bims-apauto 2022-11-27 papers

bims-apauto

Biomed News

on Apoptosis and autophagy

Issue of 2022‒11‒27
seven papers selected by
Su Hyun Lee
Seoul National University

Autophagic lysosome reformation in health and disease.
Extracellular SQSTM1 exacerbates acute pancreatitis by activating autophagy-dependent ferroptosis.
FACS-mediated isolation of native autophagic vesicles.
Autophagy promotes cell survival by maintaining NAD levels.
Defensosomes: a new role for autophagy proteins in innate immune defense.
Identification and Characterization of Physiological Pairing of E2 Ubiquitin-Conjugating Enzymes and E3 Ubiquitin Ligases.
BTN3A1 promotes tumor progression and radiation resistance in esophageal squamous cell carcinoma by regulating ULK1-mediated autophagy.

Autophagy. 2022 Nov 21. 1-18

Autophagic lysosome reformation in health and disease.

Randini Nanayakkara, Rajendra Gurung, Samuel J Rodgers, Matthew J Eramo, Georg Ramm, Christina A Mitchell, Meagan J McGrath.

  Lysosomes are the primary degradative compartment within cells and there have been significant advances over the past decade toward understanding how lysosome homeostasis is maintained. Lysosome repopulation ensures sustained autophagy function, a fundamental process that protects against disease. During macroautophagy/autophagy, cellular debris is sequestered into phagophores that mature into autophagosomes, which then fuse with lysosomes to generate autolysosomes in which contents are degraded. Autophagy cannot proceed without the sufficient generation of lysosomes, and this can be achieved via their de novo biogenesis. Alternatively, during autophagic lysosome reformation (ALR), lysosomes are generated via the recycling of autolysosome membranes. During this process, autolysosomes undergo significant membrane remodeling and scission to generate membrane fragments, that mature into functional lysosomes. By utilizing membranes already formed during autophagy, this facilitates an efficient pathway for re-deriving lysosomes, particularly under conditions of prolonged autophagic flux. ALR dysfunction is emerging as an important disease mechanism including for neurodegenerative disorders such as hereditary spastic paraplegia and Parkinson disease, neuropathies including Charcot-Marie-Tooth disease, lysosome storage disorders, muscular dystrophy, metabolic syndrome, and inflammatory and liver disorders. Here, we provide a comprehensive review of ALR, including an overview of its dynamic spatiotemporal regulation by MTOR and phosphoinositides, and the role ALR dysfunction plays in many diseases.

Keywords:  Autophagic lysosome reformation; MTOR; PtdIns(4,5)P2; PtdIns4P; lysosome; phosphoinositide

DOI:  https://doi.org/10.1080/15548627.2022.2128019
Autophagy. 2022 Nov 25.

Extracellular SQSTM1 exacerbates acute pancreatitis by activating autophagy-dependent ferroptosis.

Liangchun Yang, Fanghua Ye, Jiao Liu, Daniel J Klionsky, Daolin Tang, Rui Kang.

  Acute pancreatitis (AP) is an abdominal inflammatory disease initiated by damaged pancreatic acinar cells and developed by systemic inflammation. SQSTM1 (sequestosome 1) has an intracellular function in mediating substrate degradation during macroautophagy/autophagy, and it can be released by macrophages and monocytes to trigger lethal inflammation during bacterial infection. Here, we report that extracellular SQSTM1 acts as a mediator of AP by enhancing the sensitivity to autophagy-dependent ferroptotic cell death. Serum SQSTM1 is elevated in AP patients as well as in mice that have cerulein-induced AP. The administration of SQSTM1-neutralizing antibodies protects against experimental AP in mice. Mechanistically, recombinant SQSTM1 protein (rSQSTM1) increases AGER (advanced glycosylation end-product specific receptor)-dependent ACSL4 (acyl-CoA synthetase long chain family member 4) expression, leading to polyunsaturated fatty acid production for autophagosome formation and subsequent ferroptosis. The rSQSTM1-elicited pathological responses during AP are attenuated in mice with the conditional deletion of Ager in the pancreas. These findings may provide not only new insights into the mechanism of autophagy-dependent cell death, but also suggest that targeting the extracellular SQSTM1 pathway is a potential strategy for the treatment of AP.

Keywords:  SQSTM1; autophagy; ferroptosis; inflammation; lipid peroxidation; pancreatitis

DOI:  https://doi.org/10.1080/15548627.2022.2152209
Autophagy. 2022 Nov 23.

