadelphia, PA 19104 (United States); Shanghai Key Laboratory of Female Reproductive Endocrine Related Diseases, Obstetrics and Gynecology Hospital, Fudan University, Shanghai 200011 (China); Tanyi, Janos L.; Shen, Jianfeng [Ovarian Cancer Research Center and Department of Obstetrics and Gynecology, University of Pennsylvania, Philadelphia, PA 19104 (United States); Xu, Congjian [Shanghai Key Laboratory of Female Reproductive Endocrine Related Diseases, Obstetrics and Gynecology Hospital, Fudan University, Shanghai 200011 (China); Gao, Peng [Department of General Surgery, Gansu Provincial Hospital, Lanzhou, Gansu 710000 (China); Zheng, Tim M. [Department of Pathology and Laboratory Medicine, University of Pennsylvania, Philadelphia, PA 19104 (United States); DeMichele, Angela [Division of Hematology and Oncology, Department of Medicine, University of Pennsylvania, Philadelphia, PA 19104 (United States); Zhang, Lin, E-mail: linzhang@mail.med.upenn.edu [Ovarian Cancer Research Center and Department of Obstetrics and Gynecology, University of Pennsylvania, Philadelphia, PA 19104 (United States)

    2013-02-15

    Highlights: ► Gene set enrichment analysis indicated mir-30d might regulate the autophagy pathway. ► mir-30d represses the expression of BECN1, BNIP3L, ATG12, ATG5 and ATG2. ► BECN1, BNIP3L, ATG12, ATG5 and ATG2 are direct targets of mir-30d. ► mir-30d inhibits autophagosome formation and LC3B-I conversion to LC3B-II. ► mir-30d regulates the autophagy process. -- Abstract: In human epithelial cancers, the microRNA (miRNA) mir-30d is amplified with high frequency and serves as a critical oncomir by regulating metastasis, apoptosis, proliferation, and differentiation. Autophagy, a degradation pathway for long-lived protein and organelles, regulates the survival and death of many cell types. Increasing evidence suggests that autophagy plays an important function in epithelial tumor initiation and progression. Using a combined bioinformatics approach, gene set enrichment analysis, and miRNA target prediction, we found that mir-30d might regulate multiple genes in the autophagy pathway including BECN1, BNIP3L, ATG12, ATG5, and ATG2. Our further functional experiments demonstrated that the expression of these core proteins in the autophagy pathway was directly suppressed by mir-30d in cancer cells. Finally, we showed that mir-30d regulated the autophagy process by inhibiting autophagosome formation and LC3B-I conversion to LC3B-II. Taken together, our results provide evidence that the oncomir mir-30d impairs the autophagy process by targeting multiple genes in the autophagy pathway. This result will contribute to understanding the molecular mechanism of mir-30d in tumorigenesis and developing novel cancer therapy strategy.

  8. Sinonasal inflammation in COPD

    DEFF Research Database (Denmark)

    Håkansson, Kåre; Konge, Lars; Thomsen, Sf;

    2013-01-01

    In this review we demonstrate that patients with chronic obstructive pulmonary disease (COPD) frequently report sinonasal symptoms. Furthermore, we present evidence that smoking on its own can cause nasal disease, and that in COPD patients, nasal inflammation mimics that of the bronchi. All...... this evidence suggests that COPD related sinonasal disease does exist and that smoking on its own rather than systemic inflammation triggers the condition. However, COPD related sinonasal disease remains to be characterized in terms of symptoms and endoscopic findings. In addition, more studies are needed...

  9. Interactions between Autophagy and Bacterial Toxins: Targets for Therapy?

    Science.gov (United States)

    Mathieu, Jacques

    2015-08-04

    Autophagy is a physiological process involved in defense mechanisms for clearing intracellular bacteria. The autophagic pathway is finely regulated and bacterial toxins interact with this process in a complex manner. Bacterial toxins also interact significantly with many biochemical processes. Evaluations of the effects of bacterial toxins, such as endotoxins, pore-forming toxins and adenylate cyclases, on autophagy could support the development of new strategies for counteracting bacterial pathogenicity. Treatment strategies could focus on drugs that enhance autophagic processes to improve the clearance of intracellular bacteria. However, further in vivo studies are required to decipher the upregulation of autophagy and potential side effects limiting such approaches. The capacity of autophagy activation strategies to improve the outcome of antibiotic treatment should be investigated in the future.

  10. Role of autophagy in liver physiology and pathophysiology

    Institute of Scientific and Technical Information of China (English)

    2010-01-01

    Autophagy is a highly conserved intracellular degradation pathway by which bulk cytoplasm and superfluous or damaged organelles are enveloped by double membrane structures termed autophagosomes. The autophago-somes then fuse with lysosomes for degradation of their contents, and the resulting amino acids can then recycle back to the cytosol. Autophagy is normally activated in response to nutrient deprivation and other stressors and occurs in all eukaryotes. In addition to maintaining energy and nutrient balance in the liver, it is now clear that autophagy plays a role in liver protein aggregates related diseases, hepatocyte cell death, steatohepatitis, hepatitis virus infection and hepatocellular carcinoma. In this review, I discuss the recent findings of autophagy with a focus on its role in liver pathophysiology.

