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Recruitment of the autophagic machinery to endosomes during infection is mediated by ubiquitin.

ABSTRACT: Although ubiquitin is thought to be important for the autophagic sequestration of invading bacteria (also called xenophagy), its precise role remains largely enigmatic. Here we determined how ubiquitin is involved in this process. After invasion, ubiquitin is conjugated to host cellular proteins in endosomes that contain Salmonella or transfection reagent-coated latex (polystyrene) beads, which mimic invading bacteria. Ubiquitin is recognized by the autophagic machinery independently of the LC3-ubiquitin interaction through adaptor proteins, including a direct interaction between ubiquitin and Atg16L1. To ensure that invading pathogens are captured and degraded, Atg16L1 targeting is secured by two backup systems that anchor Atg16L1 to ubiquitin-decorated endosomes. Thus, we reveal that ubiquitin is a pivotal molecule that connects bacteria-containing endosomes with the autophagic machinery upstream of LC3.

SUBMITTER: Fujita N

PROVIDER: S-EPMC3798248 | biostudies-literature | 2013 Oct

REPOSITORIES: biostudies-literature

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Recruitment of the autophagic machinery to endosomes during infection is mediated by ubiquitin.

Fujita Naonobu N Morita Eiji E Itoh Takashi T Tanaka Atsushi A Nakaoka Megumi M Osada Yuki Y Umemoto Tetsuo T Saitoh Tatsuya T Nakatogawa Hitoshi H Kobayashi Shouhei S Haraguchi Tokuko T Guan Jun-Lin JL Iwai Kazuhiro K Tokunaga Fuminori F Saito Kazunobu K Ishibashi Koutaro K Akira Shizuo S Fukuda Mitsunori M Noda Takeshi T Yoshimori Tamotsu T

The Journal of cell biology 20131007 1

Although ubiquitin is thought to be important for the autophagic sequestration of invading bacteria (also called xenophagy), its precise role remains largely enigmatic. Here we determined how ubiquitin is involved in this process. After invasion, ubiquitin is conjugated to host cellular proteins in endosomes that contain Salmonella or transfection reagent-coated latex (polystyrene) beads, which mimic invading bacteria. Ubiquitin is recognized by the autophagic machinery independently of the LC3- ...[more]

PMID: 24100292

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Project description:Mitophagy is a conserved intracellular catabolic process responsible for the selective removal of dysfunctional or superfluous mitochondria to maintain mitochondrial quality and need in cells. Here, we examine the mechanisms of receptor-mediated mitophagy activation, with the focus on BNIP3L/NIX mitophagy receptor, proven to be indispensable for selective removal of mitochondria during the terminal differentiation of reticulocytes. The molecular mechanisms of selecting damaged mitochondria from healthy ones are still very obscure. We investigated BNIP3L dimerization as a potentially novel molecular mechanism underlying BNIP3L-dependent mitophagy. Forming stable homodimers, BNIP3L recruits autophagosomes more robustly than its monomeric form. Amino acid substitutions of key transmembrane residues of BNIP3L, BNIP3LG204A or BNIP3LG208V, led to the abolishment of dimer formation, resulting in the lower LC3A-BNIP3L recognition and subsequently lower mitophagy induction. Moreover, we identified the serine 212 as the main amino acid residue at the C-terminal of BNIP3L, which extends to the intermembrane space, responsible for dimerization. In accordance, the phosphomimetic mutation BNIP3LS212E leads to a complete loss of BNIP3L dimerization. Thus, the interplay between BNIP3L phosphorylation and dimerization indicates that the combined mechanism of LIR phosphorylation and receptor dimerization is needed for proper BNIP3L-dependent mitophagy initiation and progression.Abbreviations: AMBRA1: autophagy and beclin 1 regulator 1; Baf A1: bafilomycin A1; BH3: BCL2 homology 3; BNIP3: BCL2 interacting protein 3; BNIP3L/NIX: BCL2 interacting protein 3 like; CCCP: carbonyl cyanide 3-chlorophenylhydrazone; CoCl2: cobalt (II) chloride; FKBP8: FKBP prolyl isomerase 8; FUNDC1: FUN14 domain containing 1; GABARAP: GABA type A receptor-associated protein; GST: glutathione S-transferase; IMM: inner mitochondrial membrane; LIR: LC3-interacting region; MAP1LC3/LC3: microtubule associated protein 1 light chain 3; OMM: outer mitochondrial membrane; PHB2: prohibitin 2; PI: propidium iodide; PINK1: PTEN induced kinase 1; TM: transmembrane domain; TOMM20: translocase of outer mitochondrial membrane 20.