FACS-mediated isolation of native autophagic vesicles.

Daniel Schmitt, Süleyman Bozkurt, Pascale Henning-Domres, Heike Huesmann, Stefan Eimer, Laura Bindila, Christian Behrends, Emily Boyle, Florian Wilfling, Georg Tascher, Christian Münch, Christian Behl, Andreas Kern.

  Autophagosome isolation enables the thorough investigation of structural components and engulfed materials. Recently, we introduced a novel antibody-based FACS-mediated method for isolation of native macroautophagic/autophagic vesicles and confirmed the quality of the preparations. We performed phospholipidomic and proteomic analyses to characterize autophagic vesicle-associated phospholipids and protein cargoes under different autophagy conditions. Lipidomic analyses identified phosphoglycerides and sphingomyelins within autophagic vesicles and revealed that the lipid composition was unaffected by different rates of autophagosome formation. Proteomic analyses identified more than 4500 potential autophagy substrates and showed that in comparison to autophagic vesicles isolated under basal autophagy conditions, starvation only marginally affected the cargo profile. Proteasome inhibition, however, resulted in the enhanced degradation of ubiquitin-proteasome system components. Taken together, the novel isolation method enriched large quantities of autophagic vesicles and enabled detailed analyses of their lipid and cargo composition.

Keywords:  autophagic vesicles; autophagy; cargo profiling; lipid profiling; vesicle isolation

DOI:  https://doi.org/10.1080/15548627.2022.2151188
Dev Cell. 2022 Nov 21. pii: S1534-5807(22)00760-2. [Epub ahead of print]57(22): 2584-2598.e11

Autophagy promotes cell survival by maintaining NAD levels.

Tetsushi Kataura, Lucia Sedlackova, Elsje G Otten, Ruchika Kumari, David Shapira, Filippo Scialo, Rhoda Stefanatos, Kei-Ichi Ishikawa, George Kelly, Elena Seranova, Congxin Sun, Dorothea Maetzel, Niall Kenneth, Sergey Trushin, Tong Zhang, Eugenia Trushina, Charles C Bascom, Ryan Tasseff, Robert J Isfort, John E Oblong, Satomi Miwa, Michael Lazarou, Rudolf Jaenisch, Masaya Imoto, Shinji Saiki, Manolis Papamichos-Chronakis, Ravi Manjithaya, Oliver D K Maddocks, Alberto Sanz, Sovan Sarkar, Viktor I Korolchuk.

  Autophagy is an essential catabolic process that promotes the clearance of surplus or damaged intracellular components. Loss of autophagy in age-related human pathologies contributes to tissue degeneration through a poorly understood mechanism. Here, we identify an evolutionarily conserved role of autophagy from yeast to humans in the preservation of nicotinamide adenine dinucleotide (NAD) levels, which are critical for cell survival. In respiring mouse fibroblasts with autophagy deficiency, loss of mitochondrial quality control was found to trigger hyperactivation of stress responses mediated by NADases of PARP and Sirtuin families. Uncontrolled depletion of the NAD(H) pool by these enzymes ultimately contributed to mitochondrial membrane depolarization and cell death. Pharmacological and genetic interventions targeting several key elements of this cascade improved the survival of autophagy-deficient yeast, mouse fibroblasts, and human neurons. Our study provides a mechanistic link between autophagy and NAD metabolism and identifies targets for interventions in human diseases associated with autophagic, lysosomal, and mitochondrial dysfunction.

Keywords:  DNA damage; NAD; PARP; Sirtuins; ageing; autophagy; metabolism; mitochondria; mitophagy

DOI:  https://doi.org/10.1016/j.devcel.2022.10.008
Autophagy. 2022 Nov 21. 1-3

Defensosomes: a new role for autophagy proteins in innate immune defense.

Krystal L Ching, Victor J Torres, Ken Cadwell.

  In recent years, the contribution of exosomes to immunity, inflammation and host-pathogen interaction have been appreciated. Exosomes are small secreted extracellular vesicles from endosomal origin that contain a myriad of cellular molecules (protein, nucleic acids), including surface receptors. We have reported a pathogen-induced and macroautophagy/autophagy-dependent class of exosomes coined as "defensosomes", which protect the host from membrane-targeting toxins. In a recent study, we found that defensosomes decorated with ACE2, the SARS-CoV-2 cellular receptor, are produced in the lungs of patients with COVID-19, and that increased concentration of ACE2-loaded defensosomes is associated with decreased hospitalization length. Mechanistically, SARS-CoV-2 induces the production of ACE2-coated defensosomes, a process requiring the autophagy machinery, which in turn binds and neutralizes the virus. We propose that defensosomes represent a new form of autophagy-mediated innate immunity that contributes to the host's armamentarium against pathogens.