  11. A role for TOR complex 2 signaling in promoting autophagy.

    Science.gov (United States)

    Vlahakis, Ariadne; Powers, Ted

    2014-01-01

    The conserved target of rapamycin (TOR) kinase is a central regulator of cell growth in response to nutrient availability. TOR forms 2 structurally and functionally distinct complexes, TORC1 and TORC2, and negatively regulates autophagy via TORC1. Here we demonstrate TOR also operates independently through the TORC2 signaling pathway to promote autophagy upon amino acid limitation. Under these conditions, TORC2, through its downstream target kinase Ypk1, inhibits the Ca(2+)- and Cmd1/calmodulin-dependent phosphatase, calcineurin, to enable the activation of the amino acid-sensing EIF2S1/eIF2α kinase, Gcn2, and promote autophagy. Thus TORC2 signaling regulates autophagy in a pathway distinct from TORC1 to provide a tunable response to the cellular metabolic state.

  12. Autophagy in Skeletal Muscle Homeostasis and in Muscular Dystrophies

    Directory of Open Access Journals (Sweden)

    Paolo Bonaldo

    2012-07-01

    Full Text Available Skeletal muscles are the agent of motion and one of the most important tissues responsible for the control of metabolism. The maintenance of muscle homeostasis is finely regulated by the balance between catabolic and anabolic process. Macroautophagy (or autophagy is a catabolic process that provides the degradation of protein aggregation and damaged organelles through the fusion between autophagosomes and lysosomes. Proper regulation of the autophagy flux is fundamental for the homeostasis of skeletal muscles during physiological situations and in response to stress. Defective as well as excessive autophagy is harmful for muscle health and has a pathogenic role in several forms of muscle diseases. This review will focus on the role of autophagy in muscle homeostasis and diseases.

  13. Glucocorticoids induce autophagy in rat bone marrow mesenchymal stem cells

    DEFF Research Database (Denmark)

    Wang, L.; Fan, J.; Lin, Y. S.;

    2015-01-01

    Glucocorticoidinduced osteoporosis (GIOP) is a widespread clinical complication following glucocorticoid therapy. This irreversible damage to boneforming and resorbing cells is essential in the pathogenesis of osteoporosis. Autophagy is a physiological process involved in the regulation of cells...... and their responses to diverse stimuli, however, the role of autophagy in glucocorticoidinduced damage to bone marrow mesenchymal stem cells (BMSCs) remains unclear. The current study confirmed that glucocorticoid administration impaired the proliferation of BMSCs. Transmission electron microscopy......, immunohistochemistry and western blot analysis detected autophagy in vitro and in GIOP model rats (in vivo). With the addition of the autophagy inhibitor 3methyladenine, the proliferative ability of BMSCs was further reduced, while the number of apoptotic BMSCs was significantly increased. The data suggests...

  14. microRNA-101 is a potent inhibitor of autophagy

    DEFF Research Database (Denmark)

    Frankel, Lisa B; Wen, Jiayu; Lees, Michael;

    2011-01-01

    Autophagy is an evolutionarily conserved mechanism of cellular self-digestion in which proteins and organelles are degraded through delivery to lysosomes. Defects in this process are implicated in numerous human diseases including cancer. To further elucidate regulatory mechanisms of autophagy, we...... performed a functional screen in search of microRNAs (miRNAs), which regulate the autophagic flux in breast cancer cells. In this study, we identified the tumour suppressive miRNA, miR-101, as a potent inhibitor of basal, etoposide- and rapamycin-induced autophagy. Through transcriptome profiling, we...... identified three novel miR-101 targets, STMN1, RAB5A and ATG4D. siRNA-mediated depletion of these genes phenocopied the effect of miR-101 overexpression, demonstrating their importance in autophagy regulation. Importantly, overexpression of STMN1 could partially rescue cells from miR-101-mediated inhibition...

  15. Heat shock response and autophagy--cooperation and control.

    Science.gov (United States)

    Dokladny, Karol; Myers, Orrin B; Moseley, Pope L

    2015-01-01

    Protein quality control (proteostasis) depends on constant protein degradation and resynthesis, and is essential for proper homeostasis in systems from single cells to whole organisms. Cells possess several mechanisms and processes to maintain proteostasis. At one end of the spectrum, the heat shock proteins modulate protein folding and repair. At the other end, the proteasome and autophagy as well as other lysosome-dependent systems, function in the degradation of dysfunctional proteins. In this review, we examine how these systems interact to maintain proteostasis. Both the direct cellular data on heat shock control over autophagy and the time course of exercise-associated changes in humans support the model that heat shock response and autophagy are tightly linked. Studying the links between exercise stress and molecular control of proteostasis provides evidence that the heat shock response and autophagy coordinate and undergo sequential activation and downregulation, and that this is essential for proper proteostasis in eukaryotic systems.

