bims-auttor 2020-11-22 papers

bims-auttor

Biomed News

on Autophagy and mTOR

Issue of 2020–11–22
forty papers selected by
Viktor Korolchuk, Newcastle University

A Selective Autophagy Pathway for Phase-Separated Endocytic Protein Deposits.
Lysosomal targeting of autophagosomes by the TECPR domain of TECPR2.
Alternative mitochondrial quality control mediated by extracellular release.
Fine-tuning ER-phagy by post-translational modifications.
STING induces LC3B lipidation onto single-membrane vesicles via the V-ATPase and ATG16L1-WD40 domain.
Ubiquitination of Intramitochondrial Proteins: Implications for Metabolic Adaptability.
MAPK1/3 kinase-dependent ULK1 degradation attenuates mitophagy and promotes breast cancer bone metastasis.
A SNARE protein Syntaxin 17 captures CFTR to potentiate autophagosomal clearance under stress.
Loss of TAX1BP1-Directed Autophagy Results in Protein Aggregate Accumulation in the Brain.
Interplay between reactive oxygen species and autophagy in the course of age-related macular degeneration.
Mitochondria-ER Tethering in Neurodegenerative Diseases.
NPM1 mutant maintains ULK1 protein stability via TRAF6-dependent ubiquitination to promote autophagic cell survival in leukemia.
Naa20, the catalytic subunit of NatB complex, contributes to hepatocellular carcinoma by regulating the LKB1-AMPK-mTOR axis.
Mitochondria-Associated Endoplasmic Reticulum Membranes in the Pathogenesis of Type 2 Diabetes Mellitus.
Replicative Senescence and Expression of Autophagy Genes in Mesenchymal Stromal Cells.
A tecpr2 knockout mouse exhibits age-dependent neuroaxonal dystrophyassociated with autophagosome accumulation.
p62-Nrf2-p62 Mitophagy Regulatory Loop as a Target for Preventive Therapy of Neurodegenerative Diseases.
Functions of outer mitochondrial membrane proteins: mediating the crosstalk between mitochondrial dynamics and mitophagy.
The Interaction of Aging and Cellular Stress Contributes to Pathogenesis in Mouse and Human Huntington Disease Neurons.
Calciprotein particle-induced cytotoxicity via lysosomal dysfunction and altered cholesterol distribution in renal epithelial HK-2 cells.
TXNIP/VDUP1 attenuates steatohepatitis via autophagy and fatty acid oxidation.
Regulation of cytokine signaling through direct interaction between cytokine receptors and the ATG16L1 WD40 domain.
Design, Synthesis, and Characterization of an Orally Active Dual-Specific ULK1/2 Autophagy Inhibitor that Synergizes with the PARP Inhibitor Olaparib for the Treatment of Triple-Negative Breast Cancer.
Malvidin-3-O-arabinoside ameliorates ethyl carbamate-induced oxidative damage by stimulating AMPK-mediated autophagy.
The Host Autophagy During Toxoplasma Infection.
Autophagy: a promising therapeutic target for improving mesenchymal stem cell biological functions.
Axon Injury-Induced Autophagy Activation Is Impaired in a C. elegans Model of Tauopathy.
The E3 ubiquitin ligase Peli1 regulates the metabolic actions of mTORC1 to suppress antitumor T cell responses.
Increased telomerase improves motor function and alpha-synuclein pathology in a transgenic mouse model of Parkinson's disease associated with enhanced autophagy.
Substrate stiffness differentially impacts autophagy of endothelial cells and smooth muscle cells.
Autophagy Stimulus-Dependent Role of the Small GTPase Ras2 in Peroxisome Degradation.
ABHD5 suppresses cancer cell anabolism through lipolysis-dependent activation of the AMPK/mTORC1 pathway.
Autophagy is induced in human keratinocytes during human papillomavirus 11 pseudovirion entry.
Induction of Lysosomal Membrane Permeabilization Is a Major Event of FTY720-Mediated Non-Apoptotic Cell Death in Human Glioma Cells.
Dual Targeting of v-ATPase and mTORC1 Signaling Disarms Multidrug-Resistant Cancers.
Signaling Network Centered on mTORC1 Dominates Mammalian Intestinal Stem Cell Ageing.
TRIB3 facilitates glioblastoma progression via restraining autophagy.
Inverse sex-based expression profiles of PTEN and Klotho in mice.
Ubiquitin signaling in neurodegenerative diseases: an autophagy and proteasome perspective.
All-trans-retinoic acid inhibits the malignant behaviors of hepatocarcinoma cells by regulating autophagy.

Mol Cell. 2020 Nov 05. pii: S1097-2765(20)30737-1. [Epub ahead of print]

A Selective Autophagy Pathway for Phase-Separated Endocytic Protein Deposits.

Florian Wilfling, Chia-Wei Lee, Philipp S Erdmann, Yumei Zheng, Dawafuti Sherpa, Stefan Jentsch, Boris Pfander, Brenda A Schulman, Wolfgang Baumeister.

  Autophagy eliminates cytoplasmic content selected by autophagy receptors, which link cargo to the membrane-bound autophagosomal ubiquitin-like protein Atg8/LC3. Here, we report a selective autophagy pathway for protein condensates formed by endocytic proteins in yeast. In this pathway, the endocytic protein Ede1 functions as a selective autophagy receptor. Distinct domains within Ede1 bind Atg8 and mediate phase separation into condensates. Both properties are necessary for an Ede1-dependent autophagy pathway for endocytic proteins, which differs from regular endocytosis and does not involve other known selective autophagy receptors but requires the core autophagy machinery. Cryo-electron tomography of Ede1-containing condensates, at the plasma membrane and in autophagic bodies, shows a phase-separated compartment at the beginning and end of the Ede1-mediated selective autophagy route. Our data suggest a model for autophagic degradation of macromolecular protein complexes by the action of intrinsic autophagy receptors.

Keywords:  Atg11; Atg8; Ede1; clathrin-mediated endocytosis; intrinsic autophagy receptor; liquid-liquid phase separation; selective autophagy

DOI:  https://doi.org/10.1016/j.molcel.2020.10.030
Autophagy. 2020 Nov 19.

Lysosomal targeting of autophagosomes by the TECPR domain of TECPR2.

Milana Fraiberg, Bat-Chen Tamim-Yecheskel, Kamilya Kokabi, Nemanja Subic, Gali Heimer, Franziska Eck, Karsten Nalbach, Christian Behrends, Bruria Ben-Zeev, Oren Shatz, Zvulun Elazar.

  TECPR2 (tectonin beta-propeller repeat containing 2) is a large, multi-domain protein comprised of an amino-terminal WD domain, a middle unstructured region and a carboxy-terminal TEPCR domain comprises of six TECPR repeats followed by a functional LIR motif. Human TECPR2 mutations are linked to spastic paraplegia type 49 (SPG49), a hereditary neurodegenerative disorder. Here we show that basal macroautophagic/autophagic flux is impaired in SPG49 patient fibroblasts in the form of accumulated autophagosomes. Ectopic expression of either full length TECPR2 or the TECPR domain rescued autophagy in patient fibroblasts in a LIR-dependent manner. Moreover, this domain is recruited to the cytosolic leaflet of autophagosomal and lysosomal membranes in a LIR- and VAMP8-dependent manner, respectively. These findings provide evidence for a new role of the TECPR domain in particular, and TECPR2 in general, in lysosomal targeting of autophagosomes via association with Atg8-family proteins on autophagosomes and VAMP8 on lysosomes.

Keywords:  SPG49; TECPR2; autophagy; lysosome; neurodegeneration

DOI:  https://doi.org/10.1080/15548627.2020.1852727
Autophagy. 2020 Nov 20.

Alternative mitochondrial quality control mediated by extracellular release.

Chi-Jing Choong, Tatsusada Okuno, Kensuke Ikenaka, Kousuke Baba, Hideki Hayakawa, Masato Koike, Mutsumi Yokota, Junko Doi, Keita Kakuda, Toshihide Takeuchi, Akiko Kuma, Shuhei Nakamura, Yoshitaka Nagai, Seiichi Nagano, Tamotsu Yoshimori, Hideki Mochizuki.

  Mitochondrial quality control, which is crucial for maintaining cellular homeostasis, has been considered to be achieved exclusively through mitophagy. Here we report an alternative mitochondrial quality control pathway mediated by extracellular mitochondria release. By performing time-lapse confocal imaging on a stable cell line with fluorescent-labeled mitochondria, we observed release of mitochondria from cells into the extracellular space. Correlative light-electron microscopy revealed that majority of the extracellular mitochondria are in free form and, on rare occasions, some are enclosed in membrane-surrounded vesicles. Rotenone- and carbonyl cyanide m-chlorophenylhydrazone-induced mitochondrial quality impairment promotes the extracellular release of depolarized mitochondria. Overexpression of PRKN (parkin RBR E3 ubiquitin protein ligase), which has a pivotal role in mitophagy regulation, suppresses the extracellular mitochondria release under basal and stress condition, whereas its knockdown exacerbates it. Correspondingly, overexpression of PRKN-independent mitophagy regulators, BNIP3 (BCL2 interacting protein 3) and BNIP3L/NIX (BCL2 interacting protein 3 like), suppress extracellular mitochondria release. Autophagy-deficient cell lines show elevated extracellular mitochondria release. These results imply that perturbation of mitophagy pathway prompts mitochondria expulsion. Presence of mitochondrial protein can also be detected in mouse sera. Sera of PRKN-deficient mice contain higher level of mitochondrial protein compared to that of wild-type mice. More importantly, fibroblasts and cerebrospinal fluid samples from Parkinson disease patients carrying loss-of-function PRKN mutations show increased extracellular mitochondria compared to control subjects, providing evidence in a clinical context. Taken together, our findings suggest that extracellular mitochondria release is a comparable yet distinct quality control pathway from conventional mitophagy.

