Journal articles: 'Atg18'

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Sawa-Makarska, Justyna, Verena Baumann, Nicolas Coudevylle, Sören von Bülow, Veronika Nogellova, Christine Abert, Martina Schuschnig, Martin Graef, Gerhard Hummer, and Sascha Martens. "Reconstitution of autophagosome nucleation defines Atg9 vesicles as seeds for membrane formation." Science 369, no. 6508 (September 3, 2020): eaaz7714. http://dx.doi.org/10.1126/science.aaz7714.

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Autophagosomes form de novo in a manner that is incompletely understood. Particularly enigmatic are autophagy-related protein 9 (Atg9)–containing vesicles that are required for autophagy machinery assembly but do not supply the bulk of the autophagosomal membrane. In this study, we reconstituted autophagosome nucleation using recombinant components from yeast. We found that Atg9 proteoliposomes first recruited the phosphatidylinositol 3-phosphate kinase complex, followed by Atg21, the Atg2-Atg18 lipid transfer complex, and the E3-like Atg12–Atg5-Atg16 complex, which promoted Atg8 lipidation. Furthermore, we found that Atg2 could transfer lipids for Atg8 lipidation. In selective autophagy, these reactions could potentially be coupled to the cargo via the Atg19-Atg11-Atg9 interactions. We thus propose that Atg9 vesicles form seeds that establish membrane contact sites to initiate lipid transfer from compartments such as the endoplasmic reticulum.

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Hegedűs, Krisztina, Péter Nagy, Zoltán Gáspári, and Gábor Juhász. "The Putative HORMA Domain Protein Atg101 Dimerizes and Is Required for Starvation-Induced and Selective Autophagy inDrosophila." BioMed Research International 2014 (2014): 1–13. http://dx.doi.org/10.1155/2014/470482.

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The large-scale turnover of intracellular material including organelles is achieved by autophagy-mediated degradation in lysosomes. Initiation of autophagy is controlled by a protein kinase complex consisting of an Atg1-family kinase, Atg13, FIP200/Atg17, and the metazoan-specific subunit Atg101. Here we show that loss of Atg101 impairs both starvation-induced and basal autophagy inDrosophila. This leads to accumulation of protein aggregates containing the selective autophagy cargo ref(2)P/p62. Mapping experiments suggest that Atg101 binds to the N-terminal HORMA domain of Atg13 and may also interact with two unstructured regions of Atg1. Another HORMA domain-containing protein, Mad2, forms a conformational homodimer. We show thatDrosophilaAtg101 also dimerizes, and it is predicted to fold into a HORMA domain. Atg101 interacts with ref(2)P as well, similar to Atg13, Atg8a, Atg16, Atg18, Keap1, and RagC, a known regulator of Tor kinase which coordinates cell growth and autophagy. These results raise the possibility that the interactions and dimerization of the putative HORMA domain protein Atg101 play critical roles in starvation-induced autophagy and proteostasis, by promoting the formation of protein aggregate-containing autophagosomes.

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Efe, Jem A., Roberto J. Botelho, and Scott D. Emr. "Atg18 Regulates Organelle Morphology and Fab1 Kinase Activity Independent of Its Membrane Recruitment by Phosphatidylinositol 3,5-Bisphosphate." Molecular Biology of the Cell 18, no. 11 (November 2007): 4232–44. http://dx.doi.org/10.1091/mbc.e07-04-0301.

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The lipid kinase Fab1 governs yeast vacuole homeostasis by generating PtdIns(3,5)P2on the vacuolar membrane. Recruitment of effector proteins by the phospholipid ensures precise regulation of vacuole morphology and function. Cells lacking the effector Atg18p have enlarged vacuoles and high PtdIns(3,5)P2levels. Although Atg18 colocalizes with Fab1p, it likely does not directly interact with Fab1p, as deletion of either kinase activator—VAC7 or VAC14—is epistatic to atg18Δ: atg18Δvac7Δ cells have no detectable PtdIns(3,5)P2. Moreover, a 2xAtg18 (tandem fusion) construct localizes to the vacuole membrane in the absence of PtdIns(3,5)P2, but requires Vac7p for recruitment. Like the endosomal PtdIns(3)P effector EEA1, Atg18 membrane binding may require a protein component. When the lipid requirement is bypassed by fusing Atg18 to ALP, a vacuolar transmembrane protein, vac14Δ vacuoles regain normal morphology. Rescue is independent of PtdIns(3,5)P2, as mutation of the phospholipid-binding site in Atg18 does not prevent vacuole fission and properly regulates Fab1p activity. Finally, the vacuole-specific type-V myosin adapter Vac17p interacts with Atg18p, perhaps mediating cytoskeletal attachment during retrograde transport. Atg18p is likely a PtdIns(3,5)P2“sensor,” acting as an effector to remodel membranes as well as regulating its synthesis via feedback that might involve Vac7p.

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Strømhaug, Per E., Fulvio Reggiori, Ju Guan, Chao-Wen Wang, and Daniel J. Klionsky. "Atg21 Is a Phosphoinositide Binding Protein Required for Efficient Lipidation and Localization of Atg8 during Uptake of Aminopeptidase I by Selective Autophagy." Molecular Biology of the Cell 15, no. 8 (August 2004): 3553–66. http://dx.doi.org/10.1091/mbc.e04-02-0147.

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Delivery of proteins and organelles to the vacuole by autophagy and the cytoplasm to vacuole targeting (Cvt) pathway involves novel rearrangements of membrane resulting in the formation of vesicles that fuse with the vacuole. The mechanism of vesicle formation and the origin of the membrane are complex issues still to be resolved. Atg18 and Atg21 are proteins essential to vesicle formation and together with Ygr223c form a novel family of phosphoinositide binding proteins that are associated with the vacuole and perivacuolar structures. Their localization requires the activity of Vps34, suggesting that phosphatidylinositol(3)phosphate may be essential for their function. The activity of Atg18 is vital for all forms of autophagy, whereas Atg21 is required for the Cvt pathway but not for nitrogen starvation-induced autophagy. The loss of Atg21 results in the absence of Atg8 from the pre-autophagosomal structure (PAS), which may be ascribed to a reduced rate of conjugation of Atg8 to phosphatidylethanolamine. A similar defect in localization of a second ubiquitin-like conjugate, Atg12-Atg5, suggests that Atg21 may be involved in the recruitment of membrane to the PAS.

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Aslan, Erhan, Nurçin Küçükoğlu, and Muhittin Arslanyolu. "A comparative in-silico analysis of autophagy proteins in ciliates." PeerJ 5 (January 17, 2017): e2878. http://dx.doi.org/10.7717/peerj.2878.

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Autophagy serves as a turnover mechanism for the recycling of redundant and/or damaged macromolecules present in eukaryotic cells to re-use them under starvation conditions via a double-membrane structure known as autophagosome. A set of eukaryotic genes called autophagy-related genes (ATGs) orchestrate this highly elaborative process. The existence of these genes and the role they play in different eukaryotes are well-characterized. However, little is known of their role in some eukaryotes such as ciliates. Here, we report the computational analyses of ATG genes in five ciliate genomes to understand their diversity. Our results show that Oxytricha trifallax is the sole ciliate which has a conserved Atg12 conjugation system (Atg5-Atg12-Atg16). Interestingly, Oxytricha Atg16 protein includes WD repeats in addition to its N-terminal Atg16 domain as is the case in multicellular organisms. Additionally, phylogenetic analyses revealed that E2-like conjugating protein Atg10 is only present in Tetrahymena thermophila. We fail to find critical autophagy components Atg5, Atg7 and Atg8 in the parasitic ciliate Ichthyophthirius multifiliis. Contrary to previous reports, we also find that ciliate genomes do not encode typical Atg1 since all the candidate sequences lack an Atg1-specific C-terminal domain which is essential for Atg1 complex formation. Consistent with the absence of Atg1, ciliates also lack other members of the Atg1 complex. However, the presence of Atg6 in all ciliates examined here may rise the possibility that autophagosome formation could be operated through Atg6 in ciliates, since Atg6 has been shown as an alternative autophagy inducer. In conclusion, our results highlight that Atg proteins are partially conserved in ciliates. This may provide a better understanding for the autophagic destruction of the parental macronucleus, a developmental process also known as programmed nuclear death in ciliates.