  16. Autophagy-modulating aminosteroids isolated from the sponge Cliona celata

    NARCIS (Netherlands)

    R.A. Keyzers; J. Daoust; M.T. Davies-Coleman; R. van Soest; A. Balgi; E. Donohue; M. Roberge; R.J. Andersen

    2008-01-01

    Clionamines A−D (1−4), new aminosteroids that modulate autophagy, have been isolated from South African specimens of the sponge Cliona celata. Clionamine D (4) has an unprecedented spiro bislactone side chain.

  17. Interactions between Autophagy and Bacterial Toxins: Targets for Therapy?

    Directory of Open Access Journals (Sweden)

    Jacques Mathieu

    2015-08-01

    Full Text Available Autophagy is a physiological process involved in defense mechanisms for clearing intracellular bacteria. The autophagic pathway is finely regulated and bacterial toxins interact with this process in a complex manner. Bacterial toxins also interact significantly with many biochemical processes. Evaluations of the effects of bacterial toxins, such as endotoxins, pore-forming toxins and adenylate cyclases, on autophagy could support the development of new strategies for counteracting bacterial pathogenicity. Treatment strategies could focus on drugs that enhance autophagic processes to improve the clearance of intracellular bacteria. However, further in vivo studies are required to decipher the upregulation of autophagy and potential side effects limiting such approaches. The capacity of autophagy activation strategies to improve the outcome of antibiotic treatment should be investigated in the future.

  18. Autophagy as a Therapeutic Target in Cardiovascular Disease

    Science.gov (United States)

    Nemchenko, Andriy; Chiong, Mario; Turer, Aslan; Lavandero, Sergio; Hill, Joseph A.

    2011-01-01

    The epidemic of heart failure continues apace, and development of novel therapies with clinical efficacy has lagged. Now, important insights into the molecular circuitry of cardiovascular autophagy have raised the prospect that this cellular pathway of protein quality control may be a target of clinical relevance. Whereas basal levels of autophagy are required for cell survival, excessive levels – or perhaps distinct forms of autophagic flux – contribute to disease pathogenesis. Our challenge will be to distinguish mechanisms that drive adaptive versus maladaptive autophagy and to manipulate those pathways for therapeutic gain. Recent evidence suggests this may be possible. Here, we review the fundamental biology of autophagy and its role in a variety of forms of cardiovascular disease. We discuss ways in which this evolutionarily conserved catabolic mechanism can be manipulated, discuss studies presently underway in heart disease, and provide our perspective on where this exciting field may lead in the future. PMID:21723289

  19. Methods for the Detection of Autophagy in Mammalian Cells.

    Science.gov (United States)

    Zhang, Ziyan; Singh, Rajat; Aschner, Michael

    2016-01-01

    Macroautophagy (hereafter referred to as autophagy) is a degradation pathway that delivers cytoplasmic materials to lysosomes via double-membraned vesicles designated autophagosomes. Cytoplasmic constituents are sequestered into autophagosomes, which subsequently fuse with lysosomes, where the cargo is degraded. Autophagy is a crucial mechanism involved in many aspects of cell function, including cellular metabolism and energy balance; alterations in autophagy have been linked to various human pathological processes. Thus, methods that accurately measure autophagic activity are necessary. In this unit, we introduce several approaches to analyze autophagy in mammalian cells, including immunoblotting analysis of LC3 and p62, detection of autophagosome formation by fluorescence microscopy, and monitoring autophagosome maturation by tandem mRFP-GFP fluorescence microscopy. Overall, we recommend a combined use of multiple methods to accurately assess the autophagic activity in any given biological setting. © 2016 by John Wiley & Sons, Inc. PMID:27479363

  20. Detection of Autophagy in Caenorhabditis elegans Using GFP::LGG-1 as an Autophagy Marker.

    Science.gov (United States)

    Palmisano, Nicholas J; Meléndez, Alicia

    2016-01-04

    In yeast and mammalian cells, the autophagy protein Atg8/LC3 (microtubule-associated proteins 1A/1B light chain 3B encoded by MAP1LC3B) has been the marker of choice to detect double-membraned autophagosomes that are produced during the process of autophagy. A lipid-conjugated form of Atg8/LC3B is localized to the inner and outer membrane of the early-forming structure known as the phagophore. During maturation of autophagosomes, Atg8/LC3 bound to the inner autophagosome membrane remains in situ as the autophagosomes fuse with lysosomes. The nematode Caenorhabditis elegans is thought to conduct a similar process, meaning that tagging the nematode ortholog of Atg8/LC3-known as LGG-1-with a fluorophore has become a widely accepted method to visualize autophagosomes. Under normal growth conditions, GFP-modified LGG-1 displays a diffuse expression pattern throughout a variety of tissues, whereas, when under conditions that induce autophagy, the GFP::LGG-1 tag labels positive punctate structures, and its overall level of expression increases. Here, we present a protocol for using fluorescent reporters of LGG-1 coupled to GFP to monitor autophagosomes in vivo. We also discuss the use of alternative fluorescent markers and the possible utility of the LGG-1 paralog LGG-2.