Keywords:  Mitochondria; Parkinson disease; mitochondrial quality control; mitophagy; parkin

DOI:  https://doi.org/10.1080/15548627.2020.1848130
Bioessays. 2020 Nov 18. e2000212

Fine-tuning ER-phagy by post-translational modifications.

Mohamed A Eldeeb, Cornelia E Zorca, Mohamed A Ragheb, Fatma B Rashidi, Doaa S Salah El-Din.

  Autophagy functions in both selective and non-selective ways to maintain cellular homeostasis. Endoplasmic reticulum autophagy (ER-phagy) is a subclass of autophagy responsible for the degradation of the endoplasmic reticulum through selective encapsulation into autophagosomes. ER-phagy occurs both under physiological conditions and in response to stress cues, and plays a crucial role in maintaining the homeostatic control of the organelle. Although specific receptors that target parts of the ER membrane, as well as, internal proteins for lysosomal degradation have been identified, the molecular regulation of ER-phagy has been elusive. Recent work has uncovered novel regulators of ER-phagy that involve post-translational modifications of ER-resident proteins and functional cross-talk with other cellular processes. Herein, we discuss how morphology affects the function of the peripheral ER, and how ER-phagy modulates the turnover of this organelle. We also address how ER-phagy is regulated at the molecular level, considering implications relevant to human diseases.

Keywords:  ER-phagy; N-degron; UFMylation; autophagy; endoplasmic reticulum; lysosome; protein degradation; proteolysis

DOI:  https://doi.org/10.1002/bies.202000212
J Cell Biol. 2020 Dec 07. pii: e202009128. [Epub ahead of print]219(12):

STING induces LC3B lipidation onto single-membrane vesicles via the V-ATPase and ATG16L1-WD40 domain.

Tara D Fischer, Chunxin Wang, Benjamin S Padman, Michael Lazarou, Richard J Youle.

Following the detection of cytosolic double-stranded DNA from viral or bacterial infection in mammalian cells, cyclic dinucleotide activation of STING induces interferon β expression to initiate innate immune defenses. STING activation also induces LC3B lipidation, a classical but equivocal marker of autophagy, that promotes a cell-autonomous antiviral response that arose before evolution of the interferon pathway. We report that STING activation induces LC3B lipidation onto single-membrane perinuclear vesicles mediated by ATG16L1 via its WD40 domain, bypassing the requirement of canonical upstream autophagy machinery. This process is blocked by bafilomycin A1 that binds and inhibits the vacuolar ATPase (V-ATPase) and by SopF, a bacterial effector that catalytically modifies the V-ATPase to inhibit LC3B lipidation via ATG16L1. These results indicate that activation of the cGAS-STING pathway induces V-ATPase-dependent LC3B lipidation that may mediate cell-autonomous host defense, an unanticipated mechanism that is distinct from LC3B lipidation onto double-membrane autophagosomes.

DOI: https://doi.org/10.1083/jcb.202009128
Biomolecules. 2020 Nov 16. pii: E1559. [Epub ahead of print]10(11):

Ubiquitination of Intramitochondrial Proteins: Implications for Metabolic Adaptability.

Prasad Sulkshane, Jonathan Ram, Michael H Glickman.

  Mitochondria are constantly subjected to stressful conditions due to their unique physiology and organization. The resulting damage leads to mitochondrial dysfunction, which underlies many pathophysiological conditions. Hence, constant surveillance is required to closely monitor mitochondrial health for sound maintenance of cellular metabolism and thus, for viability. In addition to internal mitochondrial chaperones and proteases, mitochondrial health is also governed by host cell protein quality control systems. The ubiquitin-proteasome system (UPS) and autophagy constitute the main pathways for removal of damaged or superfluous proteins in the cytosol, nucleus, and from certain organelles such as the Endoplasmic Reticulum (ER) and mitochondria. Although stress-induced ubiquitin-dependent degradation of mitochondrial outer membrane proteins has been widely studied, mechanisms of intramitochondrial protein ubiquitination has remained largely elusive due to the predominantly cytosolic nature of UPS components, separated from internal mitochondrial proteins by a double membrane. However, recent research has illuminated examples of intramitochondrial protein ubiquitination pathways and highlighted their importance under basal and stressful conditions. Owing to the dependence of mitochondria on the error-prone process of protein import from the cytosol, it is imperative that the cell eliminate any accumulated proteins in the event of mitochondrial protein import deficiency. Apparently, a significant portion of this activity involves ubiquitination in one way or another. In the present review article, following a brief introduction to mitochondrial protein quality control mechanisms, we discuss our recent understanding of intramitochondrial protein ubiquitination, its importance for basal function of mitochondria, metabolic implications, and possible therapeutic applications.

Keywords:  autophagy; metabolism; mitochondria; mitophagy; proteasome; protein import; protein quality control; proteolysis; ubiquitin

DOI:  https://doi.org/10.3390/biom10111559
Autophagy. 2020 Nov 19.

MAPK1/3 kinase-dependent ULK1 degradation attenuates mitophagy and promotes breast cancer bone metastasis.

Rong Deng, Hai-Liang Zhang, Jun-Hao Huang, Rui-Zhao Cai, Yan Wang, Yu-Hong Chen, Bing-Xin Hu, Zhi-Peng Ye, Zhi-Ling Li, Jia Mai, Yun Huang, Xuan Li, Xiao-Dan Peng, Gong-Kan Feng, Jun-Dong Li, Jun Tang, Xiao-Feng Zhu.

  The function of mitophagy in cancer is controversial. ULK1 is critical for induction of macroautophagy/autophagy and has a more specific role in mitophagy in response to hypoxia. Here, we show that ULK1 deficiency induces an invasive phenotype of breast cancer cells under hypoxia and increases osteolytic bone metastasis. Mechanistically, ULK1 depletion attenuates mitophagy ability during hypoxia. As a result, the accumulation of damaged, ROS-generating mitochondria leads to activation of the NLRP3 inflammasome, which induces abnormal soluble cytokines secretion, then promotes the differentiation and maturation of osteoclasts, and ultimately results in bone metastasis. Notably, phosphorylation of ULK1 by MAPK1/ERK2-MAPK3/ERK1 kinase triggers its interaction with BTRC and subsequent K48-linked ubiquitination and proteasome degradation. Also, a clearly negative correlation between the expression levels of ULK1 and p-MAPK1/3 was observed in human breast cancer tissues. The MAP2K/MEK inhibitor trametinib is sufficient to restore mitophagy function via upregulation of ULK1, leading to inhibition of NLRP3 inflammasome activation, thereby reduces bone metastasis. These results indicate that ULK1 knockout-mediated mitophagy defect promotes breast cancer bone metastasis and provide evidence to explore MAP2K/MEK- MAPK1/3 pathway inhibitors for therapy, especially in cancers displaying low levels of ULK1.

Keywords:  MAPK1/3 kinase; NLRP3 inflammasome; ULK1; bone metastasis; breast cancer; mitophagy deficiency

DOI:  https://doi.org/10.1080/15548627.2020.1850609
FASEB J. 2020 Nov 15.

A SNARE protein Syntaxin 17 captures CFTR to potentiate autophagosomal clearance under stress.

Kavisha Arora, Pramodha Liyanage, Qing Zhong, Anjaparavanda P Naren.

  Autophagy, a cellular stress response to starvation and bacterial infection, is executed by double-membrane-bound organelles called autophagosomes. Autophagosomes transfer cytosolic material to acidified lysosomes for degradation following soluble N-ethylmaleimide-sensitive factor attachment receptor (SNARE)-dependent fusion processes. Many of the autophagy-related disorders stem from defective end-step proteolysis inside lysosomes. The role of epithelial cystic fibrosis (CF) transmembrane conductance regulator (CFTR) chloride channel has been argued to be critical for efficient lysosomal clearance; however, its context to autophagic clearance and the underlying mechanism is poorly defined. Here, we report that syntaxin17 (Stx17), an autophagic SNARE protein interacts with CFTR under nutritional stress and bacterial infection and incorporates it into mature autophagosomes to mediate an efficient lysosomal clearance. Lack of CFTR function and Stx17 and loss of CFTR-Stx17 interaction impairs bacterial clearance. We discover a specialized role of the Stx17-CFTR protein complex that is critical to prevent defective autophagy as has been the reported scenario in CF airway epithelial cells, infectious diseases, and lysosomal clearance disorders.

