RESUMEN
Tauopathies are neurodegenerative disorders characterized by the deposition of aggregates of the microtubule-associated protein tau, a main component of neurofibrillary tangles. Alzheimer's disease (AD) is the most common type of tauopathy and dementia, with amyloid-beta pathology as an additional hallmark feature of the disease. Besides its role in stabilizing microtubules, tau is localized at postsynaptic sites and can regulate synaptic plasticity. The activity-regulated cytoskeleton-associated protein (Arc) is an immediate early gene that plays a key role in synaptic plasticity, learning, and memory. Arc has been implicated in AD pathogenesis and regulates the release of amyloid-beta. We found that decreased Arc levels correlate with AD status and disease severity. Importantly, Arc protein was upregulated in the hippocampus of Tau KO mice and dendrites of Tau KO primary hippocampal neurons. Overexpression of tau decreased Arc stability in an activity-dependent manner, exclusively in neuronal dendrites, which was coupled to an increase in the expression of dendritic and somatic surface GluA1-containing α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptors. The tau-dependent decrease in Arc was found to be proteasome-sensitive, yet independent of Arc ubiquitination and required the endophilin-binding domain of Arc. Importantly, these effects on Arc stability and GluA1 localization were not observed in the commonly studied tau mutant, P301L. These observations provide a potential molecular basis for synaptic dysfunction mediated through the accumulation of tau in dendrites. Our findings confirm that Arc is misregulated in AD and further show a physiological role for tau in regulating Arc stability and AMPA receptor targeting.
Asunto(s)
Proteínas del Citoesqueleto , Dendritas , Proteínas del Tejido Nervioso , Complejo de la Endopetidasa Proteasomal , Proteínas tau , Animales , Humanos , Ratones , Enfermedad de Alzheimer/metabolismo , Enfermedad de Alzheimer/patología , Enfermedad de Alzheimer/genética , Proteínas del Citoesqueleto/metabolismo , Proteínas del Citoesqueleto/genética , Dendritas/metabolismo , Dendritas/patología , Hipocampo/metabolismo , Hipocampo/patología , Ratones Noqueados , Proteínas del Tejido Nervioso/metabolismo , Proteínas del Tejido Nervioso/genética , Neuronas/metabolismo , Neuronas/patología , Complejo de la Endopetidasa Proteasomal/metabolismo , Estabilidad Proteica , Proteínas tau/metabolismo , Proteínas tau/genética , Ubiquitina/metabolismo , UbiquitinaciónRESUMEN
The selectivity in selective macroautophagy/autophagy pathways is achieved via selective autophagy receptors (SARs) - proteins that bind a ligand on the substrate to be degraded and an Atg8-family protein on the growing autophagic membrane, phagophore, effectively bridging them. In mammals, the most common ligand of SARs is ubiquitin, a small protein modifier that tags substrates for their preferential degradation by autophagy. Consequently, most common SARs are ubiquitin-binding SARs, such as SQSTM1/p62 (sequestosome 1). Surprisingly, there is only one SAR of this type in yeast - Cue5, which acts as the receptor for aggrephagy and proteaphagy - pathways that remove ubiquitinated protein aggregates and proteasomes, respectively. However, recent studies described ubiquitin-dependent autophagic pathways that do not require Cue5, e.g. the stationary phase lipophagy for lipid droplets or nitrogen starvation-induced mitophagy for mitochondria. What is the role of ubiquitin in these pathways? Here, we propose that ubiquitinated lipid droplets and mitochondria are recognized by alternative ubiquitin-binding SARs. Our analysis identifies proteins that could potentially fulfill this role in yeast. We think that matching of ubiquitin-dependent (but Cue5-independent) autophagic pathways with ubiquitin- and Atg8-binding proteins enlisted here might uncover novel ubiquitin-binding SARs in yeast.Abbreviations: AIM: Atg8-family interacting motif; CUE: coupling of ubiquitin conjugation to ER degradation; ERMES: endoplasmic reticulum-mitochondria encounter structure; HECT: homologous to the E6-AP carboxyl terminus; LD: lipid droplet; SAR: selective autophagy receptor; SGD: Saccharomyces Genome Database; UBA: ubiquitin-associated; UBX: ubiquitin regulatory X; UIM: ubiquitin-interacting motif.