  1. Periostin in Allergic Inflammation

    Directory of Open Access Journals (Sweden)

    Kenji Izuhara

    2014-01-01

    Full Text Available Periostin, an extracellular matrix protein belonging to the fasciclin family, has been shown to play a critical role in the process of remodeling during tissue/organ development or repair. Periostin functions as a matricellular protein in cell activation by binding to their receptors on cell surface, thereby exerting its biological activities. After we found that periostin is a downstream molecule of interleukin (IL-4 and IL-13, signature cytokines of type 2 immune responses, we showed that periostin is a component of subepithelial fibrosis in bronchial asthma, the first formal proof that periostin is involved in allergic inflammation. Subsequently, a great deal of evidence has accumulated demonstrating the significance of periostin in allergic inflammation. It is of note that in skin tissues, periostin is critical for amplification and persistence of allergic inflammation by communicating between fibroblasts and keratinocytes. Furthermore, periostin has been applied to development of novel diagnostics or therapeutic agents for allergic diseases. Serum periostin can reflect local production of periostin in inflamed lesions induced by Th2-type immune responses and also can predict the efficacy of Th2 antagonists against bronchial asthma. Blocking the interaction between periostin and its receptor, αv integrin, or down-regulating the periostin expression shows improvement of periostin-induced inflammation in mouse models or in in vitro systems. It is hoped that diagnostics or therapeutic agents targeting periostin will be of practical use in the near future.

  2. Immunsystemet ved kronisk inflammation

    DEFF Research Database (Denmark)

    Bendtzen, Klaus

    2008-01-01

    Innate and adaptive immunity has evolved as a defence against infections and as an important repair mechanism after physical injury. If elimination of microbes and healing is not achieved, or if the immune system is dysregulated, chronic inflammation ensues. Immune cells become engaged in prolonged...

  3. The amino acid sensor GCN2 controls gut inflammation by inhibiting inflammasome activation.

    Science.gov (United States)

    Ravindran, Rajesh; Loebbermann, Jens; Nakaya, Helder I; Khan, Nooruddin; Ma, Hualing; Gama, Leonardo; Machiah, Deepa K; Lawson, Benton; Hakimpour, Paul; Wang, Yi-chong; Li, Shuzhao; Sharma, Prachi; Kaufman, Randal J; Martinez, Jennifer; Pulendran, Bali

    2016-03-24

    The integrated stress response (ISR) is a homeostatic mechanism by which eukaryotic cells sense and respond to stress-inducing signals, such as amino acid starvation. General controlled non-repressed (GCN2) kinase is a key orchestrator of the ISR, and modulates protein synthesis in response to amino acid starvation. Here we demonstrate in mice that GCN2 controls intestinal inflammation by suppressing inflammasome activation. Enhanced activation of ISR was observed in intestinal antigen presenting cells (APCs) and epithelial cells during amino acid starvation, or intestinal inflammation. Genetic deletion of Gcn2 (also known as Eif2ka4) in CD11c(+) APCs or intestinal epithelial cells resulted in enhanced intestinal inflammation and T helper 17 cell (TH17) responses, owing to enhanced inflammasome activation and interleukin (IL)-1β production. This was caused by reduced autophagy in Gcn2(-/-) intestinal APCs and epithelial cells, leading to increased reactive oxygen species (ROS), a potent activator of inflammasomes. Thus, conditional ablation of Atg5 or Atg7 in intestinal APCs resulted in enhanced ROS and TH17 responses. Furthermore, in vivo blockade of ROS and IL-1β resulted in inhibition of TH17 responses and reduced inflammation in Gcn2(-/-) mice. Importantly, acute amino acid starvation suppressed intestinal inflammation via a mechanism dependent on GCN2. These results reveal a mechanism that couples amino acid sensing with control of intestinal inflammation via GCN2.

  4. Are mitochondrial reactive oxygen species required for autophagy?

    Energy Technology Data Exchange (ETDEWEB)

    Jiang, Jianfei, E-mail: jjf73@pitt.edu [Center for Free Radical and Antioxidant Health, Department of Environmental and Occupational Health, University of Pittsburgh (United States); Maeda, Akihiro; Ji, Jing [Center for Free Radical and Antioxidant Health, Department of Environmental and Occupational Health, University of Pittsburgh (United States); Baty, Catherine J.; Watkins, Simon C. [Center for Biologic Imaging, Department of Cell Biology and Physiology, University of Pittsburgh (United States); Greenberger, Joel S. [Department of Radiation Oncology, University of Pittsburgh (United States); Kagan, Valerian E., E-mail: kagan@pitt.edu [Center for Free Radical and Antioxidant Health, Department of Environmental and Occupational Health, University of Pittsburgh (United States)