Keywords:  SNARE proteins; autophagy; cystic fibrosis; cystic fibrosis transmembrane conductance regulator; lysosomal clearance

DOI:  https://doi.org/10.1096/fj.201903210R
Mol Cell. 2020 Nov 12. pii: S1097-2765(20)30775-9. [Epub ahead of print]

Loss of TAX1BP1-Directed Autophagy Results in Protein Aggregate Accumulation in the Brain.

Shireen A Sarraf, Hetal V Shah, Gil Kanfer, Alicia M Pickrell, Lynne A Holtzclaw, Michael E Ward, Richard J Youle.

  Protein aggregates disrupt cellular homeostasis, causing toxicity linked to neurodegeneration. Selective autophagic elimination of aggregates is critical to protein quality control, but how aggregates are selectively targeted for degradation is unclear. We compared the requirements for autophagy receptor proteins: OPTN, NBR1, p62, NDP52, and TAX1BP1 in clearance of proteotoxic aggregates. Endogenous TAX1BP1 is recruited to and required for the clearance of stress-induced aggregates, whereas ectopic expression of TAX1BP1 increases clearance through autophagy, promoting viability of human induced pluripotent stem cell-derived neurons. In contrast, TAX1BP1 depletion sensitizes cells to several forms of aggregate-induced proteotoxicity. Furthermore, TAX1BP1 is more specifically expressed in the brain compared to other autophagy receptor proteins. In vivo, loss of TAX1BP1 results in accumulation of high molecular weight ubiquitin conjugates and premature lipofuscin accumulation in brains of young TAX1BP1 knockout mice. TAX1BP1 mediates clearance of a broad range of cytotoxic proteins indicating therapeutic potential in neurodegenerative diseases.

Keywords:  Huntington’s disease; OPTN; aggregate; aggrephagy; autophagy receptor; p62; proteostasis; proteotoxic stress; selective autophagy; ubiquitin

DOI:  https://doi.org/10.1016/j.molcel.2020.10.041
EXCLI J. 2020 ;19 1353-1371

Interplay between reactive oxygen species and autophagy in the course of age-related macular degeneration.

Malgorzata Nita, Andrzej Grzybowski.

  Pathological biomolecules such as lipofuscin, methylglyoxal-modified proteins (the major precursors of advanced glycationend products), misfolding protein deposits and dysfunctional mitochondria are source of oxidative stress and act as strong autophagic stimulators in age-related macular degeneration. Disturbed autophagy accelerates progression of the disease, since it leads to retinal cells' death and activates inflammation by the interplay with the NLRP3 inflammasome complex. Vascular dysfunction and hypoxia, as well as circulating autoantibodies against autophagy regulators (anti-S100A9, anti-ANXA5, and anti-HSPA8, A9 and B4) compromise an autophagy-mediated mechanism as well. Metformin, the autophagic stimulator, may act as a senostatic drug to inhibit the senescent phenotype in the age-related macular degeneration. PGC-1α , Sirt1 and AMPK represent new therapeutic targets for interventions in this disease.

Keywords:  age-related macular degeneration; autophagy; ophthalmology; oxidative stress; reactive oxygen species; retina disease

DOI:  https://doi.org/10.17179/excli2020-2915
Cell Mol Neurobiol. 2020 Nov 16.

Mitochondria-ER Tethering in Neurodegenerative Diseases.

Reza Raeisossadati, Merari F R Ferrari.

  Organelles juxtaposition has been detected for decades, although only recently gained importance due to a pivotal role in the regulation of cellular processes dependent on membrane contact sites. Endoplasmic reticulum (ER) and mitochondria interaction is a prime example of organelles contact sites. Mitochondria-associated membranes (MAM) are proposed to harbor ER-mitochondria tether complexes, mainly when these organelles are less than 30 nm apart. Dysfunctions of proteins located at the MAM are associated with neurodegenerative diseases such as Parkinson's, Alzheimer's and amyotrophic lateral sclerosis, as well as neurodevelopmental disorders; hence any malfunction in MAM can potentially trigger cell death. This review will focus on the role of ER-mitochondria contact sites, regarding calcium homeostasis, lipid metabolism, autophagy, morphology and dynamics of mitochondria, mainly in the context of neurodegenerative diseases. Approaches that have been employed so far to study organelles contact sites, as well as methods that were not used in neurosciences yet, but are promising and accurate ways to unveil the functions of MAM during neurodegeneration, is also discussed in the present review.

Keywords:  Autophagy; Calcium; Contact sites methodologies; Lipid metabolism; Mitochondria-associated membranes (MAM); Neurodegeneration

DOI:  https://doi.org/10.1007/s10571-020-01008-9
FASEB J. 2020 Nov 17.

NPM1 mutant maintains ULK1 protein stability via TRAF6-dependent ubiquitination to promote autophagic cell survival in leukemia.

Yuting Tang, Yao Tao, Lu Wang, Liyuan Yang, Yipei Jing, Xueke Jiang, Li Lei, Zailin Yang, Xin Wang, Meixi Peng, Qiaoling Xiao, Jun Ren, Ling Zhang.

  Nucleophosmin (NPM1) mutations are the most frequent genetic alteration in acute myeloid leukemia (AML) and aberrant cytoplasm-dislocated NPM1 mutant is a distinct biological characterization of this disease. Our group previously reported that NPM1 mutant elevated autophagy activity and autophagy activation contributed to leukemic cell survival. However, the molecular mechanisms by which cytoplasmic NPM1 mutant involving in the autophagy pathway has not been fully elucidated. Here, we showed that Unc-51-like kinase 1 (ULK1) as a core autophagy protein was highly expressed in NPM1-mA positive OCI-AML3 cells and primary NPM1-mutated AML blasts. Meanwhile, we found that NPM1-mA could interact with ULK1 protein and positively regulated ULK1 protein levels. Mechanically, NPM1-mA promoted TRAF6-dependent K63 ubiquitination and further maintained ULK1 stability and kinase activity via miR-146a. In addition, ULK1 high expression-mediated autophagy activation and facilitated to leukemic cell proliferation. Finally, we demonstrated that restoring ULK1 expression, ULK1 inhibitor SBI-0206965 treatment and using shULK1 partially rescued the effect of NPM1-mA on autophagy and cell survival. In conclusion, our findings suggest that NPM1 mutant interacts with ULK1, and thus, maintains its protein stability, which is required for NPM1 mutant-mediated autophagic cell survival. These data extend our understanding of the functions of NPM1 mutant in the regulation of autophagy activation in NPM1-mutated AML.

Keywords:  autophagy; gene mutation; kinase activity; nucleophosmin 1; unc51-like kinase 1

DOI:  https://doi.org/10.1096/fj.201903183RRR
Exp Mol Med. 2020 Nov 20.

Naa20, the catalytic subunit of NatB complex, contributes to hepatocellular carcinoma by regulating the LKB1-AMPK-mTOR axis.

Taek-Yeol Jung, Jae-Eun Ryu, Mi-Mi Jang, Soh-Yeon Lee, Gyu-Rin Jin, Chan-Woo Kim, Chae-Young Lee, Hyelee Kim, EungHan Kim, Sera Park, Seonjeong Lee, Cheolju Lee, Wankyu Kim, TaeSoo Kim, Soo-Young Lee, Bong-Gun Ju, Hyun-Seok Kim.

N-α-acetyltransferase 20 (Naa20), which is a catalytic subunit of the N-terminal acetyltransferase B (NatB) complex, has recently been reported to be implicated in hepatocellular carcinoma (HCC) progression and autophagy, but the underlying mechanism remains unclear. Here, we report that based on bioinformatic analysis of Gene Expression Omnibus and The Cancer Genome Atlas data sets, Naa20 expression is much higher in HCC tumors than in normal tissues, promoting oncogenic properties in HCC cells. Mechanistically, Naa20 inhibits the activity of AMP-activated protein kinase (AMPK) to promote the mammalian target of rapamycin signaling pathway, which contributes to cell proliferation, as well as autophagy, through its N-terminal acetyltransferase (NAT) activity. We further show that liver kinase B1 (LKB1), a major regulator of AMPK activity, can be N-terminally acetylated by NatB in vitro, but also probably by NatB and/or other members of the NAT family in vivo, which may have a negative effect on AMPK activity through downregulation of LKB1 phosphorylation at S428. Indeed, p-LKB1 (S428) and p-AMPK levels are enhanced in Naa20-deficient cells, as well as in cells expressing the nonacetylated LKB1-MPE mutant; moreover, importantly, LKB1 deficiency reverses the molecular and cellular events driven by Naa20 knockdown. Taken together, our findings suggest that N-terminal acetylation of LKB1 by Naa20 may inhibit the LKB1-AMPK signaling pathway, which contributes to tumorigenesis and autophagy in HCC.

DOI: https://doi.org/10.1038/s12276-020-00525-3
Front Cell Dev Biol. 2020 ;8 571554

Mitochondria-Associated Endoplasmic Reticulum Membranes in the Pathogenesis of Type 2 Diabetes Mellitus.