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Nakatogawa, Hitoshi. "Two ubiquitin-like conjugation systems that mediate membrane formation during autophagy." Essays in Biochemistry 55 (September 27, 2013): 39–50. http://dx.doi.org/10.1042/bse0550039.

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In autophagy, the autophagosome, a transient organelle specialized for the sequestration and lysosomal or vacuolar transport of cellular constituents, is formed via unique membrane dynamics. This process requires concerted actions of a distinctive set of proteins named Atg (autophagy-related). Atg proteins include two ubiquitin-like proteins, Atg12 and Atg8 [LC3 (light-chain 3) and GABARAP (γ-aminobutyric acid receptor-associated protein) in mammals]. Sequential reactions by the E1 enzyme Atg7 and the E2 enzyme Atg10 conjugate Atg12 to the lysine residue in Atg5, and the resulting Atg12–Atg5 conjugate forms a complex with Atg16. On the other hand, Atg8 is first processed at the C-terminus by Atg4, which is related to ubiquitin-processing/deconjugating enzymes. Atg8 is then activated by Atg7 (shared with Atg12) and, via the E2 enzyme Atg3, finally conjugated to the amino group of the lipid PE (phosphatidylethanolamine). The Atg12–Atg5–Atg16 complex acts as an E3 enzyme for the conjugation reaction of Atg8; it enhances the E2 activity of Atg3 and specifies the site of Atg8–PE production to be autophagy-related membranes. Atg8–PE is suggested to be involved in autophagosome formation at multiple steps, including membrane expansion and closure. Moreover, Atg4 cleaves Atg8–PE to liberate Atg8 from membranes for reuse, and this reaction can also regulate autophagosome formation. Thus these two ubiquitin-like systems are intimately involved in driving the biogenesis of the autophagosomal membrane.

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Noda, Nobuo. "Structural basis of Atg conjugation systems essential for autophagy." Acta Crystallographica Section A Foundations and Advances 70, a1 (August 5, 2014): C302. http://dx.doi.org/10.1107/s2053273314096971.

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Autophagy is an evolutionarily-conserved, intracellular degradation system for which two ubiquitin-like modifiers, Atg8 and Atg12, play essential roles. After processed by Atg4, the exposed C-terminal glycine of Atg8 is activated by Atg7 (E1) and is then transferred to Atg3 (E2), and is finally conjugated with a phospholipid, phosphatidylethanolamine (PE) through an amide bond. Whereas, Atg12 is activated by the same E1, Atg7, without processing, and is then transferred to Atg10 (E2), and is finally conjugated with Atg5 through an isopeptide bond. Atg12-Atg5 conjugates, together with Atg16, function as an E3-like enzyme to facilitate the conjugation reaction between Atg8 and PE. During autophagy, Atg8-PE conjugates play a critical role in selective cargo recognition in addition to autophagosome formation. We determined the structures of all these Atg proteins and their complexes mainly by X-ray crystallography, and performed structure-based biochemical analyses on them [1,2]. These studies established the molecular mechanisms of Atg8 and Atg12 modification reactions that have many unique features compared with canonical ubiquitin-like systems. Furthermore, we found a conserved motif named the Atg8-family interacting motif (AIM), through which Atg8 recognizes specific cargoes and selectively incorporates them into autophagosomes for degradation [3].

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Kabeya, Yukiko, Yoshiaki Kamada, Misuzu Baba, Hirosato Takikawa, Mitsuru Sasaki, and Yoshinori Ohsumi. "Atg17 Functions in Cooperation with Atg1 and Atg13 in Yeast Autophagy." Molecular Biology of the Cell 16, no. 5 (May 2005): 2544–53. http://dx.doi.org/10.1091/mbc.e04-08-0669.

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In eukaryotic cells, nutrient starvation induces the bulk degradation of cellular materials; this process is called autophagy. In the yeast Saccharomyces cerevisiae, most of the ATG (autophagy) genes are involved in not only the process of degradative autophagy, but also a biosynthetic process, the cytoplasm to vacuole (Cvt) pathway. In contrast, the ATG17 gene is required specifically in autophagy. To better understand the function of Atg17, we have performed a biochemical characterization of the Atg17 protein. We found that the atg17Δ mutant under starvation condition was largely impaired in autophagosome formation and only rarely contained small autophagosomes, whose size was less than one-half of normal autophagosomes in diameter. Two-hybrid analyses and coimmunoprecipitation experiments demonstrated that Atg17 physically associates with Atg1-Atg13 complex, and this binding was enhanced under starvation conditions. Atg17-Atg1 binding was not detected in atg13Δ mutant cells, suggesting that Atg17 interacts with Atg1 through Atg13. A point mutant of Atg17, Atg17C24R, showed reduced affinity for Atg13, resulting in impaired Atg1 kinase activity and significant defects in autophagy. Taken together, these results indicate that Atg17-Atg13 complex formation plays an important role in normal autophagosome formation via binding to and activating the Atg1 kinase.

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Tamura, Naoki, Masahide Oku, Moemi Ito, Nobuo N. Noda, Fuyuhiko Inagaki, and Yasuyoshi Sakai. "Atg18 phosphoregulation controls organellar dynamics by modulating its phosphoinositide-binding activity." Journal of Cell Biology 202, no. 4 (August 12, 2013): 685–98. http://dx.doi.org/10.1083/jcb.201302067.

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The PROPPIN family member Atg18 is a phosphoinositide-binding protein that is composed of a seven β-propeller motif and is part of the conserved autophagy machinery. Here, we report that the Atg18 phosphorylation in the loops in the propellar structure of blade 6 and blade 7 decreases its binding affinity to phosphatidylinositol 3,5-bisphosphate in the yeast Pichia pastoris. Dephosphorylation of Atg18 was necessary for its association with the vacuolar membrane and caused septation of the vacuole. Upon or after dissociation from the vacuolar membrane, Atg18 was rephosphorylated, and the vacuoles fused and formed a single rounded structure. Vacuolar dynamics were regulated according to osmotic changes, oxidative stresses, and nutrient conditions inducing micropexophagy via modulation of Atg18 phosphorylation. This study reveals how the phosphoinositide-binding activity of the PROPPIN family protein Atg18 is regulated at the membrane association domain and highlights the importance of such phosphoregulation in coordinated intracellular reorganization.

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Melani, Mariana, Ayelén Valko, Nuria M. Romero, Milton O. Aguilera, Julieta M. Acevedo, Zambarlal Bhujabal, Joel Perez-Perri, et al. "Zonda is a novel early component of the autophagy pathway in Drosophila." Molecular Biology of the Cell 28, no. 22 (November 2017): 3070–81. http://dx.doi.org/10.1091/mbc.e16-11-0767.

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Autophagy is an evolutionary conserved process by which eukaryotic cells undergo self-digestion of cytoplasmic components. Here we report that a novel Drosophila immunophilin, which we have named Zonda, is critically required for starvation-induced autophagy. We show that Zonda operates at early stages of the process, specifically for Vps34-mediated phosphatidylinositol 3-phosphate (PI3P) deposition. Zonda displays an even distribution under basal conditions and, soon after starvation, nucleates in endoplasmic reticulum–associated foci that colocalize with omegasome markers. Zonda nucleation depends on Atg1, Atg13, and Atg17 but does not require Vps34, Vps15, Atg6, or Atg14. Zonda interacts physically with Atg1 through its kinase domain, as well as with Atg6 and Vps34. We propose that Zonda is an early component of the autophagy cascade necessary for Vps34-dependent PI3P deposition and omegasome formation.

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Chang, Yu-Yun, and Thomas P. Neufeld. "An Atg1/Atg13 Complex with Multiple Roles in TOR-mediated Autophagy Regulation." Molecular Biology of the Cell 20, no. 7 (April 2009): 2004–14. http://dx.doi.org/10.1091/mbc.e08-12-1250.