    2011-08-19

    Highlights: {yields} Autophageal and apoptotic pathways were dissected in cytochrome c deficient cells. {yields} Staurosporine (STS)-induced autophagy was not accompanied by ROS generation. {yields} Autophagy was detectable in mitochondrial DNA deficient {rho}{sup 0} cells. {yields} Mitochondrial ROS are not required for the STS-induced autophagy in HeLa cells. -- Abstract: Reactive oxygen species (ROS) are said to participate in the autophagy signaling. Supporting evidence is obscured by interference of autophagy and apoptosis, whereby the latter heavily relies on ROS signaling. To dissect autophagy from apoptosis we knocked down expression of cytochrome c, the key component of mitochondria-dependent apoptosis, in HeLa cells using shRNA. In cytochrome c deficient HeLa1.2 cells, electron transport was compromised due to the lack of electron shuttle between mitochondrial respiratory complexes III and IV. A rapid and robust LC3-I/II conversion and mitochondria degradation were observed in HeLa1.2 cells treated with staurosporine (STS). Neither generation of superoxide nor accumulation of H{sub 2}O{sub 2} was detected in STS-treated HeLa1.2 cells. A membrane permeable antioxidant, PEG-SOD, plus catalase exerted no effect on STS-induced LC3-I/II conversion and mitochondria degradation. Further, STS caused autophagy in mitochondria DNA-deficient {rho}{sup o} HeLa1.2 cells in which both electron transport and ROS generation were completely disrupted. Counter to the widespread view, we conclude that mitochondrial ROS are not required for the induction of autophagy.

  5. Linking ER Stress to Autophagy: Potential Implications for Cancer Therapy

    OpenAIRE

    Tom Verfaillie; Maria Salazar; Guillermo Velasco; Patrizia Agostinis

    2010-01-01

    Different physiological and pathological conditions can perturb protein folding in the endoplasmic reticulum, leading to a condition known as ER stress. ER stress activates a complex intracellular signal transduction pathway, called unfolded protein response (UPR). The UPR is tailored essentially to reestablish ER homeostasis also through adaptive mechanisms involving the stimulation of autophagy. However, when persistent, ER stress can switch the cytoprotective functions of UPR and autophagy...

  6. Autophagy in Skeletal Muscle Homeostasis and in Muscular Dystrophies

    OpenAIRE

    Paolo Bonaldo; Paolo Grumati

    2012-01-01

    Skeletal muscles are the agent of motion and one of the most important tissues responsible for the control of metabolism. The maintenance of muscle homeostasis is finely regulated by the balance between catabolic and anabolic process. Macroautophagy (or autophagy) is a catabolic process that provides the degradation of protein aggregation and damaged organelles through the fusion between autophagosomes and lysosomes. Proper regulation of the autophagy flux is fundamental for the homeostasis o...

  7. Autophagy Inhibition to Increase Radiosensitization in Breast Cancer

    OpenAIRE

    Liang, Diana Hwang; El-Zein, Randa; Dave, Bhuvanesh

    2015-01-01

    Currently, many breast cancer patients with localized breast cancer undergo breast-conserving therapy, consisting of local excision followed by radiation therapy. Following radiation therapy, breast cancer cells are noted to undergo induction of autophagy, development of radioresistance, and enrichment of breast cancer stem cell subpopulations. It is hypothesized that inhibition of the cytoprotective autophagy that arises following radiation therapy increases radiosensitivity and confers long...

  8. Obesity, autophagy and the pathogenesis of liver and pancreatic cancers

    OpenAIRE

    Aghajan, Mariam; Ning LI; Karin, Michael

    2012-01-01

    Liver and pancreatic cancers are both highly lethal diseases with limited to no therapeutic options for patients. Recent studies suggest that deregulated autophagy plays a role in the pathogenesis of these diseases by perturbing cellular homeostasis and laying the foundation for disease development. While accumulation of p62 upon impaired autophagy has been implicated in hepatocellular carcinoma, it’s role in pancreatic adenocarcinoma remains less clear. This review will focus on recent studi...

  9. Autophagy as a new therapeutic target in Duchenne muscular dystrophy

    OpenAIRE

    Palma, C.; F. Morisi; Cheli, S; S. Pambianco; Cappello, V; Vezzoli, M; Rovere-Querini, P; Moggio, M; Ripolone, M.; Francolini, M; Sandri, M.; Clementi, E

    2012-01-01

    A resolutive therapy for Duchene muscular dystrophy, a severe degenerative disease of the skeletal muscle, is still lacking. Because autophagy has been shown to be crucial in clearing dysfunctional organelles and in preventing tissue damage, we investigated its pathogenic role and its suitability as a target for new therapeutic interventions in Duchenne muscular dystrophy (DMD). Here we demonstrate that autophagy is severely impaired in muscles from patients affected by DMD and mdx mice, a mo...