Shanshan Yang, Ruixue Zhou, Caixia Zhang, Siyuan He, Zhiguang Su.

  The endoplasmic reticulum (ER) and mitochondria are essential intracellular organelles that actively communicate via temporally and spatially formed contacts called mitochondria-associated membranes (MAMs). These mitochondria-ER contacts are not only necessary for the physiological function of the organelles and their coordination with each other, but they also control the intracellular lipid exchange, calcium signaling, cell survival, and homeostasis in cellular metabolism. Growing evidence strongly supports the role of the mitochondria-ER connection in the insulin resistance of peripheral tissues, pancreatic β cell dysfunction, and the consequent development of type 2 diabetes mellitus (T2DM). In this review, we summarize current advances in the understanding of the mitochondria-ER connection and specifically focus on addressing a new perspective of the alterations in mitochondria-ER communication in insulin signaling and β cell maintenance.

Keywords:  diabetes; endoplasmic reticulum; insulin resistance; mitochondria; mitochondria-associated endoplasmic reticulum membranes; pancreatic β cell

DOI:  https://doi.org/10.3389/fcell.2020.571554
Biochemistry (Mosc). 2020 Oct;85(10): 1169-1177

Replicative Senescence and Expression of Autophagy Genes in Mesenchymal Stromal Cells.

A Y Ratushnyy, Y V Rudimova, L B Buravkova.

Cell senescence leads to a number of changes in the properties of mesenchymal stromal cells (MSCs). In particular, the number of damaged structures is increased producing negative effect on intracellular processes. Elimination of the damaged molecules and organelles occurs via autophagy that can be important in the context of aging. Cultivation under low oxygen level can be used as an approach for enhancement of MSC therapeutic properties and "slowing down" cell senescence. The goal of this work was to study some morphological and functional characteristics and expression of autophagy-associated genes during replicative senescence of MSCs under different oxygen concentration. The study revealed changes in the regulation of autophagy at the transcriptional level. Upregulation of the expression of autophagosome membrane growth genes ATG9A and ULK1, of the autophagosome maturation genes CTSD, CLN3, GAA, and GABARAPL1, of the autophagy regulation genes TP53, TGFB1, BCL2L1, FADD, and HTT was shown. These changes were accompanied by downregulation of IGF1 and TGM2 expression. Increase of the lysosomal compartment volume was observed in the senescent MSCs that also indicated increase of their degradation activity. The number of lysosomes was decreased following prolonged cultivation under low oxygen concentration (5%). The replicative senescence of MSCs under conditions of different oxygen levels led to the similar modifications in the expression of the autophagy-associated genes.

DOI: https://doi.org/10.1134/S0006297920100053
Autophagy. 2020 Nov 20.

A tecpr2 knockout mouse exhibits age-dependent neuroaxonal dystrophyassociated with autophagosome accumulation.

Bat-Chen Tamim-Yecheskel, Milana Fraiberg, Kamilya Kokabi, Saskia Freud, Oren Shatz, Letizia Marvaldi, Nemanja Subic, Ori Brenner, Michael Tsoory, Raya Eilam-Altstadter, Inbal Biton, Alon Savidor, Nili Dezorella, Gali Heimer, Christian Behrends, Bruria Ben-Zeev, Zvulun Elazar.

  Mutations in the coding sequence of human TECPR2 were recently linked to spastic paraplegia type 49 (SPG49), a hereditary neurodegenerative disorder involving intellectual disability, autonomic-sensory neuropathy, chronic respiratory disease and decreased pain sensitivity. Here, we report the generation of a novel CRISPR-Cas9 tecpr2 knockout (tecpr2 -/-) mouse that exhibits behavioral pathologies observed in SPG49 patients. tecpr2 -/- mice develop neurodegenerative patterns in an age-dependent manner, manifested predominantly as neuroaxonal dystrophy in the gracile (GrN) and cuneate nuclei (CuN) of the medulla oblongata in the brainstem and dorsal white matter column of the spinal cord. Age-dependent correlation with accumulation of autophagosomes suggests compromised targeting to lysosome. Taken together, our findings establish the tecpr2 knockout mouse as a potential model for SPG49 and ascribe a new role to TECPR2 in macroautophagy/autophagy-related neurodegenerative disorders.

Keywords:  SPG49; TECPR2; autophagy; axonal dystrophy; lysosome; neurodegeneration

DOI:  https://doi.org/10.1080/15548627.2020.1852724
Brain Sci. 2020 Nov 12. pii: E847. [Epub ahead of print]10(11):

p62-Nrf2-p62 Mitophagy Regulatory Loop as a Target for Preventive Therapy of Neurodegenerative Diseases.

Artem P Gureev, Irina S Sadovnikova, Natalia N Starkov, Anatoly A Starkov, Vasily N Popov.

  Turnover of the mitochondrial pool due to coordinated processes of mitochondrial biogenesis and mitophagy is an important process in maintaining mitochondrial stability. An important role in this process is played by the Nrf2/ARE signaling pathway, which is involved in the regulation of the expression of genes responsible for oxidative stress protection, regulation of mitochondrial biogenesis, and mitophagy. The p62 protein is a multifunctional cytoplasmic protein that functions as a selective mitophagy receptor for the degradation of ubiquitinated substrates. There is evidence that p62 can positively regulate Nrf2 by binding to its negative regulator, Keap1. However, there is also strong evidence that Nrf2 up-regulates p62 expression. Thereby, a regulatory loop is formed between two important signaling pathways, which may be an important target for drugs aimed at treating neurodegeneration. Constitutive activation of p62 in parallel with Nrf2 would most likely result in the activation of mTORC1-mediated signaling pathways that are associated with the development of malignant neoplasms. The purpose of this review is to describe the p62-Nrf2-p62 regulatory loop and to evaluate its role in the regulation of mitophagy under various physiological conditions.

Keywords:  Nrf2; mitochondria; mitophagy; neurodegenerative disease; p62; regulatory loop

DOI:  https://doi.org/10.3390/brainsci10110847
Cell Death Differ. 2020 Nov 18.

Functions of outer mitochondrial membrane proteins: mediating the crosstalk between mitochondrial dynamics and mitophagy.

Hongxu Xian, Yih-Cherng Liou.

Most cellular stress responses converge on the mitochondria. Consequently, the mitochondria must rapidly respond to maintain cellular homeostasis and physiological demands by fine-tuning a plethora of mitochondria-associated processes. The outer mitochondrial membrane (OMM) proteins are central to mediating mitochondrial dynamics, coupled with continuous fission and fusion. These OMM proteins also have vital roles in controlling mitochondrial quality and serving as mitophagic receptors for autophagosome enclosure during mitophagy. Mitochondrial fission segregates impaired mitochondria in smaller sizes from the mother mitochondria and may favor mitophagy for eliminating damaged mitochondria. Conversely, mitochondrial fusion mixes dysfunctional mitochondria with healthy ones to repair the damage by diluting the impaired components and consequently prevents mitochondrial clearance via mitophagy. Despite extensive research efforts into deciphering the interplay between fission-fusion and mitophagy, it is still not clear whether mitochondrial fission essentially precedes mitophagy. In this review, we summarize recent breakthroughs concerning OMM research, and dissect the functions of these proteins in mitophagy from their traditional roles in fission-fusion dynamics, in response to distinct context, at the intersection of the OMM platform. These insights into the OMM proteins in mechanistic researches would lead to new aspects of mitochondrial quality control and better understanding of mitochondrial homeostasis intimately tied to pathological impacts.

DOI: https://doi.org/10.1038/s41418-020-00657-z
Front Aging Neurosci. 2020 ;12 524369

The Interaction of Aging and Cellular Stress Contributes to Pathogenesis in Mouse and Human Huntington Disease Neurons.

Emily Machiela, Ritika Jeloka, Nicholas S Caron, Shagun Mehta, Mandi E Schmidt, Helen J E Baddeley, Colton M Tom, Nalini Polturi, Yuanyun Xie, Virginia B Mattis, Michael R Hayden, Amber L Southwell.

  Huntington disease (HD) is a fatal, inherited neurodegenerative disorder caused by a mutation in the huntingtin (HTT) gene. While mutant HTT is present ubiquitously throughout life, HD onset typically occurs in mid-life. Oxidative damage accumulates in the aging brain and is a feature of HD. We sought to interrogate the roles and interaction of age and oxidative stress in HD using primary Hu97/18 mouse neurons, neurons differentiated from HD patient induced pluripotent stem cells (iPSCs), and the brains of HD mice. We find that primary neurons must be matured in culture for canonical stress responses to occur. Furthermore, when aging is accelerated in mature HD neurons, mutant HTT accumulates and sensitivity to oxidative stress is selectively enhanced. Furthermore, we observe HD-specific phenotypes in neurons and mouse brains that have undergone accelerated aging, including a selective increase in DNA damage. These findings suggest a role for aging in HD pathogenesis and an interaction between the biological age of HD neurons and sensitivity to exogenous stress.