Asunto(s)
Saccharomyces cerevisiae , Ubiquitina , Animales , Proteínas Adaptadoras Transductoras de Señales/metabolismo , Autofagia , Familia de las Proteínas 8 Relacionadas con la Autofagia/metabolismo , Ligandos , Mamíferos/metabolismo , Saccharomyces cerevisiae/metabolismo , Ubiquitina/metabolismoRESUMEN
SQSTM1/p62 (sequestosome 1) is a well-established indicator of macroautophagic/autophagic flux. It was initially characterized as the ubiquitin-binding autophagic receptor in aggrephagy, the selective autophagy of ubiquitinated protein aggregates. Recently, several studies correlated its levels with the abundance of intracellular lipid droplets (LDs). In the absence of a bona fide receptor for the selective autophagy of LDs (lipophagy), a few studies demonstrated the role of SQSTM1 in lipophagy. Our analysis of these studies shows that SQSTM1 colocalizes with LDs, bridges them with phagophores, is co-degraded with them in the lysosomes, and affects LD abundance in a variety of cells and under diverse experimental conditions. Although only one study reported all these functions together, the overwhelming and complementary evidence from other studies suggests that the role of SQSTM1 in lipophagy via tagging, movement, aggregation/clustering and sequestration of LDs is rather a common phenomenon in mammalian cells. As ubiquitination of the LD-associated proteins under stress conditions is increasingly recognized as another common phenomenon, some other ubiquitin-binding autophagic receptors, such as NBR1 and OPTN, might soon join SQSTM1 on a list of the non-exclusive lipophagy receptors.Abbreviations: LD: lipid droplet; LIR: LC3-interacting region; PAT: Perilipin, ADRP and TIP47 domain; SAR: selective autophagy receptor.
Asunto(s)
Autofagia , Gotas Lipídicas , Animales , Autofagia/fisiología , Proteína Sequestosoma-1/metabolismo , Gotas Lipídicas/metabolismo , Proteínas/metabolismo , Ubiquitina/metabolismo , Proteínas Portadoras/metabolismo , Mamíferos/metabolismoRESUMEN
Recently, we developed Komagataella phaffii (formerly Pichia pastoris) as a model for lipophagy, the selective autophagy of lipid droplets (LDs). We found that lipophagy pathways induced by acute nitrogen (N) starvation and in stationary (S) phase have different molecular mechanisms. Moreover, both types of lipophagy are independent of Atg11, the scaffold protein that interacts with most autophagic receptors and, therefore, is essential for most types of selective autophagy in yeast. Since yeast aggrephagy, the selective autophagy of ubiquitinated protein aggregates, is also independent of Atg11 and utilizes the ubiquitin-binding receptor, Cue5, we studied the relationship of K. phaffii Cue5 with differentially induced LDs and lipophagy. While there was no relationship of Cue5 with LDs and lipophagy under N-starvation conditions, Cue5 accumulated on LDs in S-phase and degraded together with LDs via S-phase lipophagy. The accumulation of Cue5 on LDs and its degradation by S-phase lipophagy strongly depended on the ubiquitin-binding CUE domain and Prl1, the positive regulator of lipophagy 1. However, unlike Prl1, which is required for S-phase lipophagy, Cue5 was dispensable for it suggesting that Cue5 is rather a new substrate of this pathway. We propose that a similar mechanism (Prl1-dependent accumulation on LDs) might be employed by Prl1 to recruit another ubiquitin-binding protein that is essential for S-phase lipophagy.
Asunto(s)
Autofagia , Proteínas Fúngicas/metabolismo , Gotas Lipídicas/metabolismo , Proteolisis , Saccharomycetales/metabolismo , Vacuolas/metabolismo , Modelos BiológicosRESUMEN
Coronavirus disease 2019 (COVID-19), caused by a new severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has instantaneously emerged as a worldwide pandemic. However, humans encountered other coronaviruses in the past, and they caused a broad range of symptoms, from mild to life-threatening, depending on the virus and immunocompetence of the host. Most human coronaviruses interact with the proteins and/or double-membrane vesicles of autophagy, the membrane trafficking pathway that degrades and recycles the intracellular protein aggregates, organelles, and pathogens, including viruses. However, coronaviruses often neutralize and hijack this pathway to complete their life cycle. In this review, we discuss the interactions of human coronaviruses and autophagy, including recent data from SARS-CoV-2-related studies. Some of these interactions (for example, viral block of the autophagosome-lysosome fusion), while being conserved across multiple coronaviruses, are accomplished via different molecular mechanisms. Therefore, it is important to understand the molecular interplay between human coronaviruses and autophagy for developing efficient therapies against coronaviral diseases.