  10. Autophagy in herpesvirus immune control and immune escape

    OpenAIRE

    Taylor, G. S.; Mautner, J.; Münz, C

    2011-01-01

    Autophagy delivers cytoplasmic constituents for lysosomal degradation, and thereby facilitates pathogen degradation and pathogen fragment loading onto MHC molecules for antigen presentation to T cells. Herpesviruses have been used to demonstrate these novel functions of autophagy, which previously has been primarily appreciated for its pro-survival role during starvation. In this review, we summarize recent findings how macroautophagy restricts herpesvirus infections directly, how macroautoph...

  11. Mastermind-Like 1 Is Ubiquitinated: Functional Consequences for Notch Signaling.

    Directory of Open Access Journals (Sweden)

    Mozhgan Farshbaf

    Full Text Available Early studies demonstrated the involvement of ubiquitination of the Notch intracellular domain for rapid turnover of the transcriptional complex at Notch target genes. It was shown that this ubiquitination was promoted by the co-activator Mastermind like 1 (MAML1. MAML1 also contains numerous lysine residues that may also be ubiquitinated and necessary for protein regulation. In this study, we show that over-expressed MAML1 is ubiquitinated and identify eight conserved lysine residues which are required for ubiquitination. We also show that p300 stimulates ubiquitination and that Notch inhibits ubiquitination. Furthermore, we show that a mutant MAML1 that has decreased ubiquitination shows increased output from a HES1 reporter gene assay. Therefore, we speculate that ubiquitination of MAML1 might be a mechanism to maintain low levels of the protein until needed for transcriptional activation. In summary, this study identifies that MAML1 is ubiquitinated in the absence of Notch signaling to maintain low levels of MAML1 in the cell. Our data supports the notion that a precise and tight regulation of the Notch pathway is required for this signaling pathway.

  12. Regulation of B cell differentiation by the ubiquitin-binding protein TAX1BP1

    Science.gov (United States)

    Matsushita, Nobuko; Suzuki, Midori; Ikebe, Emi; Nagashima, Shun; Inatome, Ryoko; Asano, Kenichi; Tanaka, Masato; Matsushita, Masayuki; Kondo, Eisaku; Iha, Hidekatsu; Yanagi, Shigeru

    2016-01-01

    Tax1-binding protein 1 (TAX1BP1) is a ubiquitin-binding protein that restricts nuclear factor-κB (NF-κB) activation and facilitates the termination of aberrant inflammation. However, its roles in B-cell activation and differentiation are poorly understood. To evaluate the function of TAX1BP1 in B cells, we established TAX1BP1-deficient DT40 B cells that are hyper-responsive to CD40-induced extracellular signal-regulated kinase (ERK) activation signaling, exhibit prolonged and exaggerated ERK phosphorylation and show enhanced B lymphocyte-induced maturation protein 1 (Blimp-1; a transcription factor inducing plasma cell differentiation) expression that is ERK-dependent. Furthermore, TAX1BP1-deficient cells exhibit significantly decreased surface IgM expression and increased IgM secretion. Moreover, TAX1BP1-deficient mice display reduced germinal center formation and antigen-specific antibody production. These findings show that TAX1BP1 restricts ERK activation and Blimp-1 expression and regulates germinal center formation. PMID:27515252

  13. Ataxia, Dementia, and Hypogonadotropism Caused by Disordered Ubiquitination

    DEFF Research Database (Denmark)

    Margolin, David H.; Kousi, Maria; Chan, Yee-Ming;

    2013-01-01

    affected patients. Neurologic and reproductive endocrine phenotypes were characterized in detail. The effects of sequence variants and the presence of an epistatic interaction were tested in a zebrafish model. RESULTS Digenic homozygous mutations in RNF216 and OTUD4, which encode a ubiquitin E3 ligase...... in zebrafish embryos induced defects in the eye, optic tectum, and cerebellum; combinatorial suppression of both genes exacerbated these phenotypes, which were rescued by nonmutant, but not mutant, human RNF216 or OTUD4 messenger RNA. All patients had progressive ataxia and dementia. Neuronal loss was observed...... in cerebellar pathways and the hippocampus; surviving hippocampal neurons contained ubiquitin-immunoreactive intranuclear inclusions. Defects were detected at the hypothalamic and pituitary levels of the reproductive endocrine axis. CONCLUSIONS The syndrome of hypogonadotropic hypogonadism, ataxia, and dementia...

  14. Role of the Ubiquitin Proteasome System in Regulating Skin Pigmentation

    Directory of Open Access Journals (Sweden)

    Hideya Ando

    2009-10-01

    Full Text Available Pigmentation of the skin, hair and eyes is regulated by tyrosinase, the critical rate-limiting enzyme in melanin synthesis by melanocytes. Tyrosinase is degraded endogenously, at least in part, by the ubiquitin proteasome system (UPS. Several types of inherited hypopigmentary diseases, such as oculocutaneous albinism and Hermansky-Pudlak syndrome, involve the aberrant processing and/or trafficking of tyrosinase and its subsequent degradation which can occur due to the quality-control machinery. Studies on carbohydrate modifications have revealed that tyrosinase in the endoplasmic reticulum (ER is proteolyzed via ER-associated protein degradation and that tyrosinase degradation can also occur following its complete maturation in the Golgi. Among intrinsic factors that regulate the UPS, fatty acids have been shown to modulate tyrosinase degradation in contrasting manners through increased or decreased amounts of ubiquitinated tyrosinase that leads to its accelerated or decelerated degradation by proteasomes.