Keywords:  DNA damage; Huntington (disease); iPSC (induced pluripotent stem cell); mouse models of disease; neurodegenaration; primary neuron culture

DOI:  https://doi.org/10.3389/fnagi.2020.524369
Sci Rep. 2020 Nov 18. 10(1): 20125

Calciprotein particle-induced cytotoxicity via lysosomal dysfunction and altered cholesterol distribution in renal epithelial HK-2 cells.

Rina Kunishige, Mai Mizoguchi, Asako Tsubouchi, Kenjiro Hanaoka, Yutaka Miura, Hiroshi Kurosu, Yasuteru Urano, Makoto Kuro-O, Masayuki Murata.

Dietary phosphate overload induces chronic kidney disease (CKD), and calciprotein particles (CPPs), a form of nanoparticle comprising calcium phosphate and serum proteins, has been proposed to cause renal toxicity. However, the mechanism of CPP cytotoxicity in renal tubular cells is unknown. Here we show that in renal proximal tubular epithelial HK-2 cells, endocytosed CPPs accumulate in late endosomes/lysosomes (LELs) and increase their luminal pH by ~ 1.0 unit. This results in a decrease in lysosomal hydrolase activity and autophagic flux blockage without lysosomal rupture and reactive oxygen species generation. CPP treatment led to vulnerability to H2O2-induced oxidative stress and plasma membrane injury, probably because of autophagic flux blockage and decreased plasma membrane cholesterol, respectively. CPP-induced disruption of lysosomal homeostasis, autophagy flux and plasma membrane integrity might trigger a vicious cycle, leading to progressive nephron loss.

DOI: https://doi.org/10.1038/s41598-020-77308-3
Autophagy. 2020 Nov 16. 1-16

TXNIP/VDUP1 attenuates steatohepatitis via autophagy and fatty acid oxidation.

Hee-Seon Park, Ji-Won Song, Jin-Ho Park, Byung-Kwan Lim, Og-Sung Moon, Hwa-Young Son, Jae-Hyuk Lee, Bin Gao, Young-Suk Won, Hyo-Jung Kwon.

  Impaired macroautophagy/autophagy has been implicated in experimental and human nonalcoholic steatohepatitis (NASH). However, the mechanism underlying autophagy dysregulation in NASH is largely unknown. Here, we investigated the role and mechanism of TXNIP/VDUP1 (thioredoxin interacting protein), a key mediator of cellular stress responses, in the pathogenesis of NASH. Hepatic TXNIP expression was upregulated in nonalcoholic fatty liver disease (NAFLD) patients and in methionine choline-deficient (MCD) diet-fed mice, as well as in palmitic acid (PA)-treated hepatocytes. Upregulation of hepatic TXNIP was positively correlated with impaired autophagy, as evidenced by a decreased number of MAP1LC3B/LC3B (microtubule-associated protein 1 light chain 3 beta) puncta and increased SQSTM1/p62 (sequestosome 1) expression. Deletion of the Txnip gene enhanced hepatic steatosis, inflammation, and fibrosis, accompanied by impaired autophagy and fatty acid oxidation (FAO) in MCD diet-fed mice. Mechanistically, TXNIP directly interacted with and positively regulated p-PRKAA, leading to inactivation of MTOR (mechanistic target of rapamycin kinase) complex 1 (MTORC1) and nuclear translocation of TFEB (transcription factor EB), which in turn promoted autophagy. Inhibition of MTORC1 by rapamycin induced autophagy and increased the expression levels of FAO-related genes and concomitantly attenuated lipid accumulation in PA-treated txnip-knockout (KO) hepatocytes, which was further abolished by silencing of Atg7. Rapamycin treatment also attenuated MCD diet-induced steatosis, inflammation, and fibrosis with increased TFEB nuclear translocation and restored FAO in txnip-KO mice. Our findings suggest that elevated TXNIP ameliorates steatohepatitis by interacting with PRKAA and thereby inducing autophagy and FAO. Targeting TXNIP may be a potential therapeutic approach for NASH. Abbreviations: ACOX1: acyl-Coenzyme A oxidase 1, palmitoyl; ACSL1: acyl-CoA synthetase long-chain family member 1; ACTA2/α-SMA: actin, alpha 2, smooth muscle, aorta; ACTB: actin beta; ADGRE1/F4/80: adhesion G protein-coupled receptor E1; AMPK: AMP-activated protein kinase; ATG: autophagy-related; BafA1: bafilomycin A1; COL1A1/Col1α1: collagen, type I, alpha 1; CPT1A: carnitine palmitoyltransferase 1a, liver; CQ: chloroquine; DGAT1: diacylglycerol O-acyltransferase 1; DGAT2: diacylglycerol O-acyltransferase 2; ECI2/Peci: enoyl-Coenzyme A isomerase 2; EHHADH: enoyl-Coenzyme A, hydratase/3-hydroxyacyl Coenzyme A dehydrogenase; FAO: fatty acid oxidation; FASN: fatty acid synthase; FFA: free fatty acids; GFP: green fluorescent protein; GK/GYK: glycerol kinase; GOT1/AST: glutamic-oxaloacetic transaminase 1, soluble; GPAM: glycerol-3-phosphate acyltransferase, mitochondrial; GPT/ALT: glutamic pyruvic transaminase, soluble; H&E: hematoxylin and eosin; IL1B/IL-1β: interleukin 1 beta; IL6: interleukin 6; IOD: integral optical density; KO: knockout; Leu: leupeptin; LPIN1: lipin 1; MAP1LC3B/LC3B: microtubule-associated protein 1 light chain 3 beta; MCD: methionine choline-deficient; MMP9: matrix metallopeptidase 9; mRNA: messenger RNA; MTORC1: mechanistic target of rapamycin kinase complex 1; NAFLD: nonalcoholic fatty liver diseases; NASH: nonalcoholic steatohepatitis; PA: palmitic acid; PPARA/PPARα: peroxisome proliferator activated receptor alpha; PPARG/PPARγ: peroxisome proliferator activated receptor gamma; qRT-PCR: quantitative real-time PCR; RPS6KB1/p70S6K1: ribosomal protein S6 kinase, polypeptide 1; RPTOR: regulatory associated protein of MTOR complex 1; SCD1: stearoyl-Coenzyme A desaturase 1; SEM: standard error of the mean; siRNA: small interfering RNA; SQSTM1/p62: sequestosome 1; TFEB: transcription factor EB; TG: triglyceride; TGFB/TGF-β: transforming growth factor, beta; TIMP1: tissue inhibitor of metalloproteinase 1; TNF/TNF-α: tumor necrosis factor; TXNIP/VDUP1: thioredoxin interacting protein; WT: wild-type.

Keywords:  Lipid metabolism; MAP1LC3B; NASH; PRKAA; SQSTM1; liver

DOI:  https://doi.org/10.1080/15548627.2020.1834711
Nat Commun. 2020 Nov 20. 11(1): 5919

Regulation of cytokine signaling through direct interaction between cytokine receptors and the ATG16L1 WD40 domain.

Inmaculada Serramito-Gómez, Emilio Boada-Romero, Raquel Villamuera, Álvaro Fernández-Cabrera, José Luis Cedillo, Ángela Martín-Regalado, Simon Carding, Uli Mayer, Penny P Powell, Thomas Wileman, Irene García-Higuera, Felipe X Pimentel-Muiños.

ATG16L1, an autophagy mediator that specifies the site of LC3 lipidation, includes a C-terminal domain formed by 7 WD40-type repeats (WD40 domain, WDD), the function of which is unclear. Here we show that the WDD interacts with the intracellular domain of cytokine receptors to regulate their signaling output in response to ligand stimulation. Using a refined version of a previously described WDD-binding amino acid motif, here we show that this element is present in the intracellular domain of cytokine receptors. Two of these receptors, IL-10RB and IL-2Rγ, recognize the WDD through the motif and exhibit WDD-dependent LC3 lipidation activity. IL-10 promotes IL-10RB/ATG16L1 interaction through the WDD, and IL-10 signaling is suboptimal in cells lacking the WDD owing to delayed endocytosis and inefficient early trafficking of IL10/IL-10R complexes. Our data reveal WDD-dependent roles of ATG16L1 in the regulation of cytokine receptor trafficking and signaling, and provide a WDD-binding motif that might be used to identify additional WDD activators.

DOI: https://doi.org/10.1038/s41467-020-19670-4
J Med Chem. 2020 Nov 17.

Design, Synthesis, and Characterization of an Orally Active Dual-Specific ULK1/2 Autophagy Inhibitor that Synergizes with the PARP Inhibitor Olaparib for the Treatment of Triple-Negative Breast Cancer.

Huiyu Ren, Nicole A Bakas, Mitchell Vamos, Apirat Chaikuad, Allison S Limpert, Carina D Wimer, Sonja N Brun, Lester J Lambert, Lutz Tautz, Maria Celeridad, Douglas J Sheffler, Stefan Knapp, Reuben J Shaw, Nicholas D P Cosford.