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The TOR kinases are conserved negative regulators of autophagy in response to nutrient conditions, but the signaling mechanisms are poorly understood. Here we describe a complex containing the protein kinase Atg1 and the phosphoprotein Atg13 that functions as a critical component of this regulation in Drosophila. We show that knockout of Atg1 or Atg13 results in a similar, selective defect in autophagy in response to TOR inactivation. Atg1 physically interacts with TOR and Atg13 in vivo, and both Atg1 and Atg13 are phosphorylated in a nutrient-, TOR- and Atg1 kinase-dependent manner. In contrast to yeast, phosphorylation of Atg13 is greatest under autophagic conditions and does not preclude Atg1-Atg13 association. Atg13 stimulates both the autophagic activity of Atg1 and its inhibition of cell growth and TOR signaling, in part by disrupting the normal trafficking of TOR. In contrast to the effects of normal Atg13 levels, increased expression of Atg13 inhibits autophagosome expansion and recruitment of Atg8/LC3, potentially by decreasing the stability of Atg1 and facilitating its inhibitory phosphorylation by TOR. Atg1-Atg13 complexes thus function at multiple levels to mediate and adjust nutrient-dependent autophagic signaling.

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Gómez-Sánchez, Rubén, Jaqueline Rose, Rodrigo Guimarães, Muriel Mari, Daniel Papinski, Ester Rieter, Willie J. Geerts, et al. "Atg9 establishes Atg2-dependent contact sites between the endoplasmic reticulum and phagophores." Journal of Cell Biology 217, no. 8 (May 30, 2018): 2743–63. http://dx.doi.org/10.1083/jcb.201710116.

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The autophagy-related (Atg) proteins play a key role in the formation of autophagosomes, the hallmark of autophagy. The function of the cluster composed by Atg2, Atg18, and transmembrane Atg9 is completely unknown despite their importance in autophagy. In this study, we provide insights into the molecular role of these proteins by identifying and characterizing Atg2 point mutants impaired in Atg9 binding. We show that Atg2 associates to autophagosomal membranes through lipid binding and independently from Atg9. Its interaction with Atg9, however, is key for Atg2 confinement to the growing phagophore extremities and subsequent association of Atg18. Assembly of the Atg9–Atg2–Atg18 complex is important to establish phagophore–endoplasmic reticulum (ER) contact sites. In turn, disruption of the Atg2–Atg9 interaction leads to an aberrant topological distribution of both Atg2 and ER contact sites on forming phagophores, which severely impairs autophagy. Altogether, our data shed light in the interrelationship between Atg9, Atg2, and Atg18 and highlight the possible functional relevance of the phagophore–ER contact sites in phagophore expansion.

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Cheong, Heesun, Usha Nair, Jiefei Geng, and Daniel J. Klionsky. "The Atg1 Kinase Complex Is Involved in the Regulation of Protein Recruitment to Initiate Sequestering Vesicle Formation for Nonspecific Autophagy in Saccharomyces cerevisiae." Molecular Biology of the Cell 19, no. 2 (February 2008): 668–81. http://dx.doi.org/10.1091/mbc.e07-08-0826.

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Autophagy is the major degradative process for recycling cytoplasmic constituents and eliminating unnecessary organelles in eukaryotic cells. Most autophagy-related (Atg) proteins are recruited to the phagophore assembly site (PAS), a proposed site for vesicle formation during either nonspecific or specific types of autophagy. Therefore, appropriate recruitment of Atg proteins to this site is critical for their function in autophagy. Atg11 facilitates PAS recruitment for the cytoplasm-to-vacuole targeting pathway, which is a specific, autophagy-like process that occurs under vegetative conditions. In contrast, it is not known how Atg proteins are recruited to the PAS, nor which components are involved in PAS formation under nonspecific autophagy-inducing, starvation conditions. Here, we studied PAS assembly during nonspecific autophagy, using an atg11Δ mutant background to eliminate the PAS formation that occurs during vegetative growth. We found that protein complexes containing the Atg1 kinase have two roles for PAS formation during nonspecific autophagy. The Atg1 C terminus mediates an interaction with Atg13 and Atg17, facilitating a structural role of Atg1 that is needed to efficiently organize an initial step of PAS assembly, whereas Atg1 kinase activity affects the dynamics of protein movement at the PAS involved in Atg protein cycling.

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Kotani, Tetsuya, Hiromi Kirisako, Michiko Koizumi, Yoshinori Ohsumi, and Hitoshi Nakatogawa. "The Atg2-Atg18 complex tethers pre-autophagosomal membranes to the endoplasmic reticulum for autophagosome formation." Proceedings of the National Academy of Sciences 115, no. 41 (September 25, 2018): 10363–68. http://dx.doi.org/10.1073/pnas.1806727115.

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The biogenesis of double-membrane vesicles called autophagosomes, which sequester and transport intracellular material for degradation in lysosomes or vacuoles, is a central event in autophagy. This process requires a unique set of factors called autophagy-related (Atg) proteins. The Atg proteins assemble to organize the preautophagosomal structure (PAS), at which a cup-shaped membrane, the isolation membrane (or phagophore), forms and expands to become the autophagosome. The molecular mechanism of autophagosome biogenesis remains poorly understood. Previous studies have shown that Atg2 forms a complex with the phosphatidylinositol 3-phosphate (PI3P)-binding protein Atg18 and localizes to the PAS to initiate autophagosome biogenesis; however, the molecular function of Atg2 remains unknown. In this study, we show that Atg2 has two membrane-binding domains in the N- and C-terminal regions and acts as a membrane tether during autophagosome formation in the budding yeast Saccharomyces cerevisiae. An amphipathic helix in the C-terminal region binds to membranes and facilitates Atg18 binding to PI3P to target the Atg2-Atg18 complex to the PAS. The N-terminal region of Atg2 is also involved in the membrane binding of this protein but is dispensable for the PAS targeting of the Atg2-Atg18 complex. Our data suggest that this region associates with the endoplasmic reticulum (ER) and is responsible for the formation of the isolation membrane at the PAS. Based on these results, we propose that the Atg2-Atg18 complex tethers the PAS to the ER to initiate membrane expansion during autophagosome formation.

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Chang, Chiung-Ying, and Wei-Pang Huang. "Atg19 Mediates a Dual Interaction Cargo Sorting Mechanism in Selective Autophagy." Molecular Biology of the Cell 18, no. 3 (March 2007): 919–29. http://dx.doi.org/10.1091/mbc.e06-08-0683.

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Autophagy is a catabolic membrane-trafficking mechanism conserved in all eukaryotic cells. In addition to the nonselective transport of bulk cytosol, autophagy is responsible for efficient delivery of the vacuolar enzyme Ape1 precursor (prApe1) in the budding yeast Saccharomyces cerevisiae, suggesting the presence of a prApe1 sorting machinery. Sequential interactions between Atg19-Atg11 and Atg19-Atg8 pairs are thought responsible for targeting prApe1 to the vesicle formation site, the preautophagosomal structure (PAS), and loading it into transport vesicles, respectively. However, the different patterns of prApe1 transport defect seen in the atg11Δ and atg19Δ strains seem to be incompatible with this model. Here we report that prApe1 could not be targeted to the PAS and failed to be delivered into the vacuole in atg8Δ atg11Δ double knockout cells regardless of the nutrient conditions. We postulate that Atg19 mediates a dual interaction prApe1-sorting mechanism through independent, instead of sequential, interactions with Atg11 and Atg8. In addition, to efficiently deliver prApe1 to the vacuole, a proper interaction between Atg11 and Atg9 is indispensable. We speculate that Atg11 may elicit a cargo-loading signal and induce Atg9 shuttling to a specific PAS site, where Atg9 relays the signal and recruits other Atg proteins to induce vesicle formation.

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Sudhakar, Renu, Divya Das, Subramanian Thanumalayan, Somesh Gorde, and Puran Singh Sijwali. "Plasmodium falciparum Atg18 localizes to the food vacuole via interaction with the multi-drug resistance protein 1 and phosphatidylinositol 3-phosphate." Biochemical Journal 478, no. 9 (May 10, 2021): 1705–32. http://dx.doi.org/10.1042/bcj20210001.

