bims-auttor 2019-09-01 papers

Issue of 2019–09–01
six papers selected by
Viktor Korolchuk, Newcastle University

Cells. 2019 Aug 25. pii: E973. [Epub ahead of print]8(9):

Mechanisms and Pathophysiological Roles of the ATG8 Conjugation Machinery.

Since their initial discovery around two decades ago, the yeast autophagy-related (Atg)8 protein and its mammalian homologues of the light chain 3 (LC3) and γ-aminobutyric acid receptor associated proteins (GABARAP) families have been key for the tremendous expansion of our knowledge about autophagy, a process in which cytoplasmic material become targeted for lysosomal degradation. These proteins are ubiquitin-like proteins that become directly conjugated to a lipid in the autophagy membrane upon induction of autophagy, thus providing a marker of the pathway, allowing studies of autophagosome biogenesis and maturation. Moreover, the ATG8 proteins function to recruit components of the core autophagy machinery as well as cargo for selective degradation. Importantly, comprehensive structural and biochemical in vitro studies of the machinery required for ATG8 protein lipidation, as well as their genetic manipulation in various model organisms, have provided novel insight into the molecular mechanisms and pathophysiological roles of the mATG8 proteins. Recently, it has become evident that the ATG8 proteins and their conjugation machinery are also involved in intracellular pathways and processes not related to autophagy. This review focuses on the molecular functions of ATG8 proteins and their conjugation machinery in autophagy and other pathways, as well as their links to disease.

Keywords: ATG16L1; ATG5; ATG7; ATG8; GABARAP; LAP; LC3; autophagy

DOI: https://doi.org/10.3390/cells8090973

J Mol Biol. 2019 Aug 23. pii: S0022-2836(19)30519-4. [Epub ahead of print]

Repair or Lysophagy: Dealing with Damaged Lysosomes.

Chrisovalantis Papadopoulos, Bojana Kravic, Hemmo Meyer.

Lysosomal membrane permeabilization or full rupture of lysosomes is a common and severe stress condition that is relevant for degenerative disease, infection and cancer. If damage is limited, cells can repair lysosomes by means of the endosomal sorting complex required for transport (ESCRT) machinery. Presumably if repair fails, lysosomes are tagged with ubiquitin to initiate clearance by selective macroautophagy, termed lysophagy. Accumulating evidence suggests damage-induced exposure of luminal glycans to the cytosol as the key trigger for ubiquitination. In this review, we discuss recent data on cellular damage sensing, the underlying ubiquitination and autophagy machinery as well as additional layers of regulation such as processing of ubiquitinated proteins by the AAA-ATPase VCP/p97. We conclude with thoughts on how these mechanisms may regulate decision-making between lysosome repair and lysophagy.

DOI: https://doi.org/10.1016/j.jmb.2019.08.010

Nat Cell Biol. 2019 Aug 26.

Protein kinase N controls a lysosomal lipid switch to facilitate nutrient signalling via mTORC1.

Alexander Wallroth, Philipp A Koch, Andrea L Marat, Eberhard Krause, Volker Haucke.

Mechanistic target of rapamycin (mTOR) kinase functions in two multiprotein complexes: lysosomal mTOR complex 1 (mTORC1) and mTORC2 at the plasma membrane. mTORC1 modulates the cell response to growth factors and nutrients by increasing protein synthesis and cell growth, and repressing the autophagy-lysosomal pathway1-4; however, dysfunction in mTORC1 is implicated in various diseases3,5,6. mTORC1 activity is regulated by phosphoinositide lipids7-10. Class I phosphatidylinositol-3-kinase (PI3K)-mediated production of phosphatidylinositol-3,4,5-trisphosphate6,11 at the plasma membrane stimulates mTORC1 signalling, while local synthesis of phosphatidylinositol-3,4-bisphosphate by starvation-induced recruitment of class II PI3K-β (PI3KC2-β) to lysosomes represses mTORC1 activity12. How the localization and activity of PI3KC2-β are regulated by mitogens is unknown. We demonstrate that protein kinase N (PKN) facilitates mTORC1 signalling by repressing PI3KC2-β-mediated phosphatidylinositol-3,4-bisphosphate synthesis downstream of mTORC2. Active PKN2 phosphorylates PI3KC2-β to trigger PI3KC2-β complex formation with inhibitory 14-3-3 proteins. Conversely, loss of PKN2 or inactivation of its target phosphorylation site in PI3KC2-β represses nutrient signalling via mTORC1. These results uncover a mechanism that couples mTORC2-dependent activation of PKN2 to the regulation of mTORC1-mediated nutrient signalling by local lipid signals.

DOI: https://doi.org/10.1038/s41556-019-0377-3

Autophagy. 2019 Aug 26. 1-15

m6A mRNA methylation controls autophagy and adipogenesis by targeting Atg5 and Atg7.

Xinxia Wang, Ruifan Wu, Youhua Liu, Yuanling Zhao, Zhen Bi, Yongxi Yao, Qing Liu, Hailing Shi, Fengqin Wang, Yizhen Wang.