Inhibition of autophagy, the major cellular recycling pathway in mammalian cells, is a promising strategy for the treatment of triple-negative breast cancer (TNBC). We previously reported SBI-0206965, a small molecule inhibitor of unc-51-like autophagy activating kinase 1 (ULK1), which is a key regulator of autophagy initiation. Herein, we describe the design, synthesis, and characterization of new dual inhibitors of ULK1 and ULK2 (ULK1/2). One inhibitor, SBP-7455 (compound 26), displayed improved binding affinity for ULK1/2 compared with SBI-0206965, potently inhibited ULK1/2 enzymatic activity in vitro and in cells, reduced the viability of TNBC cells and had oral bioavailability in mice. SBP-7455 inhibited starvation-induced autophagic flux in TNBC cells that were dependent on autophagy for survival and displayed synergistic cytotoxicity with the poly (ADP-ribose) polymerase (PARP) inhibitor olaparib against TNBC cells. These data suggest that combining ULK1/2 and PARP inhibition may have clinical utility for the treatment of TNBC.

DOI: https://doi.org/10.1021/acs.jmedchem.0c00873
Food Funct. 2020 Nov 20.

Malvidin-3-O-arabinoside ameliorates ethyl carbamate-induced oxidative damage by stimulating AMPK-mediated autophagy.

Yuting Li, Yang Xu, Jiahong Xie, Wei Chen.

Ethyl carbamate (EC) is an environmental toxin, commonly present in various fermented foods and beverages, as well as tobacco and polluted ambient air. However, studies on the effects of EC-induced toxicity on the intestines and potential protection methods are limited. In this study, we show that EC could cause severe toxicity in intestinal epithelial cells (IECs) triggering the induction of decreased cell viability, ROS accumulation and glutathione (GSH) depletion in a dose-dependent manner. Based on these results, we established an EC-treated IEC model to screen the potential protective effects of 12 kinds of anthocyanins extracted from blueberry. Interestingly, we found that malvidin-3-O-arabinoside (M3A) significantly reversed the oxidative damage caused by EC exposure by stimulating autophagy flux, which was determined by the LC3-II level and GFP-RFP-LC3 transfection experiment. Enhancement of autophagy was mainly ascribed to the regulation of lysosomes. M3A pretreatment remarkably upregulated LAMP-1 expression, which indicated elevated lysosomal mass. Besides, M3A also successfully restored lysosomal acidity and subsequently strengthened lysosomal functions. Furthermore, M3A stimulated phosphorylation of AMP-activated protein kinase (AMPK), a master regulator of autophagy. Furthermore, our study indicated the possibility of EC-caused oxidative damage to the intestines and unveiled the remarkably protective benefits of M3A-induced AMPK-mediated autophagy against this toxicity.

DOI: https://doi.org/10.1039/d0fo01562h
Front Microbiol. 2020 ;11 589604

The Host Autophagy During Toxoplasma Infection.

Minmin Wu, Obed Cudjoe, Jilong Shen, Ying Chen, Jian Du.

  Autophagy is an important homeostatic mechanism, in which lysosomes degrade and recycle cytosolic components. As a key defense mechanism against infections, autophagy is involved in the capture and elimination of intracellular parasites. However, intracellular parasites, such as Toxoplasma gondii, have developed several evasion mechanisms to manipulate the host cell autophagy for their growth and establish a chronic infection. This review provides an insight into the autophagy mechanism used by the host cells in the control of T. gondii and the host exploitation by the parasite. First, we summarize the mechanism of autophagy, xenophagy, and LC3-associated phagocytosis. Then, we illustrate the process of autophagy proteins-mediated T. gondii clearance. Furthermore, we discuss how the parasite blocks and exploits this process for its survival.

Keywords:  IFN-γ mediated pathogen elimination; LC3-associated phagocytosis; Toxoplasma gondii; autophagy; xenophagy

DOI:  https://doi.org/10.3389/fmicb.2020.589604
Mol Cell Biochem. 2020 Nov 16.

Autophagy: a promising therapeutic target for improving mesenchymal stem cell biological functions.

Jiaqiang Deng, Lijun Zhong, Zihan Zhou, Congwei Gu, Xiaoya Huang, Liuhong Shen, Suizhong Cao, Zhihua Ren, Zhicai Zuo, Junliang Deng, Shumin Yu.

  Mesenchymal stem cells (MSCs) are considered to be a promising therapeutic material due to their capacities for self-renewal, multilineage differentiation, and immunomodulation and have attracted great attention in regenerative medicine. However, MSCs may lose their biological functions because of donor age or disease and environmental pressure before and after transplantation, which hinders the application of MSC-based therapy. As a major intracellular lysosome-dependent degradative process, autophagy plays a pivotal role in maintaining cellular homeostasis and withstanding environmental pressure and may become a potential therapeutic target for improving MSC functions. Recent studies have demonstrated that the regulation of autophagy is a promising approach for improving the biological properties of MSCs. More in-depth investigations about the role of autophagy in MSC biology are required to contribute to the clinical application of MSCs. In this review, we focus on the role of autophagy regulation by various physical and chemical factors on the biological functions of MSCs in vitro and in vivo, and provide some strategies for enhancing the therapeutic efficacy of MSCs.

Keywords:  Angiogenesis; Autophagy; Differentiation; Immunosuppression; MSCs; Survival

DOI:  https://doi.org/10.1007/s11010-020-03978-2
Int J Mol Sci. 2020 Nov 13. pii: E8559. [Epub ahead of print]21(22):

Axon Injury-Induced Autophagy Activation Is Impaired in a C. elegans Model of Tauopathy.

Su-Hyuk Ko, Gilberto Gonzalez, Zhijie Liu, Lizhen Chen.

  Autophagy is a conserved pathway that plays a key role in cell homeostasis in normal settings, as well as abnormal and stress conditions. Autophagy dysfunction is found in various neurodegenerative diseases, although it remains unclear whether autophagy impairment is a contributor or consequence of neurodegeneration. Axonal injury is an acute neuronal stress that triggers autophagic responses in an age-dependent manner. In this study, we investigate the injury-triggered autophagy response in a C. elegans model of tauopathy. We found that transgenic expression of pro-aggregant Tau, but not the anti-aggregant Tau, abolished axon injury-induced autophagy activation, resulting in a reduced axon regeneration capacity. Furthermore, axonal trafficking of autophagic vesicles were significantly reduced in the animals expressing pro-aggregant F3ΔK280 Tau, indicating that Tau aggregation impairs autophagy regulation. Importantly, the reduced number of total or trafficking autophagic vesicles in the tauopathy model was not restored by the autophagy activator rapamycin. Loss of PTL-1, the sole Tau homologue in C. elegans, also led to impaired injury-induced autophagy activation, but with an increased basal level of autophagic vesicles. Therefore, we have demonstrated that Tau aggregation as well as Tau depletion both lead to disruption of injury-induced autophagy responses, suggesting that aberrant protein aggregation or microtubule dysfunction can modulate autophagy regulation in neurons after injury.

Keywords:  Tau; aggregation; autophagy; axon injury; axon regeneration; ptl-1; tauopathy

DOI:  https://doi.org/10.3390/ijms21228559
EMBO J. 2020 Nov 20. e104532

The E3 ubiquitin ligase Peli1 regulates the metabolic actions of mTORC1 to suppress antitumor T cell responses.

Chun-Jung Ko, Lingyun Zhang, Zuliang Jie, Lele Zhu, Xiaofei Zhou, Xiaoping Xie, Tianxiao Gao, Jin-Young Yang, Xuhong Cheng, Shao-Cong Sun.

  Metabolic fitness of T cells is crucial for immune responses against infections and tumorigenesis. Both the T cell receptor (TCR) signal and environmental cues contribute to the induction of T cell metabolic reprogramming, but the underlying mechanism is incompletely understood. Here, we identified the E3 ubiquitin ligase Peli1 as an important regulator of T cell metabolism and antitumor immunity. Peli1 ablation profoundly promotes tumor rejection, associated with increased tumor-infiltrating CD4 and CD8 T cells. The Peli1-deficient T cells display markedly stronger metabolic activities, particularly glycolysis, than wild-type T cells. Peli1 controls the activation of a metabolic kinase, mTORC1, stimulated by both the TCR signal and growth factors, and this function of Peli1 is mediated through regulation of the mTORC1-inhibitory proteins, TSC1 and TSC2. Peli1 mediates non-degradative ubiquitination of TSC1, thereby promoting TSC1-TSC2 dimerization and TSC2 stabilization. These results establish Peli1 as a novel regulator of mTORC1 and downstream mTORC1-mediated actions on T cell metabolism and antitumor immunity.

Keywords:  Peli1; T cell metabolism; antitumor immunity; mTORC1; ubiquitination

DOI:  https://doi.org/10.15252/embj.2020104532
Prog Neurobiol. 2020 Nov 11. pii: S0301-0082(20)30208-2. [Epub ahead of print] 101953

Increased telomerase improves motor function and alpha-synuclein pathology in a transgenic mouse model of Parkinson's disease associated with enhanced autophagy.

Tengfei Wan, Emma J Weir, Mary Johnson, Viktor I Korolchuk, Gabriele C Saretzki.