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Autophagy, a lysosome-dependent degradative process, does not appear to be a major degradative process in malaria parasites and has a limited repertoire of genes. To better understand the autophagy process, we investigated Plasmodium falciparum Atg18 (PfAtg18), a PROPPIN family protein, whose members like S. cerevisiae Atg18 (ScAtg18) and human WIPI2 bind PI3P and play an essential role in autophagosome formation. Wild type and mutant PfAtg18 were expressed in P. falciparum and assessed for localization, the effect of various inhibitors and antimalarials on PfAtg18 localization, and identification of PfAtg18-interacting proteins. PfAtg18 is expressed in asexual erythrocytic stages and localized to the food vacuole, which was also observed with other Plasmodium Atg18 proteins, indicating that food vacuole localization is likely a shared feature. Interaction of PfAtg18 with the food vacuole-associated PI3P is essential for localization, as PfAtg18 mutants of PI3P-binding motifs neither bound PI3P nor localized to the food vacuole. Interestingly, wild type ScAtg18 interacted with PI3P, but its expression in P. falciparum showed complete cytoplasmic localization, indicating additional requirement for food vacuole localization. The food vacuole multi-drug resistance protein 1 (MDR1) was consistently identified in the immunoprecipitates of PfAtg18 and P. berghei Atg18, and also interacted with PfAtg18. In contrast with PfAtg18, ScAtg18 did not interact with MDR1, which, in addition to PI3P, could play a critical role in localization of PfAtg18. Chloroquine and amodiaquine caused cytoplasmic localization of PfAtg18, suggesting that these target PfAtg18 transport pathway. Thus, PI3P and MDR1 are critical mediators of PfAtg18 localization.

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Fang, Yao, Zhang, Tian, Wang, Li, and Cai. "iTRAQ-Based Proteomics Analysis of Autophagy-Mediated Responses against MeJA in Laticifers of Euphorbia kansui L." International Journal of Molecular Sciences 20, no. 15 (August 1, 2019): 3770. http://dx.doi.org/10.3390/ijms20153770.

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Autophagy is a well-defined catabolic mechanism whereby cytoplasmic materials are engulfed into a structure termed the autophagosome. Methyl jasmonate (MeJA), a plant hormone, mediates diverse developmental process and defense responses which induce a variety of metabolites. In plants, little is known about autophagy-mediated responses against MeJA. In this study, we used high-throughput comparative proteomics to identify proteins of latex in the laticifers. The isobaric tags for relative and absolute quantification (iTRAQ) MS/MS proteomics were performed, and 298 proteins among MeJA treated groups and the control group of Euphorbia kansui were identified. It is interesting to note that 29 significant differentially expressed proteins were identified and their associations with autophagy and ROS pathway were verified for several selected proteins as follows: α-L-fucosidase, β-galactosidase, cysteine proteinase, and Cu/Zn superoxide dismutase. Quantitative real-time PCR analysis of the selected genes confirmed the fact that MeJA might enhance the expression of some genes related to autophagy. The western blotting and immunofluorescence results of ATG8 and ATG18a which are two important proteins for the formation of autophagosomes also demonstrated that MeJA could promote autophagy at the protein level. Using the electron microscope, we observed an increase in autophagosomes after MeJA treatment. These results indicated that MeJA might promote autophagy in E. kansui laticifers; and it was speculated that MeJA mediated autophagy through two possible ways: the increase of ROS induces ATG8 accumulation and then aotophagosome formation, and MeJA promotes ATG18 accumulation and then autophagosome formation. Taken together, our results provide several novel insights for understanding the mechanism between autophagy and MeJA treatment. However, the specific mechanism remains to be further studied in the future.

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Lin, Mary G., Johannes Schöneberg, Christopher W. Davies, Xuefeng Ren, and James H. Hurley. "The dynamic Atg13-free conformation of the Atg1 EAT domain is required for phagophore expansion." Molecular Biology of the Cell 29, no. 10 (May 15, 2018): 1228–37. http://dx.doi.org/10.1091/mbc.e17-04-0258.

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Yeast macroautophagy begins with the de novo formation of a double-membrane phagophore at the preautophagosomal structure/phagophore assembly site (PAS), followed by its expansion into the autophagosome responsible for cargo engulfment. The kinase Atg1 is recruited to the PAS by Atg13 through interactions between the EAT domain of the former and the tMIM motif of the latter. Mass-spectrometry data have shown that, in the absence of Atg13, the EAT domain structure is strikingly dynamic, but the function of this Atg13-free dynamic state has been unclear. We used structure-based mutational analysis and quantitative and superresolution microscopy to show that Atg1 is present on autophagic puncta at, on average, twice the stoichiometry of Atg13. Moreover, Atg1 colocalizes with the expanding autophagosome in a manner dependent on Atg8 but not Atg13. We used isothermal titration calorimetry and crystal structure information to design an EAT domain mutant allele ATG1DD that selectively perturbs the function of the Atg13-free state. Atg1DD shows reduced PAS formation and does not support phagophore expansion, showing that the EAT domain has an essential function that is separate from its Atg13-dependent role in autophagy initiation.

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Gopaldass, Navin, Bruno Fauvet, Hilal Lashuel, Aurélien Roux, and Andreas Mayer. "Membrane scission driven by the PROPPIN Atg18." EMBO Journal 36, no. 22 (October 13, 2017): 3274–91. http://dx.doi.org/10.15252/embj.201796859.

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van Zutphen, Tim, Virginia Todde, Rinse de Boer, Martin Kreim, Harald F. Hofbauer, Heimo Wolinski, Marten Veenhuis, Ida J. van der Klei, and Sepp D. Kohlwein. "Lipid droplet autophagy in the yeast Saccharomyces cerevisiae." Molecular Biology of the Cell 25, no. 2 (January 15, 2014): 290–301. http://dx.doi.org/10.1091/mbc.e13-08-0448.

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Cytosolic lipid droplets (LDs) are ubiquitous organelles in prokaryotes and eukaryotes that play a key role in cellular and organismal lipid homeostasis. Triacylglycerols (TAGs) and steryl esters, which are stored in LDs, are typically mobilized in growing cells or upon hormonal stimulation by LD-associated lipases and steryl ester hydrolases. Here we show that in the yeast Saccharomyces cerevisiae, LDs can also be turned over in vacuoles/lysosomes by a process that morphologically resembles microautophagy. A distinct set of proteins involved in LD autophagy is identified, which includes the core autophagic machinery but not Atg11 or Atg20. Thus LD autophagy is distinct from endoplasmic reticulum–autophagy, pexophagy, or mitophagy, despite the close association between these organelles. Atg15 is responsible for TAG breakdown in vacuoles and is required to support growth when de novo fatty acid synthesis is compromised. Furthermore, none of the core autophagy proteins, including Atg1 and Atg8, is required for LD formation in yeast.

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Suzuki, Sho W., Hayashi Yamamoto, Yu Oikawa, Chika Kondo-Kakuta, Yayoi Kimura, Hisashi Hirano, and Yoshinori Ohsumi. "Atg13 HORMA domain recruits Atg9 vesicles during autophagosome formation." Proceedings of the National Academy of Sciences 112, no. 11 (March 3, 2015): 3350–55. http://dx.doi.org/10.1073/pnas.1421092112.