N6-methyladenosine (m6A), the most abundant internal modification on mRNAs in eukaryotes, play roles in adipogenesis. However, the underlying mechanism remains largely unclear. Here, we show that m6A plays a critical role in regulating macroautophagy/autophagy and adipogenesis through targeting Atg5 and Atg7. Mechanistically, knockdown of FTO, a well-known m6A demethylase, decreased the expression of ATG5 and ATG7, leading to attenuation of autophagosome formation, thereby inhibiting autophagy and adipogenesis. We proved that FTO directly targeted Atg5 and Atg7 transcripts and mediated their expression in an m6A-dependent manner. Further study identified that Atg5 and Atg7 were the targets of YTHDF2 (YTH N6-methyladenosine RNA binding protein 2). Upon FTO silencing, Atg5 and Atg7 transcripts with higher m6A levels were captured by YTHDF2, which resulted in mRNA degradation and reduction of protein expression, thus alleviating autophagy and adipogenesis. Furthermore, we generated an adipose-selective fto knockout mouse and find that FTO deficiency decreased white fat mass and impairs ATG5- and ATG7-dependent autophagy in vivo. Together, these findings unveil the functional importance of the m6A methylation machinery in autophagy and adipogenesis regulation, which expands our understanding of such interplay that is essential for development of therapeutic strategies in the prevention and treatment of obesity. Abbreviations: 3-MA: 3-methyladenine; ACTB: actin, beta; ATG: autophagy-related; Baf A1: bafilomycin A1; CEBPA: CCAAT/enhancer binding protein (C/EBP), alpha; CEBPB: CCAAT/enhancer binding protein (C/EBP), beta; FABP4: fatty acid binding protein 4, adipocyte; FTO: fat mass and obesity associated; HFD: high-fat diet; LC-MS/MS: liquid chromatography-tandem mass spectrometry; MAP1LC3B/LC3: microtubule-associated protein 1 light chain 3 beta; m6A: N6-methyladenosine; MEFs: mouse embryo fibroblasts; MeRIP-qPCR: methylated RNA immunoprecipitation-qPCR; PPARG: peroxisome proliferator activated receptor gamma; RIP: RNA-immunoprecipitation; SAT: subcutaneous adipose tissue; siRNA: small interfering RNA; SQSTM1/p62: sequestosome 1; TEM: transmission electron microscopy; ULK1: unc-51 like kinase 1; VAT: visceral adipose tissue; WAT: white adipose tissue; YTHDF: YTH N6-methyladenosine RNA binding protein.

Keywords: ATG5; ATG7; Adipogenesis; FTO; autophagy; mA

DOI: https://doi.org/10.1080/15548627.2019.1659617

EMBO Rep. 2019 Aug 26. e47734

Targeting of early endosomes by autophagy facilitates EGFR recycling and signalling.

Jane Fraser, Joanne Simpson, Rosa Fontana, Chieko Kishi-Itakura, Nicholas T Ktistakis, Noor Gammoh.

Despite recently uncovered connections between autophagy and the endocytic pathway, the role of autophagy in regulating endosomal function remains incompletely understood. Here, we find that the ablation of autophagy-essential players disrupts EGF-induced endocytic trafficking of EGFR. Cells lacking ATG7 or ATG16L1 exhibit increased levels of phosphatidylinositol-3-phosphate (PI(3)P), a key determinant of early endosome maturation. Increased PI(3)P levels are associated with an accumulation of EEA1-positive endosomes where EGFR trafficking is stalled. Aberrant early endosomes are recognised by the autophagy machinery in a TBK1- and Gal8-dependent manner and are delivered to LAMP2-positive lysosomes. Preventing this homeostatic regulation of early endosomes by autophagy reduces EGFR recycling to the plasma membrane and compromises downstream signalling and cell survival. Our findings uncover a novel role for the autophagy machinery in maintaining early endosome function and growth factor sensing.

Keywords: EGFR; autophagy; early endosomes; galectin; signalling

DOI: https://doi.org/10.15252/embr.201947734

Biochem Biophys Res Commun. 2019 Aug 21. pii: S0006-291X(19)31626-2. [Epub ahead of print]

UVA-induced photoaging inhibits autophagic degradation by impairing lysosomal function in dermal fibroblasts.

Yunfen Huang, Yuying Li, Yingying Qu, Yue Zheng, Mengting Ouyang, Yunqing Zhang, Wei Lai, Qingfang Xu.

Autophagy has been associated with a variety of diseases especially aging. Human dermal fibroblasts (HDFs) can internalize and then degrade elastin, collagen and advanced glycation end products (AGEs) in lysosomes, which plays prominent roles in extracellular matrix homeostasis and AGEs removal in the dermis. Although autophagy has been reported to be decreased in photoaged fibroblasts, the underlying mechanism and its relevance to photoaging remain elusive. Here, we showed that GFP-LC3 puncta per cell, LC3Ⅰ/Ⅱ conversion and p62 expression were significantly increased, whereas beclin1 expression was not altered in UVA-induced photoaged fibroblasts compared with non-photoaged control. Moreover, autophagic flux was not significantly affected by chloroquine treatment, but was remarkably induced by rapamycin treatment in photoaged fibroblasts, suggesting that UVA-induced photoaging might inhibit autophagy at the degradation stage. Further lysosomal function studies demonstrated that degradation of formed autophagosomes, LC3Ⅱprotein and DQ-Green BSA was all dramatically decreased in photoaged fibroblasts. LysoSensor yellow/blue DND 160 staining and flow cytometry assays demonstrated that photoaging obviously attenuated lysosomal acidification. Also, decreased expression of cathepsin B, L and D was found in photoaged fibroblasts. These data suggest that lowered lysosomal acidity and decreased cathepsins expression might contribute to the inhibition of autophagic degradation, which might be crucial in the development of photoaging through impairing intracellular degradation.

Keywords: Autophagy; Cathepsin; Dermal fibroblasts; Lysosome; Photoaging

DOI: https://doi.org/10.1016/j.bbrc.2019.08.103