  Protective effects of the telomerase protein TERT have been shown in neurons and brain. We previously demonstrated that TERT protein can accumulate in mitochondria of Alzheimer's disease (AD) brains and protect from pathological tau in primary mouse neurons. This prompted us to employ telomerase activators in order to boost telomerase expression in a mouse model of Parkinson's disease (PD) overexpressing human wild type α-synuclein. Our aim was to test whether increased Tert expression levels were able to ameliorate PD symptoms and to activate protein degradation. We found increased Tert expression in brain for both activators which correlated with a substantial improvement of motor functions such as gait and motor coordination while telomere length in the analysed region was not changed. Interestingly, only one activator (TA-65) resulted in a decrease of reactive oxygen species from brain mitochondria. Importantly, we demonstrate that total, phosphorylated and aggregated α-synuclein were significantly decreased in the hippocampus and neocortex of activator-treated mice corresponding to enhanced markers of autophagy suggesting an improved degradation of toxic alpha-synuclein. We conclude that increased Tert expression caused by telomerase activators is associated with decreased α-synuclein protein levels either by activating autophagy or by preventing or delaying degradation mechanisms which are impaired during disease progression. This encouraging preclinical data could be translated into novel therapeutic options for neurodegenerative disorders such as PD.

Keywords:  Parkinson’s disease; Telomerase; Telomerase activator; alpha-synuclein; autophagy; motor function; transgenic mouse model

DOI:  https://doi.org/10.1016/j.pneurobio.2020.101953
Bioact Mater. 2021 May;6(5): 1413-1422

Substrate stiffness differentially impacts autophagy of endothelial cells and smooth muscle cells.

Mi Hu, Fan Jia, Wei-Pin Huang, Xu Li, Deng-Feng Hu, Jing Wang, Ke-Feng Ren, Guo-Sheng Fu, Yun-Bing Wang, Jian Ji.

  Stiffening of blood vessels is one of the most important characteristics in the process of many cardiovascular pathologies such as atherosclerosis, angiosteosis, and vascular aging. Increased stiffness of the vascular extracellular matrix drives artery pathology and alters phenotypes of vascular cell. Understanding how substrate stiffness impacts vascular cell behaviors is of great importance to the biomaterial design in tissue engineering, regenerative medicine, and medical devices. Here we report that changing substrate stiffness has a significant impact on the autophagy of vascular endothelial cells (VECs) and smooth muscle cells (VSMCs). Interestingly, our findings demonstrate that, with the increase of substrate stiffness, the autophagy level of VECs and VSMCs showed differential changes: endothelial autophagy levels reduced, leading to the reductions in a range of gene expression associated with endothelial function; while, autophagy levels of VSMCs increased, showing a transition from contractile to the synthetic phenotype. We further demonstrate that, by inhibiting cell autophagy, the expressions of endothelial functional gene were further reduced and the expression of VSMC calponin increased, suggesting an important role of autophagy in response of the cells to the challenge of microenvironment stiffness changing. Although the underlying mechanism requires further study, this work highlights the relationship of substrate stiffness, autophagy, and vascular cell behaviors, and enlightening the design principles of surface stiffness of biomaterials in cardiovascular practical applications.

Keywords:  Autophagy; Endothelial cells; Polyelectrolyte film; Smooth muscle cells; Substrate stiffness

DOI:  https://doi.org/10.1016/j.bioactmat.2020.10.013
Biomolecules. 2020 Nov 14. pii: E1553. [Epub ahead of print]10(11):

Autophagy Stimulus-Dependent Role of the Small GTPase Ras2 in Peroxisome Degradation.

Fahd Boutouja, Harald W Platta.

  The changing accessibility of nutrient resources induces the reprogramming of cellular metabolism in order to adapt the cell to the altered growth conditions. The nutrient-depending signaling depends on the kinases mTOR (mechanistic target of rapamycin), which is mainly activated by nitrogen-resources, and PKA (protein kinase A), which is mainly activated by glucose, as well as both of their associated factors. These systems promote protein synthesis and cell proliferation, while they inhibit degradation of cellular content by unselective bulk autophagy. Much less is known about their role in selective autophagy pathways, which have a more regulated cellular function. Especially, we were interested to analyse the central Ras2-module of the PKA-pathway in the context of peroxisome degradation. Yeast Ras2 is homologous to the mammalian Ras proteins, whose mutant forms are responsible for 33% of human cancers. In the present study, we were able to demonstrate a context-dependent role of Ras2 activity depending on the type of mTOR-inhibition and glucose-sensing situation. When mTOR was inhibited directly via the macrolide rapamycin, peroxisome degradation was still partially suppressed by Ras2, while inactivation of Ras2 resulted in an enhanced degradation of peroxisomes, suggesting a role of Ras2 in the inhibition of peroxisome degradation in glucose-grown cells. In contrast, the inhibition of mTOR by shifting cells from oleate-medium, which lacks glucose, to pexophagy-medium, which contains glucose and is limited in nitrogen, required Ras2-activity for efficient pexophagy, strongly suggesting that the role of Ras2 in glucose sensing-associated signaling is more important in this context than its co-function in mTOR-related autophagy-inhibition.

Keywords:  Ras2; autophagy; mTOR; peroxisomes; pexophagy; rapamycin

DOI:  https://doi.org/10.3390/biom10111553
J Biol Chem. 2020 Nov 20. pii: jbc.RA120.014682. [Epub ahead of print]

ABHD5 suppresses cancer cell anabolism through lipolysis-dependent activation of the AMPK/mTORC1 pathway.

Guohua Chen, Guoli Zhou, Aaron Lotvola, James G Granneman, Jian Wang.

  ABHD5 is an essential coactivator of ATGL, the rate-limiting triglyceride (TG) lipase in many cell types. Importantly, ABHD5 also functions as a tumor suppressor, and ABHD5 mRNA expression levels correlate with patient survival for several cancers. Nevertheless, the mechanisms involved in ABHD5-dependent tumor suppression are not known. We found that overexpression of ABHD5 induces cell-cycle arrest at the G1 phase and causes growth retardation in a panel of prostate cancer cells. Transcriptomic profiling and biochemical analysis revealed that genetic or pharmacological activation of lipolysis by ABHD5 potently inhibits mTORC1 signaling, leading to a significant downregulation of protein synthesis. Mechanistically, we found that ABHD5 elevates intracellular AMP content, which activates AMPK, leading to inhibition of mTORC1. Interestingly, ABHD5-dependent suppression of mTORC1 was abrogated by pharmacological inhibition of DGAT1 or DGAT2, isoenzymes that re-esterify fatty acids in a process that consumes ATP. Collectively, this study maps out a novel molecular pathway crucial for limiting cancer cell proliferation, in which ABHD5-mediated lipolysis creates an energy-consuming futile cycle between TG hydrolysis and resynthesis, leading to inhibition of mTORC1 and cancer cell growth arrest.

Keywords:  lipid signaling; lipolysis; metabolic regulation; tumor cell biology; tumor metabolism

DOI:  https://doi.org/10.1074/jbc.RA120.014682
Aging (Albany NY). 2020 Nov 16. 12

Autophagy is induced in human keratinocytes during human papillomavirus 11 pseudovirion entry.

Rui Han, Chunting Hua, Siyuan Sun, Boya Zhang, Yinjing Song, Stijn van der Veen, Hao Cheng.

  Human papillomavirus type 11 (HPV11) is one of the main causes of condyloma acuminatum, a widespread sexually transmitted disease. During infection of its primary target cell, keratinocytes, it is likely to encounter the autophagy pathway, which is an intracellular maintenance process that is also able to target invading pathogens. It is currently unknown whether HPV11 is targeted by autophagy or whether it is able to escape autophagy-mediated killing. Here, we investigated the autophagy response during HPV11 pseudovirion (PsV) entry in human keratinocytes. Transmission electron microscopy showed that intracellular PsVs were sequestered in lumen of double-membrane autophagosomes that subsequently appeared to fuse with lysosomes, while confocal microscopy showed induction LC3 puncta, the hallmark of induced autophagy activity. Furthermore, quantitative infection assays showed that high autophagy activity resulted in reduced HPV11 PsV infectivity. Therefore, the autophagy pathway seemed to actively target invading HPV11 PsVs for destruction in the autolysosome. Western analysis on the phosphorylation state of autophagy regulators and upstream pathways indicated that autophagy was activated through interplay between Erk and Akt signaling. In conclusion, autophagy functions as a cellular protection mechanism against intracellular HPV11 and therefore therapies that stimulate autophagy may prevent recurrent condyloma acuminatum by helping eliminate latent HPV11 infections.

Keywords:  autophagy; entry; human papillomavirus 11; keratinocyte; pseudovirion

DOI:  https://doi.org/10.18632/aging.104046
Cancers (Basel). 2020 Nov 16. pii: E3388. [Epub ahead of print]12(11):

Induction of Lysosomal Membrane Permeabilization Is a Major Event of FTY720-Mediated Non-Apoptotic Cell Death in Human Glioma Cells.

Kyoung-Jin Min, Taeg Kyu Kwon.