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During autophagosome formation, autophagosome-related (Atg) proteins are recruited hierarchically to organize the preautophagosomal structure (PAS). Atg13, which plays a central role in the initial step of PAS formation, consists of two structural regions, the N-terminal HORMA (from Hop1, Rev7, and Mad2) domain and the C-terminal disordered region. The C-terminal disordered region of Atg13, which contains the binding sites for Atg1 and Atg17, is essential for the initiation step in which the Atg1 complex is formed to serve as a scaffold for the PAS. The N-terminal HORMA domain of Atg13 is also essential for autophagy, but its molecular function has not been established. In this study, we searched for interaction partners of the Atg13 HORMA domain and found that it binds Atg9, a multispanning membrane protein that exists on specific cytoplasmic vesicles (Atg9 vesicles). After the Atg1 complex is formed, Atg9 vesicles are recruited to the PAS and become part of the autophagosomal membrane. HORMA domain mutants, which are unable to interact with Atg9, impaired the PAS localization of Atg9 vesicles and exhibited severe defects in starvation-induced autophagy. Thus, Atg9 vesicles are recruited to the PAS via the interaction with the Atg13 HORMA domain. Based on these findings, we propose that the two distinct regions of Atg13 play crucial roles in distinct steps of autophagosome formation: In the first step, Atg13 forms a scaffold for the PAS via its C-terminal disordered region, and subsequently it recruits Atg9 vesicles via its N-terminal HORMA domain.

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Short, Ben. "Phosphorylation helps Atg18 get the vacuole in shape." Journal of Cell Biology 202, no. 4 (August 12, 2013): 600. http://dx.doi.org/10.1083/jcb.2024iti3.

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Mesquita, Ana, Luis C. Tábara, Oscar Martinez-Costa, Natalia Santos-Rodrigo, Olivier Vincent, and Ricardo Escalante. "Dissecting the function of Atg1 complex in Dictyostelium autophagy reveals a connection with the pentose phosphate pathway enzyme transketolase." Open Biology 5, no. 8 (August 2015): 150088. http://dx.doi.org/10.1098/rsob.150088.

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The network of protein–protein interactions of the Dictyostelium discoideum autophagy pathway was investigated by yeast two-hybrid screening of the conserved autophagic proteins Atg1 and Atg8. These analyses confirmed expected interactions described in other organisms and also identified novel interactors that highlight the complexity of autophagy regulation. The Atg1 kinase complex, an essential regulator of autophagy, was investigated in detail here. The composition of the Atg1 complex in D. discoideum is more similar to mammalian cells than to Saccharomyces cerevisiae as, besides Atg13, it contains Atg101, a protein not conserved in this yeast. We found that Atg101 interacts with Atg13 and genetic disruption of these proteins in Dictyostelium leads to an early block in autophagy, although the severity of the developmental phenotype and the degree of autophagic block is higher in Atg13-deficient cells. We have also identified a protein containing zinc-finger B-box and FNIP motifs that interacts with Atg101. Disruption of this protein increases autophagic flux, suggesting that it functions as a negative regulator of Atg101. We also describe the interaction of Atg1 kinase with the pentose phosphate pathway enzyme transketolase (TKT). We found changes in the activity of endogenous TKT activity in strains lacking or overexpressing Atg1, suggesting the presence of an unsuspected regulatory pathway between autophagy and the pentose phosphate pathway in Dictyostelium that seems to be conserved in mammalian cells.

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Koefinger, Juergen, Michael J. Ragusa, Gerhard Hummer, and James H. Hurley. "Autophagy: Solution Structure of the Atg17-Atg29-Atg31-Atg1-Atg13 Complex." Biophysical Journal 108, no. 2 (January 2015): 343a. http://dx.doi.org/10.1016/j.bpj.2014.11.1882.

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Baskaran, Sulochanadevi, Michael J. Ragusa, and James H. Hurley. "How Atg18 and the WIPIs sense phosphatidylinositol 3-phosphate." Autophagy 8, no. 12 (December 2012): 1851–52. http://dx.doi.org/10.4161/auto.22077.

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Stjepanovic, Goran, and James H. Hurley. "Architecture and Dynamics of the Autophagic ATG2-ATG18 Complex." Biophysical Journal 114, no. 3 (February 2018): 426a. http://dx.doi.org/10.1016/j.bpj.2017.11.2360.

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Kawamata, Tomoko, Yoshiaki Kamada, Yukiko Kabeya, Takayuki Sekito, and Yoshinori Ohsumi. "Organization of the Pre-autophagosomal Structure Responsible for Autophagosome Formation." Molecular Biology of the Cell 19, no. 5 (May 2008): 2039–50. http://dx.doi.org/10.1091/mbc.e07-10-1048.

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Autophagy induced by nutrient depletion is involved in survival during starvation conditions. In addition to starvation-induced autophagy, the yeast Saccharomyces cerevisiae also has a constitutive autophagy-like system, the Cvt pathway. Among 31 autophagy-related (Atg) proteins, the function of Atg17, Atg29, and Atg31 is required specifically for autophagy. In this study, we investigated the role of autophagy-specific (i.e., non-Cvt) proteins under autophagy-inducing conditions. For this purpose, we used atg11Δ cells in which the Cvt pathway is abrogated. The autophagy-unique proteins are required for the localization of Atg proteins to the pre-autophagosomal structure (PAS), the putative site for autophagosome formation, under starvation condition. It is likely that these Atg proteins function as a ternary complex, because Atg29 and Atg31 bind to Atg17. The Atg1 kinase complex (Atg1–Atg13) is also essential for recruitment of Atg proteins to the PAS. The assembly of Atg proteins to the PAS is observed only under autophagy-inducing conditions, indicating that this structure is specifically involved in autophagosome formation. Our results suggest that Atg1 complex and the autophagy-unique Atg proteins cooperatively organize the PAS in response to starvation signals.

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Fischer, Sarah, Ramesh Rijal, Peter Frommolt, Prerana Wagle, Roman Konertz, Jan Faix, Susanne Meßling, and Ludwig Eichinger. "Functional Characterization of Ubiquitin-Like Core Autophagy Protein ATG12 in Dictyostelium discoideum." Cells 8, no. 1 (January 19, 2019): 72. http://dx.doi.org/10.3390/cells8010072.

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Autophagy is a highly conserved intracellular degradative pathway that is crucial for cellular homeostasis. During autophagy, the core autophagy protein ATG12 plays, together with ATG5 and ATG16, an essential role in the expansion of the autophagosomal membrane. In this study we analyzed gene replacement mutants of atg12 in Dictyostelium discoideum AX2 wild-type and ATG16‾ cells. RNAseq analysis revealed a strong enrichment of, firstly, autophagy genes among the up-regulated genes and, secondly, genes implicated in cell motility and phagocytosis among the down-regulated genes in the generated ATG12‾, ATG16‾ and ATG12‾/16‾ cells. The mutant strains showed similar defects in fruiting body formation, autolysosome maturation, and cellular viability, implying that ATG12 and ATG16 act as a functional unit in canonical autophagy. In contrast, ablation of ATG16 or of ATG12 and ATG16 resulted in slightly more severe defects in axenic growth, macropinocytosis, and protein homeostasis than ablation of only ATG12, suggesting that ATG16 fulfils an additional function in these processes. Phagocytosis of yeast, spore viability, and maximal cell density were much more affected in ATG12‾/16‾ cells, indicating that both proteins also have cellular functions independent of each other. In summary, we show that ATG12 and ATG16 fulfil autophagy-independent functions in addition to their role in canonical autophagy.

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Krick, Roswitha, Sandra Henke, Joern Tolstrup, and Michael Thumm. "Dissecting the localization and function of Atg18, Atg21 and Ygr223c." Autophagy 4, no. 7 (October 2008): 896–910. http://dx.doi.org/10.4161/auto.6801.

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Yorimitsu, Tomohiro, and Daniel J. Klionsky. "Atg11 Links Cargo to the Vesicle-forming Machinery in the Cytoplasm to Vacuole Targeting Pathway." Molecular Biology of the Cell 16, no. 4 (April 2005): 1593–605. http://dx.doi.org/10.1091/mbc.e04-11-1035.