Asunto(s)
Autofagia , Infecciones por Coronavirus/fisiopatología , Coronavirus/metabolismo , COVID-19/metabolismo , COVID-19/fisiopatología , Infecciones por Coronavirus/metabolismo , Humanos , Lisosomas , SARS-CoV-2/metabolismoRESUMEN
PROBLEM: Candida albicans infection induces damage in host cells that results in the activation of cell death pathways such as apoptosis and necrosis. Autophagy acts as a pro-survival mechanism during various infections and has cross talks with apoptosis. The objective of our study was to delineate the effect of autophagy knockdown in human vaginal epithelial cells (VECs) during Candida infection-induced apoptosis. METHOD OF STUDY: We overexpressed wild-type or mutant form of autophagy-related gene 5 (ATG5) in human VECs and determined the levels of autophagy and lysosome marker genes upon C. albicans infection. We analyzed the expression of apoptosis and necrosis markers using confocal microscopy and flow cytometry. RESULTS: Mutant ATG5 overexpressing VECs were unable to form Atg5-Atg12 complex, which is required for functional autophagy pathway. Upon Candida albicans infection, wild-type ATG5 overexpressing cells showed upregulation of autophagy marker genes, ATG5, LC3-I, LC3-II, and LAMP-1. Mutant ATG5 overexpressing cells could not upregulate LC3-II and LAMP-1 expression, indicating a defective autophagy pathway. Confocal microscopy and flow cytometry results revealed that wild-type ATG5 overexpressing VECs showed significantly lower number of apoptotic and necrotic cells as compared to mutant ATG5 overexpressing cells. CONCLUSION: Vaginal epithelial cells with active autophagy were able to survive the damage caused by C. albicans infection, whereas those with defective autophagy succumbed to the infection. This suggests that autophagy plays a critical role in human VECs survival during C. albicans infection.
Asunto(s)
Proteína 5 Relacionada con la Autofagia/metabolismo , Candida albicans/fisiología , Candidiasis/metabolismo , Células Epiteliales/metabolismo , Vagina/patología , Apoptosis/genética , Autofagia/genética , Proteína 12 Relacionada con la Autofagia/metabolismo , Proteína 5 Relacionada con la Autofagia/genética , Células Cultivadas , Resistencia a la Enfermedad , Células Epiteliales/microbiología , Células Epiteliales/patología , Femenino , Citometría de Flujo , Técnicas de Silenciamiento del Gen , Humanos , Microscopía Confocal , Mutación/genética , Unión Proteica , Transgenes/genéticaRESUMEN
The data presented here are related to the research article entitled "Knockout of autophagy gene, ATG5 in mice vaginal cells abrogates cytokine response and pathogen clearance during vaginal infection of Candida albicans" (Shroff et al., 2018) [1]. The cited research article describes the role of autophagy in host immune response against C. albicans infection of mice vagina. In this data report wild-type C57BL/6 mice were infected intravaginally with C. albicans. Vaginal cells were analyzed for the expression of autophagy marker genes LC3 & ATG5 and lysosome marker LAMP1 at the transcript and protein level. Vaginal lavages were also obtained from these infected mice. The levels of pro-inflammatory and T-helper cell related cytokines were determined in these lavages.
RESUMEN
The female reproductive tract (FRT) presents a unique challenge to the mucosal immune system as it needs to monitor constantly for the presence of opportunistic pathogens amidst its commensal flora. During infection, autophagy plays a critical role in pathogen clearance, presentation of antigens and production of pro-inflammatory cytokines. However, no information is available that describes the role of autophagy in mouse vaginal infection of Candida albicans. The objective of our study is to evaluate the effect of autophagy gene, ATG5 knockout in vaginal cells in response to vaginal C. albicans infection. Mice having knockout of ATG5 in the vaginal cells (PR-ATG5-KO mice) were infected intra-vaginally with the yeast form of Candida albicans. Vaginal lavages were collected once in a week until the infection was cleared. We detected the expression of autophagy marker genes (LC3, ATG5 and LAMP1) in the vaginal cells. We determined the levels of various cytokines (IL-1α, IL-1ß, IL-6, IL-10, IL-17A, IL-22, IL-23p19, TNF-α and G-CSF) involved in anti-candida response. The levels of cytokines in the vaginal lavages were quantified using Aimplex Premixed analyte kit. The vaginal lavages were checked for polymorphonuclear leucocytes (PMNLs) infiltration. The candida clearance rate from the vaginal lumen was determined by Colony Forming Units (CFUs) assay. The results revealed that PR-ATG5-KO mice failed to induce the expression of LC3, ATG5 and LAMP1 indicating an impaired autophagy pathway. The levels of all the cytokines (except IL-10) in C. albicans infected PR-ATG5-KO mice were significantly reduced as compared to the wild type infected C57BL/6 mice. The number of PMNLs infiltrated into the vaginal lavages of infected PR-ATG5-KO mice was reduced. The clearance of C. albicans from the vaginal lumen was also considerably delayed in PR-ATG5-KO mice. In conclusion, the results revealed that impaired autophagy in vaginal cells influences host response during vaginal infection of C. albicans by affecting anti-Candida cytokine levels in the vaginal lavage resulting in reduction of pathogen clearance rate.