Keywords:  ATG16L1; COVID-19; SARS-CoV-2; defensosomes; extracellular vesicles; innate immunity; staphylococcus aureus; toll-like receptor

DOI:  https://doi.org/10.1080/15548627.2022.2146894
Methods Mol Biol. 2023 ;2581 13-29

Identification and Characterization of Physiological Pairing of E2 Ubiquitin-Conjugating Enzymes and E3 Ubiquitin Ligases.

Carla Brillada, Marco Trujillo.

  The posttranslational attachment of the small protein modifier ubiquitin (Ub) is best known for its function in targeting proteins for degradation by the proteasome. However, ubiquitination also serves as a signal determining protein localization, activity, and interaction. Ubiquitination requires the sequential activity of E1 ubiquitin-activating enzyme (UBA), E2 ubiquitin-conjugating enzyme (UBC), and E3 ubiquitin ligase. Recognition of a target protein by an Ub-E2-E3 complex can result in its mono-ubiquitination (attachment of a single Ub moiety) or poly-ubiquitination, i.e., attachment of Ub chains. While the E3 ligase is important for the reaction specificity, the E2s catalyze the attachment of Ub to the target and to Ub itself to generate chains. In Arabidopsis thaliana, there are two E1s, 37 UBCs (and two ubiquitin-like conjugating enzymes) and more than 1400 E3 ligases, working in a combinatorial way. Therefore, in order to understand E3 ligase function, it is important to frame it within its possible E2s interactors. In this chapter, we propose a two-step identification and characterization of physiological E2-E3 pairs. In a first step, in vivo interacting E2s are identified through bimolecular fluorescence complementation (BiFC) using transient expression in Arabidopsis protoplast. In the second step, the activity of E2-E3 pairs is analyzed by a synthetic biology approach in which autoubiquitination is reconstituted in bacteria.

Keywords:  Autoubiquitination; BiFC; E2-E3 pairing; E3 ubiquitin ligase; Ubiquitin; Ubiquitin-conjugating enzyme (E2)

DOI:  https://doi.org/10.1007/978-1-0716-2784-6_2
Cell Death Dis. 2022 Nov 22. 13(11): 984

BTN3A1 promotes tumor progression and radiation resistance in esophageal squamous cell carcinoma by regulating ULK1-mediated autophagy.

Wenjing Yang, Bo Cheng, Pengxiang Chen, Xiaozheng Sun, Zhihua Wen, Yufeng Cheng.

Radiotherapy is one of the most effective treatments for esophageal squamous cell carcinoma (ESCC); however, radioresistance is a clinical problem that must urgently be solved. Here, we found that butyrophilin subfamily 3 member A1 (BTN3A1) is upregulated in ESCC tumor tissues compared with nontumor tissues. We also evaluated BTN3A1 expression in patients with ESCC receiving adjuvant radiotherapy. The results demonstrated that BTN3A1 upregulation predicts a poor prognosis for ESCC patients. BTN3A1 overexpression promotes ESCC cell proliferation in vitro and in vivo. Moreover, BTN3A1 knockdown sensitized ESCC cells to radiation. We further explored the mode of death involved in BTN3A1-mediated radioresistance. Previous studies have shown that apoptosis, autophagy, necrosis, pyroptosis and ferroptosis are important for the survival of ESCC cells. We performed an RT-PCR array and western blotting (WB) to identify the mode of death and revealed for the first time that BTN3A1 promotes cell radioresistance by activating autophagy. In addition, by performing immunoprecipitation and mass spectrometry analyses, we found that BTN3A1 regulated the expression of UNC-51-like autophagy activating kinase 1(ULK1) and promoted its phosphorylation to subsequently initiate autophagy. Chromatin immunoprecipitation (ChIP) and luciferase reporter assay results indicated that BTN3A1 is a novel direct target of hypoxia inducible factor-1α (HIF-1α). HIF-1α, a transcription factor, promotes BTN3A1 transcription upon irradiation. Overall, the present study is the first to show that BTN3A1 plays a key role in radioresistance and that targeting BTN3A1 might be a promising strategy to improve radiotherapy efficacy in patients with ESCC.

DOI: https://doi.org/10.1038/s41419-022-05429-w