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Proteins are selectively packaged into vesicles at specific sites and then delivered correctly to the various organelles where they function, which is critical to the proper physiology of each organelle. The precursor form of the vacuolar hydrolase aminopeptidase I is a selective cargo molecule of the cytoplasm to vacuole targeting (Cvt) pathway and autophagy. Precursor Ape1 along with its receptor Atg19 forms the Cvt complex, which is transported to the pre-autophagosomal structure (PAS), the putative site of Cvt vesicle formation, in a process dependent on Atg11. Here, we show that this interaction occurs through the Atg11 C terminus; subsequent recruitment of the Cvt complex to the PAS depends on central regions within Atg11. Atg11 was shown to physically link several proteins, although the timing of these interactions and their importance are unknown. Our mapping shows that the Atg11 coiled-coil domains are involved in self-assembly and the interaction with other proteins, including two previously unidentified partners, Atg17 and Atg20. Atg11 mutants defective in the transport of the Cvt complex to the PAS affect the localization of other Atg components, supporting the idea that the cargo facilitates the organization of the PAS in selective autophagy. These findings suggest that Atg11 plays an integral role in connecting cargo molecules with components of the vesicle-forming machinery.

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Memisoglu, Gonen, Vinay V. Eapen, Ying Yang, Daniel J. Klionsky, and James E. Haber. "PP2C phosphatases promote autophagy by dephosphorylation of the Atg1 complex." Proceedings of the National Academy of Sciences 116, no. 5 (January 17, 2019): 1613–20. http://dx.doi.org/10.1073/pnas.1817078116.

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Macroautophagy is orchestrated by the Atg1-Atg13 complex in budding yeast. Under nutrient-rich conditions, Atg13 is maintained in a hyperphosphorylated state by the TORC1 kinase. After nutrient starvation, Atg13 is dephosphorylated, triggering Atg1 kinase activity and macroautophagy induction. The phosphatases that dephosphorylate Atg13 remain uncharacterized. Here, we show that two redundant PP2C phosphatases, Ptc2 and Ptc3, regulate macroautophagy by dephosphorylating Atg13 and Atg1. In the absence of these phosphatases, starvation-induced macroautophagy and the cytoplasm-to-vacuole targeting pathway are inhibited, and the recruitment of the essential autophagy machinery to the phagophore assembly site is impaired. Expressing a genomic ATG13-8SA allele lacking key TORC1 phosphorylation sites partially bypasses the macroautophagy defect in ptc2Δ ptc3Δ strains. Moreover, Ptc2 and Ptc3 interact with the Atg1-Atg13 complex. Taken together, these results suggest that PP2C-type phosphatases promote macroautophagy by regulating the Atg1 complex.

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Lippai, Mónika, and Péter Lőw. "The Role of the Selective Adaptor p62 and Ubiquitin-Like Proteins in Autophagy." BioMed Research International 2014 (2014): 1–11. http://dx.doi.org/10.1155/2014/832704.

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The ubiquitin-proteasome system and autophagy were long viewed as independent, parallel degradation systems with no point of intersection. By now we know that these degradation pathways share certain substrates and regulatory molecules and show coordinated and compensatory function. Two ubiquitin-like protein conjugation pathways were discovered that are required for autophagosome biogenesis: the Atg12-Atg5-Atg16 and Atg8 systems. Autophagy has been considered to be essentially a nonselective process, but it turned out to be at least partially selective. Selective substrates of autophagy include damaged mitochondria, intracellular pathogens, and even a subset of cytosolic proteins with the help of ubiquitin-binding autophagic adaptors, such as p62/SQSTM1, NBR1, NDP52, and Optineurin. These proteins selectively recognize autophagic cargo and mediate its engulfment into autophagosomes by binding to the small ubiquitin-like modifiers that belong to the Atg8/LC3 family.

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Wang, Mengya, Jingjing Jing, Hao Li, Jingwei Liu, Yuan Yuan, and Liping Sun. "The expression characteristics and prognostic roles of autophagy-related genes in gastric cancer." PeerJ 9 (February 3, 2021): e10814. http://dx.doi.org/10.7717/peerj.10814.

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Background Autophagy is an evolutionally highly conserved process, accompanied by the dynamic changes of various molecules, which is necessary for the orderly degradation and recycling of cellular components. The aim of the study was to identify the role of autophagy-related (ATG) genes in the occurrence and development of gastric cancer (GC). Methods Data from Oncomine dataset was used for the differential expression analysis between cancer and normal tissues. The association of ATG genes expression with clinicopathologic indicators was evaluated by The Cancer Genome Atlas (TCGA) database and Gene Expression Omnibus (GEO) database. Moreover, using the TCGA datasets, the prognostic role of ATG genes was assessed. A nomogram was further built to assess the independent prognostic factors. Results The expression of autophagy-related genes AMBRA1, ATG4B, ATG7, ATG10, ATG12, ATG16L2, GABARAPL2, GABARAPL1, ULK4 and WIPI2 showed differences between cancer and normal tissues. After verification, ATG14 and ATG4D were significantly associated with TNM stage. ATG9A, ATG2A, and ATG4D were associated with T stage. VMP1 and ATG4A were low-expressed in patients without lymph node metastasis. No gene in autophagy pathway was associated with M stage. Further multivariate analysis suggested that ATG4D and MAP1LC3C were independent prognostic factors for GC. The C-index of nomogram was 0.676 and the 95% CI was 0.628 to 0.724. Conclusion Our study provided a comprehensive illustration of ATG genes expression characteristics in GC. Abnormal expressions of the ubiquitin-like conjugated system in ATG genes plays a key role in the occurrence of GC. ATG8/LC3 sub-system may play an important role in development and clinical outcome of GC. In the future, it is necessary to further elucidate the alterations of specific ATG8/LC3 forms in order to provide insights for the discovery, diagnosis, or targeting for GC.

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Kumar, Ravinder, Muhammad Arifur Rahman, and Taras Y. Nazarko. "Nitrogen Starvation and Stationary Phase Lipophagy Have Distinct Molecular Mechanisms." International Journal of Molecular Sciences 21, no. 23 (November 29, 2020): 9094. http://dx.doi.org/10.3390/ijms21239094.

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In yeast, the selective autophagy of intracellular lipid droplets (LDs) or lipophagy can be induced by either nitrogen (N) starvation or carbon limitation (e.g., in the stationary (S) phase). We developed the yeast, Komagataella phaffii (formerly Pichia pastoris), as a new lipophagy model and compared the N-starvation and S-phase lipophagy in over 30 autophagy-related mutants using the Erg6-GFP processing assay. Surprisingly, two lipophagy pathways had hardly overlapping stringent molecular requirements. While the N-starvation lipophagy strictly depended on the core autophagic machinery (Atg1-Atg9, Atg18, and Vps15), vacuole fusion machinery (Vam7 and Ypt7), and vacuolar proteolysis (proteinases A and B), only Atg6 and proteinases A and B were essential for the S-phase lipophagy. The rest of the proteins were only partially required in the S-phase. Moreover, we isolated the prl1 (for the positive regulator of lipophagy 1) mutant affected in the S-phase lipophagy, but not N-starvation lipophagy. The prl1 defect was at a stage of delivery of the LDs from the cytoplasm to the vacuole, further supporting the mechanistically different nature of the two lipophagy pathways. Taken together, our results suggest that N-starvation and S-phase lipophagy have distinct molecular mechanisms.

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Cheong, Heesun, Tomohiro Yorimitsu, Fulvio Reggiori, Julie E. Legakis, Chao-Wen Wang, and Daniel J. Klionsky. "Atg17 Regulates the Magnitude of the Autophagic Response." Molecular Biology of the Cell 16, no. 7 (July 2005): 3438–53. http://dx.doi.org/10.1091/mbc.e04-10-0894.

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Autophagy is a catabolic process used by eukaryotic cells for the degradation and recycling of cytosolic proteins and excess or defective organelles. In yeast, autophagy is primarily a response to nutrient limitation, whereas in higher eukaryotes it also plays a role in developmental processes. Due to its essentially unlimited degradative capacity, it is critical that regulatory mechanisms are in place to modulate the timing and magnitude of the autophagic response. One set of proteins that seems to function in this regard includes a complex that contains the Atg1 kinase. Aside from Atg1, the proteins in this complex participate primarily in either nonspecific autophagy or specific types of autophagy, including the cytoplasm to vacuole targeting pathway, which operates under vegetative growth conditions, and peroxisome degradation. Accordingly, these proteins are prime candidates for factors that regulate the conversion between these pathways, including the change in size of the sequestering vesicle, the most obvious morphological difference. The atg17Δ mutant forms a reduced number of small autophagosomes. As a result, it is defective in peroxisome degradation and is partially defective for autophagy. Atg17 interacts with both Atg1 and Atg13, via two coiled-coil domains, and these interactions facilitate its inclusion in the Atg1 complex.