  15. Cell fate determination by ubiquitin-dependent regulation of translation

    Science.gov (United States)

    Werner, Achim; Iwasaki, Shintaro; McGourty, Colleen; Medina-Ruiz, Sofia; Teerikorpi, Nia; Fedrigo, Indro; Ingolia, Nicholas T.; Rape, Michael

    2015-01-01

    Metazoan development depends on accurate execution of differentiation programs that allow pluripotent stem cells to adopt specific fates 1. Differentiation requires changes to chromatin architecture and transcriptional networks, yet whether other regulatory events support cell fate determination is less well understood. Here, we have identified the vertebrate-specific ubiquitin ligase CUL3KBTBD8 as an essential regulator of neural crest specification. CUL3KBTBD8 monoubiquitylates NOLC1 and its paralog TCOF1, whose mutation underlies the neurocristopathy Treacher Collins Syndrome 2,3. Ubiquitylation drives formation of a TCOF1-NOLC1 platform that connects RNA polymerase I with ribosome modification enzymes and remodels the translational program of differentiating cells in favor of neural crest specification. We conclude that ubiquitin-dependent regulation of translation is an important feature of cell fate determination. PMID:26399832

  16. Direct observation of silver nanoparticle-ubiquitin corona formation

    CERN Document Server

    Ding, Feng; Choudhary, Poonam; Chen, Ran; Brown, Jared M; Ke, Pu Chun

    2012-01-01

    Upon entering physiological environments, nanoparticles readily assume the form of a nanoparticle-protein corona that dictates their biological identity. Understanding the structure and dynamics of nanoparticle-protein corona is essential for predicting the fate, transport, and toxicity of nanomaterials in living systems and for enabling the vast applications of nanomedicine. We combined multiscale molecular dynamics simulations and complementary experiments to characterize the silver nanoparticle-ubiquitin corona formation. Specifically, ubiquitins competed with citrates for the nanoparticle surface and bound to the particle in a specific manner. Under a high protein/nanoparticle stoichiometry, ubiquitions formed a multi-layer corona on the particle surface. The binding exhibited an unusual stretched-exponential behavior, suggesting a rich kinetics originated from protein-protein, protein-citrate, and protein-nanoparticle interactions. Furthermore, the binding destabilized the {\\alpha}-helices while increasi...

  17. Ubiquitin-dependent trafficking and turnover of ionotropic glutamate receptors

    Directory of Open Access Journals (Sweden)

    Marisa S Goo

    2015-10-01

    Full Text Available Changes in synaptic strength underlie the basis of learning and memory and are controlled, in part, by the insertion or removal of AMPA-type glutamate receptors at the postsynaptic membrane of excitatory synapses. Once internalized, these receptors may be recycled back to the plasma membrane by subunit-specific interactions with other proteins or by post-translational modifications such as phosphorylation. Alternatively, these receptors may be targeted for destruction by multiple degradation pathways in the cell. Ubiquitination, another post-translational modification, has recently emerged as a key signal that regulates the recycling and trafficking of glutamate receptors. In this review, we will discuss recent findings on the role of ubiquitination in the trafficking and turnover of ionotropic glutamate receptors and plasticity of excitatory synapses.

  18. 细胞自噬及其在鼻咽癌中的作用%Autophagy and its role in nasopharyngeal carcinoma

    Institute of Scientific and Technical Information of China (English)

    王佳(综述); 周文胜(审校)

    2015-01-01

    Autophagy, an important homeostatic cellular recycling mechanism, is now emerging as a crucial player in response to metabolic and therapeutic stresses, which attempts to maintain/restore metabolic homeostasis through the catabolic lysis of excessive or unnecessary proteins and injured or aged organelles.Autophagy is particularly active during metabolic stress. In the cancer cell it fulfils a dual role, having tumor-promoting and tumor-suppressing properties. Functional autophagy prevents necrosis and inflammation, which can lead to genetic instability. On the other hand, autophagy might be important for tumor progression by providing energy through its recycling mechanism during unfavorable metabolic circumstances. Recently, several studies have shown that autophagy play an important role in treatment of nasopharyngeal carcinoma. hTis review will summarize the signaltransductionpathway of autophagy, its relationship with tumor and its role in the nasopharyngeal carcinoma treatment in order to give a comprehensive understanding of the function of autophagy.%自噬作为一个重要的自我平衡的细胞回收机制,现逐渐发现在新陈代谢和治疗应激中起了关键作用。它通过消除细胞内过多不必要的蛋白质及受损或衰老的细胞器成分来维持/恢复新陈代谢的稳定。在代谢应激的情况下,自噬被激活。在癌细胞中自噬有两个方面的作用:肿瘤促进作用和肿瘤抑制作用。有功能的自噬可防止会导致遗传不稳定性发生的细胞坏死和炎症反应。另一方面,自噬通过不利代谢环境的循环机制提供能量对肿瘤进程起了重要作用。最近,许多研究表明,自噬在鼻咽癌的治疗中起了重要作用。本文将对自噬所依赖的信号转导途经、自噬与肿瘤的关系、自噬在鼻咽癌治疗中的作用进行综述,以便对自噬功能有个全面的理解。