Asunto(s)
Proteína 5 Relacionada con la Autofagia/genética , Candida albicans/inmunología , Candidiasis Vulvovaginal/inmunología , Citocinas/inmunología , Animales , Autofagia/genética , Proteína 5 Relacionada con la Autofagia/inmunología , Candida albicans/patogenicidad , Candidiasis Vulvovaginal/genética , Candidiasis Vulvovaginal/microbiología , Candidiasis Vulvovaginal/patología , Femenino , Técnicas de Inactivación de Genes , Interleucinas/inmunología , Ratones , Ratones Noqueados , Neutrófilos/inmunología , Células Th17/inmunologíaRESUMEN
PROBLEM: Autophagy plays an important role in clearance of intracellular pathogens. However, no information is available on its involvement in vaginal infections such as vulvo-vaginal candidiasis (VVC). VVC is intimately associated with the immune status of the human vaginal epithelial cells (VECs). The objective of our study is to decipher if autophagy process is involved during Candida albicans infection of VECs. METHODS OF STUDY: In this study, C. albicans infection system was established using human VEC line (VK2/E6E7). Infection-induced change in the expression of autophagy markers like LC3 and LAMP-1 were analyzed by RT-PCR, q-PCR, Western blot, immunofluorescence and transmission electron microscopy (TEM) studies were carried out to ascertain the localization of autophagosomes. Multiplex ELISA was carried out to determine the cytokine profiles. RESULTS: Analysis of LC3 and LAMP-1 expression at mRNA and protein levels at different time points revealed up-regulation of these markers 6 hours post C. albicans infection. LC3 and LAMP-1 puncti were observed in infected VECs after 12 hours. TEM studies showed C. albicans entrapped in autophagosomes. Cytokines-TNF-α and IL-1ß were up-regulated in culture supernatants of VECs at 12 hours post-infection. CONCLUSION: The results suggest that C. albicans invasion led to the activation of autophagy as a host defense mechanism of VECs.
Asunto(s)
Autofagia/fisiología , Candidiasis Vulvovaginal/patología , Células Epiteliales/microbiología , Vagina/microbiología , Biomarcadores/análisis , Western Blotting , Línea Celular , Ensayo de Inmunoadsorción Enzimática , Células Epiteliales/patología , Femenino , Técnica del Anticuerpo Fluorescente , Humanos , Microscopía Electrónica de Transmisión , Reacción en Cadena de la Polimerasa , Vagina/citología , Vagina/patologíaRESUMEN
Hemoglobin (Hb) is a major protein involved in transport of oxygen (O2). Red blood cells (RBCs) contain maximum amount of Hb and because of their unique structure and plasticity they transport O2 to various tissues of the body at an optimal concentration. Recently, it has been reported that, apart from RBCs, Hb is also expressed by nonerythroid cells such as epithelial cells of different origin. The cells expressing Hb are from the tissues where maintenance of O2 homeostasis is of paramount importance. Hb expression has been observed in the epithelial cells from human tissues including lungs, neurons, retina, and endometrium. Our group has recently demonstrated that Hb is expressed by the cervicovaginal epithelial cells. We further showed that, apart from maintaining O2 homeostasis, Hb and the peptides derived from it play an indispensable role in the protection of vaginal epithelium by exhibiting antimicrobial activity. In this review, we discuss the significance of Hb expression in vaginal epithelial cells and its role in the recognition of pathogens thereby reducing the risk and/or severity of inflammation and/or infections and the possible mechanism by which Hb exhibits antimicrobial and antioxidative functions.