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Rieter, E., F. Vinke, D. Bakula, E. Cebollero, C. Ungermann, T. Proikas-Cezanne, and F. Reggiori. "Atg18 function in autophagy is regulated by specific sites within its -propeller." Journal of Cell Science 126, no. 2 (December 10, 2012): 593–604. http://dx.doi.org/10.1242/jcs.115725.

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López-Martínez, Gema, Mar Margalef-Català, Francisco Salinas, Gianni Liti, and Ricardo Cordero-Otero. "ATG18 and FAB1 Are Involved in Dehydration Stress Tolerance in Saccharomyces cerevisiae." PLOS ONE 10, no. 3 (March 24, 2015): e0119606. http://dx.doi.org/10.1371/journal.pone.0119606.

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Nazarko, Taras Y., Jean-Claude Farré, and Suresh Subramani. "Peroxisome Size Provides Insights into the Function of Autophagy-related Proteins." Molecular Biology of the Cell 20, no. 17 (September 2009): 3828–39. http://dx.doi.org/10.1091/mbc.e09-03-0221.

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Autophagy is a major pathway of intracellular degradation mediated by formation of autophagosomes. Recently, autophagy was implicated in the degradation of intracellular bacteria, whose size often exceeds the capacity of normal autophagosomes. However, the adaptations of the autophagic machinery for sequestration of large cargos were unknown. Here we developed a yeast model system to study the effect of cargo size on the requirement of autophagy-related (Atg) proteins. We controlled the size of peroxisomes before their turnover by pexophagy, the selective autophagy of peroxisomes, and found that peroxisome size determines the requirement of Atg11 and Atg26. Small peroxisomes can be degraded without these proteins. However, Atg26 becomes essential for degradation of medium peroxisomes. Additionally, the pexophagy-specific phagophore assembly site, organized by the dual interaction of Atg30 with functionally active Atg11 and Atg17, becomes essential for degradation of large peroxisomes. In contrast, Atg28 is partially required for all autophagy-related pathways independent of cargo size, suggesting it is a component of the core autophagic machinery. As a rule, the larger the cargo, the more cargo-specific Atg proteins become essential for its sequestration.

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Araki, Yasuhiro, Wei-Chi Ku, Manami Akioka, Alexander I. May, Yu Hayashi, Fumio Arisaka, Yasushi Ishihama, and Yoshinori Ohsumi. "Atg38 is required for autophagy-specific phosphatidylinositol 3-kinase complex integrity." Journal of Cell Biology 203, no. 2 (October 28, 2013): 299–313. http://dx.doi.org/10.1083/jcb.201304123.

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Autophagy is a conserved eukaryotic process of protein and organelle self-degradation within the vacuole/lysosome. Autophagy is characterized by the formation of an autophagosome, for which Vps34-dervied phosphatidylinositol 3-phosphate (PI3P) is essential. In yeast, Vps34 forms two distinct protein complexes: complex I, which functions in autophagy, and complex II, which is involved in protein sorting to the vacuole. Here we identify and characterize Atg38 as a stably associated subunit of complex I. In atg38Δ cells, autophagic activity was significantly reduced and PI3-kinase complex I dissociated into the Vps15–Vps34 and Atg14–Vps30 subcomplexes. We find that Atg38 physically interacted with Atg14 and Vps34 via its N terminus. Further biochemical analyses revealed that Atg38 homodimerizes through its C terminus and that this homodimer formation is indispensable for the integrity of complex I. These data suggest that the homodimer of Atg38 functions as a physical linkage between the Vps15–Vps34 and Atg14–Vps30 subcomplexes to facilitate complex I formation.

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Xu, Peng, Deena Damschroder, Mei Zhang, Karen A. Ryall, Paul N. Adler, Jeffrey J. Saucerman, Robert J. Wessells, and Zhen Yan. "Atg2, Atg9 and Atg18 in mitochondrial integrity, cardiac function and healthspan in Drosophila." Journal of Molecular and Cellular Cardiology 127 (February 2019): 116–24. http://dx.doi.org/10.1016/j.yjmcc.2018.12.006.

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Polson, Hannah E. J., Jane de Lartigue, Daniel J. Rigden, Marco Reedijk, Sylvie Urbé, Michael J. Clague, and Sharon A. Tooze. "Mammalian Atg18 (WIPI2) localizes to omegasome-anchored phagophores and positively regulates LC3 lipidation." Autophagy 6, no. 4 (May 16, 2010): 506–22. http://dx.doi.org/10.4161/auto.6.4.11863.

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Watanabe, Yasunori, Takafumi Kobayashi, Hayashi Yamamoto, Hisashi Hoshida, Rinji Akada, Fuyuhiko Inagaki, Yoshinori Ohsumi, and Nobuo N. Noda. "Structure-based Analyses Reveal Distinct Binding Sites for Atg2 and Phosphoinositides in Atg18." Journal of Biological Chemistry 287, no. 38 (July 31, 2012): 31681–90. http://dx.doi.org/10.1074/jbc.m112.397570.

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Kamada, Yoshiaki, Ken-ichi Yoshino, Chika Kondo, Tomoko Kawamata, Noriko Oshiro, Kazuyoshi Yonezawa, and Yoshinori Ohsumi. "Tor Directly Controls the Atg1 Kinase Complex To Regulate Autophagy." Molecular and Cellular Biology 30, no. 4 (December 7, 2009): 1049–58. http://dx.doi.org/10.1128/mcb.01344-09.

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ABSTRACT Autophagy is a bulk proteolytic process that is indispensable for cell survival during starvation. Autophagy is induced by nutrient deprivation via inactivation of the rapamycin-sensitive Tor complex1 (TORC1), a protein kinase complex regulating cell growth in response to nutrient conditions. However, the mechanism by which TORC1 controls autophagy and the direct target of TORC1 activity remain unclear. Atg13 is an essential regulatory component of autophagy upstream of the Atg1 kinase complex, and here we show that yeast TORC1 directly phosphorylates Atg13 at multiple Ser residues. Additionally, expression of an unphosphorylatable Atg13 mutant bypasses the TORC1 pathway to induce autophagy through activation of Atg1 in cells growing under nutrient-rich conditions. Our findings suggest that the direct control of the Atg1 complex by TORC1 induces autophagy.

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Sou, Yu-shin, Satoshi Waguri, Jun-ichi Iwata, Takashi Ueno, Tsutomu Fujimura, Taichi Hara, Naoki Sawada, et al. "The Atg8 Conjugation System Is Indispensable for Proper Development of Autophagic Isolation Membranes in Mice." Molecular Biology of the Cell 19, no. 11 (November 2008): 4762–75. http://dx.doi.org/10.1091/mbc.e08-03-0309.

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Autophagy is an evolutionarily conserved bulk-protein degradation pathway in which isolation membranes engulf the cytoplasmic constituents, and the resulting autophagosomes transport them to lysosomes. Two ubiquitin-like conjugation systems, termed Atg12 and Atg8 systems, are essential for autophagosomal formation. In addition to the pathophysiological roles of autophagy in mammals, recent mouse genetic studies have shown that the Atg8 system is predominantly under the control of the Atg12 system. To clarify the roles of the Atg8 system in mammalian autophagosome formation, we generated mice deficient in Atg3 gene encoding specific E2 enzyme for Atg8. Atg3-deficient mice were born but died within 1 d after birth. Conjugate formation of mammalian Atg8 homologues was completely defective in the mutant mice. Intriguingly, Atg12–Atg5 conjugation was markedly decreased in Atg3-deficient mice, and its dissociation from isolation membranes was significantly delayed. Furthermore, loss of Atg3 was associated with defective process of autophagosome formation, including the elongation and complete closure of the isolation membranes, resulting in malformation of the autophagosomes. The results indicate the essential role of the Atg8 system in the proper development of autophagic isolation membranes in mice.