  FTY720, a sphingosine-1-phosphate (S1P) receptor modulator, is a synthetic compound produced by the modification of a metabolite from I. sinclairii. Here, we found that FTY720 induced non-apoptotic cell death in human glioma cells (U251MG, U87MG, and U118MG). FTY720 (10 µM) dramatically induced cytoplasmic vacuolation in glioma cells. However, FTY720-mediated vacuolation and cell death are not associated with autophagy. Genetic or pharmacological inhibition of autophagy did not inhibit FTY720-induced cell death. Herein, we detected that FTY720-induced cytoplasmic vacuoles were stained with lysotracker red, and FTY720 induced lysosomal membrane permeabilization (LMP). Interestingly, cathepsin inhibitors (E64D and pepstatin A) and ectopic expression of heat shock protein 70 (HSP70), which is an endogenous inhibitor of LMP, markedly inhibited FTY720-induced cell death. Our results demonstrated that FTY720 induced non-apoptotic cell death via the induction of LMP in human glioma cells.

Keywords:  FTY720; LMP; cathepsins; glioma; non-apoptotic cell death

DOI:  https://doi.org/10.3390/cancers12113388
Cell Chem Biol. 2020 Nov 19. pii: S2451-9456(20)30429-3. [Epub ahead of print]27(11): 1329-1331

Dual Targeting of v-ATPase and mTORC1 Signaling Disarms Multidrug-Resistant Cancers.

Tayah Turocy, Jason M Crawford.

Chemotherapeutic treatments are frequently impeded by the development of multidrug resistance (MDR). In this issue of Cell Chemical Biology, Wang et al. (2020) identify the natural product verucopeptin as having therapeutic potential toward MDR cancer cell types by targeting v-ATPase and mTORC1 signaling.

DOI: https://doi.org/10.1016/j.chembiol.2020.10.013
Stem Cell Rev Rep. 2020 Nov 17.

Signaling Network Centered on mTORC1 Dominates Mammalian Intestinal Stem Cell Ageing.

Shao-Jie Liang, Jia-Yi Zhou, Xiu-Qi Wang.

  The intestine integrates the function of digestion, absorption, and barrier, which is easily damaged by the external factors upon ageing. The intestinal stem cells (ISCs) exist at the intestinal crypt base and play an indispensable role in intestinal homeostasis and regeneration. The intestine ageing contributes to malabsorption and other associated illnesses, which were considered to be related to ISCs. Here, we summarize the current research progress of mammalian ISCs ageing and pay more attention to the central regulatory role of the mTORC1 signaling pathway in regulating mammalian ISCs ageing, and its related AMPK, FOXO, Wnt signaling pathways. Furthermore, we also discuss the interventions aimed at mTORC1 and its associated signaling pathways, which may provide potential strategies for rejuvenating aged ISCs and the therapy of age-related intestinal diseases. Graphical abstract Many signaling pathways are altered in the ageing ISCs, thereby inducing the decrease of ISC self-renewal, differentiation, and regeneration, an increasing of oxidative stress may contribute to damage to the ISCs. Interventions such as calorie restriction, fasting and so on can effectively alleviate these adverse effects.

Keywords:  AMPK; Ageing; FOXO; Intestinal stem cells; Wnt; mTORC1

DOI:  https://doi.org/10.1007/s12015-020-10073-y
Aging (Albany NY). 2020 Nov 16. 12

TRIB3 facilitates glioblastoma progression via restraining autophagy.

Zhanbin Tang, Hongping Chen, Di Zhong, Wan Wei, Lili Liu, Qiong Duan, Baichao Han, Guozhong Li.

  The pseudokinase Tribble 3 (TRIB3) is known as a regulator in cellular responses to a variety of stresses, such as glucose insufficiency and endoplasmic reticulum (ER) stress. TRIB3 is upregulated in various cancer tissues and is closely connected to the poor prognosis of patients. However, the underlying regulation and function of TRIB3 in glioblastoma (GBM) is still largely unknown. In this study, the upregulation of TRIB3 was confirmed both in primary specimens from GBM patients and in vitro with GBM cell lines. Overexpression of specific TRIB3 transcripts promoted cell growth and migration in vitro, while knockdown of TRIB3 expression exerted a repressive effect on these cellular processes. The growth-promoting effect of TRIB3 was also demonstrated in a xenograft mouse model. Mechanistic studies further revealed that TRIB3 was able to suppress autophagic flux and that this suppression was responsible for TRIB3 silencing-induced proliferation and migration of GBM cells. These findings indicate that the suppression of autophagic flux by TRIB3 drives the invasion and proliferation of GBM cells, thus suggesting that TRIB3 is a potential novel therapeutic target for the treatment of glioma.

Keywords:  autophagy; glioblastoma; tribbles pseudokinase 3

DOI:  https://doi.org/10.18632/aging.103969
Sci Rep. 2020 Nov 19. 10(1): 20189

Inverse sex-based expression profiles of PTEN and Klotho in mice.

Natalia Prudente de Mello, Diana Zukas Andreotti, Ana Maria Orellana, Cristoforo Scavone, Elisa Mitiko Kawamoto.

Sex differences are considered predictive factors in the development of several neurological diseases, which are also known to coincide with impaired phosphoinositide 3-kinase (PI3K)-AKT pathway activity, an essential signaling cascade involved in the control of several cellular functions such as autophagy and apoptosis. Here, under physiological conditions, we show important sex differences in the underlying balancing mechanisms that lead to similar AKT activity levels and autophagy and apoptosis processes in the two sexes. We demonstrate inverse sex-based expression of PTEN and Klotho, two important proteins that are known to negatively regulate the AKT pathway, and inverse sex-dependent levels of mTOR and FoxO3a activity. Taken together, our findings indicate that inverse sex-based regulation may be one of the underlying balancing mechanisms that differ between the sexes and a possible cause of sex-based autophagic and apoptotic responses to triggering situations that can lead to a sex-based predisposition to some neurological diseases.

DOI: https://doi.org/10.1038/s41598-020-77217-5
Cell Death Differ. 2020 Nov 18.

Ubiquitin signaling in neurodegenerative diseases: an autophagy and proteasome perspective.

François Le Guerroué, Richard J Youle.

Ubiquitin signaling is a sequence of events driving the fate of a protein based on the type of ubiquitin modifications attached. In the case of neurodegenerative diseases, ubiquitin signaling is mainly associated with degradation signals to process aberrant proteins, which form aggregates often fatal for the brain cells. This signaling is often perturbed by the aggregates themselves and leads to the accumulation of toxic aggregates and inclusion bodies that are deleterious due to a toxic gain of function. Decrease in quality control pathways is often seen with age and is a critical onset for the development of neurodegeneration. Many aggregates are now thought to propagate in a prion-like manner, where mutated proteins acting like seeds are transitioning from cell to cell, converting normal proteins to toxic aggregates. Modulation of ubiquitin signaling, by stimulating ubiquitin ligase activation, is a potential therapeutic strategy to treat patients with neurodegeneration diseases.

DOI: https://doi.org/10.1038/s41418-020-00667-x
Am J Transl Res. 2020 ;12(10): 6793-6810

All-trans-retinoic acid inhibits the malignant behaviors of hepatocarcinoma cells by regulating autophagy.

Shuyu Fang, Chaoqun Hu, Lei Xu, Jiejie Cui, Li Tao, Mengjia Gong, Yi Wang, Yun He, Tongchuan He, Yang Bi.

Hepatocellular carcinoma is the fourth leading cause of cancer-related deaths due to its high rate of recurrence and metastasis. All-trans-retinoic acid (ATRA) can inhibit the malignant behaviors of hepatocarcinoma cells. Autophagy is reportedly involved in the migration and metastasis of various cancer cells. This study aimed to investigate the effect of autophagy on the function of ATRA on hepatocarcinoma cells, and to explore its possible underlying mechanism. Hepatocarcinoma cell lines, Hepa1-6 and HepG2, were treated with ATRA and autophagy inhibitors, including 3-methyladenine (3-MA) and Bafilomycin (Baf). Transmission electron microscopy, laser scanning, western blot, and real-time PCR demonstrated that ATRA induces autophagy in hepatocarcinoma cells. Trypan blue staining, a wound healing assay, and a transwell assay showed that 3-MA and Baf reverses the inhibitory functions of ATRA on the proliferation, migration, and invasion of hepatocarcinoma cells. Flow cytometry, Hoechst staining, periodic acid-Schiff staining, and indocyanine green uptake validated that 3-MA and Baf reverses the function of ATRA on apoptosis and the differentiation of hepatocarcinoma cells. Real-time PCR, western blot, and an immunofluorescence assay demonstrated that the reversal of the epithelial-mesenchymal transition (EMT) process by ATRA is weakened when autophagy is inhibited. Additionally, we confirmed that Bcl-2 is associated with the induction of ATRA-induced autophagy instead of the PI3K/Akt/mTOR pathway. These findings suggest that ATRA induces autophagy and autophagic cell death through the Bcl-2/Beclin1 pathway. Furthermore, ATRA-induced autophagy is involved in the inhibitory effect of ATRA on the malignant behaviors of hepatocarcinoma cells by reversing the EMT process.

Keywords: Bcl-2; Hepatocellular carcinoma; all-trans-retinoic acid; autophagy; epithelial-mesenchymal transition