  19. Ubiquitination of inducible nitric oxide synthase is required for its degradation

    Science.gov (United States)

    Kolodziejski, Pawel J.; Musial, Aleksandra; Koo, Ja-Seok; Eissa, N. Tony

    2002-01-01

    Inducible nitric oxide synthase (iNOS) is responsible for nitric oxide (NO) synthesis from l-arginine in response to inflammatory mediators. We have previously shown that iNOS is degraded through the 26S proteasome. Targeting of proteins for proteasomal degradation may or may not require their covalent linkage to multiubiquitin chains (ubiquitination). In addition, ubiquitination of a protein can serve functions other than signaling proteolysis. In this context, it is not known whether iNOS is subject to ubiquitination or whether ubiquitination is required for its degradation. In this study, we show that iNOS, expressed in HEK293 cells or induced in primary bronchial epithelial cells, A549 cells, or murine macrophages, is subject to ubiquitination. To investigate whether iNOS ubiquitination is required for its degradation, HEK293T cells were cotransfected with plasmids containing cDNAs of human iNOS and of the dominant negative ubiquitin mutant K48R. Disruption of ubiquitination by K48R ubiquitin resulted in inhibition of iNOS degradation. ts20 is a mutant cell line that contains a thermolabile ubiquitin-activating enzyme (E1) that is inactivated at elevated temperature, preventing ubiquitination. Incubation of ts20 cells, stably expressing human iNOS, at the nonpermissive temperature (40°C) resulted in inhibition of iNOS degradation and marked accumulation of iNOS. These studies indicate that iNOS is subject to ubiquitination and that ubiquitination is required for its degradation. PMID:12221289

  20. Retinoid receptor signaling and autophagy in acute promyelocytic leukemia

    Energy Technology Data Exchange (ETDEWEB)

    Orfali, Nina [Cork Cancer Research Center, University College Cork, Cork (Ireland); Department of Pharmacology, Weill Cornell Medical College, New York, NY 10065, USA. (United States); McKenna, Sharon L. [Cork Cancer Research Center, University College Cork, Cork (Ireland); Cahill, Mary R. [Department of Hematology, Cork University Hospital, Cork (Ireland); Gudas, Lorraine J., E-mail: ljgudas@med.cornell.edu [Department of Pharmacology, Weill Cornell Medical College, New York, NY 10065, USA. (United States); Mongan, Nigel P., E-mail: nigel.mongan@nottingham.ac.uk [Faculty of Medicine and Health Science, School of Veterinary Medicine and Science, University of Nottingham, LE12 5RD (United Kingdom); Department of Pharmacology, Weill Cornell Medical College, New York, NY 10065, USA. (United States)

    2014-05-15

    Retinoids are a family of signaling molecules derived from vitamin A with well established roles in cellular differentiation. Physiologically active retinoids mediate transcriptional effects on cells through interactions with retinoic acid (RARs) and retinoid-X (RXR) receptors. Chromosomal translocations involving the RARα gene, which lead to impaired retinoid signaling, are implicated in acute promyelocytic leukemia (APL). All-trans-retinoic acid (ATRA), alone and in combination with arsenic trioxide (ATO), restores differentiation in APL cells and promotes degradation of the abnormal oncogenic fusion protein through several proteolytic mechanisms. RARα fusion-protein elimination is emerging as critical to obtaining sustained remission and long-term cure in APL. Autophagy is a degradative cellular pathway involved in protein turnover. Both ATRA and ATO also induce autophagy in APL cells. Enhancing autophagy may therefore be of therapeutic benefit in resistant APL and could broaden the application of differentiation therapy to other cancers. Here we discuss retinoid signaling in hematopoiesis, leukemogenesis, and APL treatment. We highlight autophagy as a potential important regulator in anti-leukemic strategies. - Highlights: • Normal and aberrant retinoid signaling in hematopoiesis and leukemia is reviewed. • We suggest a novel role for RARα in the development of X-RARα gene fusions in APL. • ATRA therapy in APL activates transcription and promotes onco-protein degradation. • Autophagy may be involved in both onco-protein degradation and differentiation. • Pharmacologic autophagy induction may potentiate ATRA's therapeutic effects.