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Hibshman, Jonathan D., Tess C. Leuthner, Chelsea Shoben, Danielle F. Mello, David R. Sherwood, Joel N. Meyer, and L. Ryan Baugh. "Nonselective autophagy reduces mitochondrial content during starvation in Caenorhabditis elegans." American Journal of Physiology-Cell Physiology 315, no. 6 (December 1, 2018): C781—C792. http://dx.doi.org/10.1152/ajpcell.00109.2018.

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Starvation significantly alters cellular physiology, and signs of aging have been reported to occur during starvation. Mitochondria are essential to the regulation of cellular energetics and aging. We sought to determine whether mitochondria exhibit signs of aging during starvation and whether quality control mechanisms regulate mitochondrial physiology during starvation. We describe effects of starvation on mitochondria in the first and third larval stages of the nematode Caenorhabditis elegans. When starved, C. elegans larvae enter developmental arrest. We observed fragmentation of the mitochondrial network, a reduction in mitochondrial DNA (mtDNA) copy number, and accumulation of DNA damage during starvation-induced developmental arrest. Mitochondrial function was also compromised by starvation. Starved worms had lower basal, maximal, and ATP-linked respiration. These observations are consistent with reduced mitochondrial quality, similar to mitochondrial phenotypes during aging. Using pharmacological and genetic approaches, we found that worms deficient for autophagy were short-lived during starvation and recovered poorly from extended starvation, indicating sensitivity to nutrient stress. Autophagy mutants unc-51/Atg1 and atg-18/Atg18 maintained greater mtDNA content than wild-type worms during starvation, suggesting that autophagy promotes mitochondrial degradation during starvation. unc-51 mutants also had a proportionally smaller reduction in oxygen consumption rate during starvation, suggesting that autophagy also contributes to reduced mitochondrial function. Surprisingly, mutations in genes involved in mitochondrial fission and fusion as well as selective mitophagy of damaged mitochondria did not affect mitochondrial content during starvation. Our results demonstrate the profound influence of starvation on mitochondrial physiology with organismal consequences, and they show that these physiological effects are influenced by autophagy.

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Karow, Malte, Sarah Fischer, Susanne Meßling, Roman Konertz, Jana Riehl, Qiuhong Xiong, Ramesh Rijal, Prerana Wagle, Christoph S. Clemen, and Ludwig Eichinger. "Functional Characterisation of the Autophagy ATG12~5/16 Complex in Dictyostelium discoideum." Cells 9, no. 5 (May 9, 2020): 1179. http://dx.doi.org/10.3390/cells9051179.

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Macroautophagy, a highly conserved and complex intracellular degradative pathway, involves more than 20 core autophagy (ATG) proteins, among them the hexameric ATG12~5/16 complex, which is part of the essential ubiquitin-like conjugation systems in autophagy. Dictyostelium discoideum atg5 single, atg5/12 double, and atg5/12/16 triple gene knock-out mutant strains displayed similar defects in the conjugation of ATG8 to phosphatidylethanolamine, development, and cell viability upon nitrogen starvation. This implies that ATG5, 12 and 16 act as a functional unit in canonical autophagy. Macropinocytosis of TRITC dextran and phagocytosis of yeast were significantly decreased in ATG5¯ and ATG5¯/12¯ and even further in ATG5¯/12¯/16¯ cells. In contrast, plaque growth on Klebsiella aerogenes was about twice as fast for ATG5¯ and ATG5¯/12¯/16¯ cells in comparison to AX2, but strongly decreased for ATG5¯/12¯ cells. Along this line, phagocytic uptake of Escherichia coli was significantly reduced in ATG5¯/12¯ cells, while no difference in uptake, but a strong increase in membrane association of E. coli, was seen for ATG5¯ and ATG5¯/12¯/16¯ cells. Proteasomal activity was also disturbed in a complex fashion, consistent with an inhibitory activity of ATG16 in the absence of ATG5 and/or ATG12. Our results confirm the essential function of the ATG12~5/16 complex in canonical autophagy, and furthermore are consistent with autophagy-independent functions of the complex and its individual components. They also strongly support the placement of autophagy upstream of the ubiquitin-proteasome system (UPS), as a fully functional UPS depends on autophagy.

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Li, Yi-Ning, Ji-An Hu, and Hui-Ming Wang. "Inhibition of HIF-1αAffects Autophagy Mediated Glycosylation in Oral Squamous Cell Carcinoma Cells." Disease Markers 2015 (2015): 1–9. http://dx.doi.org/10.1155/2015/239479.

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Abstract:

Purpose. To validate the function of autophagy with the regulation of hypoxia inhibitor-induced glycosylation in oral squamous cell carcinoma cell.Methods. Human Tca8113 cell line was used to detect autophagy and glycosylation related protein expression by western blotting and immunofluorescence with HIF-1αinhibitor. Short interfering RNA (siRNA) transfection blocked human ATG12 and ATG1.Results. HIF-1αinhibitor PX-478 reduced the amount of LC3-II and LC3-I in Tca8113 cells. PX-478 decreased the expression of O-GlcNAc and OGT and increased OGA expression. The tendency of O-GlcNAc showed a similar pattern to OGT. PX-478 gradually decreased OGT expression in Tca8113 cells. Protein level of O-GlcNAc and OGT increased in ATG12 and ATG1 depletion. The expression of OGT decreased at first and then rose slowly with the treatment of Atg12 and Atg1 siRNA and PX-478 fluctuant. Autophagy affected the stability of OGT when HIF-1αsignaling was blocked.Conclusions. Autophagy reduced by hypoxic stress inhibited. HIF-1αinhibitor decreased glycosylation. OGT became unstable in the absence of autophagy when HIF-1αsignaling was blocked.

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Fujita, Naonobu, Takashi Itoh, Hiroko Omori, Mitsunori Fukuda, Takeshi Noda, and Tamotsu Yoshimori. "The Atg16L Complex Specifies the Site of LC3 Lipidation for Membrane Biogenesis in Autophagy." Molecular Biology of the Cell 19, no. 5 (May 2008): 2092–100. http://dx.doi.org/10.1091/mbc.e07-12-1257.

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Two ubiquitin-like molecules, Atg12 and LC3/Atg8, are involved in autophagosome biogenesis. Atg12 is conjugated to Atg5 and forms an ∼800-kDa protein complex with Atg16L (referred to as Atg16L complex). LC3/Atg8 is conjugated to phosphatidylethanolamine and is associated with autophagosome formation, perhaps by enabling membrane elongation. Although the Atg16L complex is required for efficient LC3 lipidation, its role is unknown. Here, we show that overexpression of Atg12 or Atg16L inhibits autophagosome formation. Mechanistically, the site of LC3 lipidation is determined by the membrane localization of the Atg16L complex as well as the interaction of Atg12 with Atg3, the E2 enzyme for the LC3 lipidation process. Forced localization of Atg16L to the plasma membrane enabled ectopic LC3 lipidation at that site. We propose that the Atg16L complex is a new type of E3-like enzyme that functions as a scaffold for LC3 lipidation by dynamically localizing to the putative source membranes for autophagosome formation.

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Nguyen, Hoa Mai, Shuxian Liu, Wassim Daher, Feng Tan, and Sébastien Besteiro. "Characterisation of two Toxoplasma PROPPINs homologous to Atg18/WIPI suggests they have evolved distinct specialised functions." PLOS ONE 13, no. 4 (April 16, 2018): e0195921. http://dx.doi.org/10.1371/journal.pone.0195921.

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Oku, Masahide, Naoki Tamura, and Yasuyoshi Sakai. "Atg18 lifts up from and lands on the vacuolar membrane mediated by phosphorylation of its propellers." Autophagy 9, no. 12 (December 5, 2013): 2161–62. http://dx.doi.org/10.4161/auto.26379.

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Journal articles on the topic 'Atg18'

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