Project description:Intracellular pathogens have evolved various efficient molecular armaments to subvert innate defenses. Cellular ubiquitination, a normal physiological process to maintain homeostasis, is emerging one such exploited mechanism. Ubiquitin (Ub), a small protein modifier, is conjugated to diverse protein substrates to regulate many functions. Structurally diverse linkages of poly-Ub to target proteins allow enormous functional diversity with specificity being governed by evolutionarily conserved enzymes (E3-Ub ligases). The Ub-binding domain (UBD) and LC3-interacting region (LIR) are critical features of macroautophagy/autophagy receptors that recognize Ub-conjugated on protein substrates. Emerging evidence suggests that E3-Ub ligases unexpectedly protect against intracellular pathogens by tagging poly-Ub on their surfaces and targeting them to phagophores. Two E3-Ub ligases, PRKN and SMURF1, provide immunity against Mycobacterium tuberculosis (M. tb). Both enzymes conjugate K63 and K48-linked poly-Ub to M. tb for successful delivery to phagophores. Intriguingly, M. tb exploits virulence factors to effectively dampen host-directed autophagy utilizing diverse mechanisms. Autophagy receptors contain LIR-motifs that interact with conserved Atg8-family proteins to modulate phagophore biogenesis and fusion to the lysosome. Intracellular pathogens have evolved a vast repertoire of virulence effectors to subdue host-immunity via hijacking the host ubiquitination process. This review highlights the xenophagy-mediated clearance of M. tb involving host E3-Ub ligases and counter-strategy of autophagy inhibition by M. tb using virulence factors. The role of Ub-binding receptors and their mode of autophagy regulation is also explained. We also discuss the co-opting and utilization of the host Ub system by M. tb for its survival and virulence.Abbreviations: APC: anaphase promoting complex/cyclosome; ATG5: autophagy related 5; BCG: bacille Calmette-Guerin; C2: Ca2+-binding motif; CALCOCO2: calcium binding and coiled-coil domain 2; CUE: coupling of ubiquitin conjugation to ER degradation domains; DUB: deubiquitinating enzyme; GABARAP: GABA type A receptor-associated protein; HECT: homologous to the E6-AP carboxyl terminus; IBR: in-between-ring fingers; IFN: interferon; IL1B: interleukin 1 beta; KEAP1: kelch like ECH associated protein 1; LAMP1: lysosomal associated membrane protein 1; LGALS: galectin; LIR: LC3-interacting region; MAPK11/p38: mitogen-activated protein kinase 11; MAP1LC3/LC3: microtubule associated protein 1 light chain 3; MAP3K7/TAK1: mitogen-activated protein kinase kinase kinase 7; MAPK8/JNK: mitogen-activated protein kinase 8; MHC-II: major histocompatibility complex-II; MTOR: mechanistic target of rapamycin kinase; NBR1: NBR1 autophagy cargo receptor; NFKB1/p50: nuclear factor kappa B subunit 1; OPTN: optineurin; PB1: phox and bem 1; PE/PPE: proline-glutamic acid/proline-proline-glutamic acid; PknG: serine/threonine-protein kinase PknG; PRKN: parkin RBR E3 ubiquitin protein ligase; RBR: RING-in between RING; RING: really interesting new gene; RNF166: RING finger protein 166; ROS: reactive oxygen species; SMURF1: SMAD specific E3 ubiquitin protein ligase 1; SQSTM1: sequestosome 1; STING1: stimulator of interferon response cGAMP interactor 1; TAX1BP1: Tax1 binding protein 1; TBK1: TANK binding kinase 1; TNF: tumor necrosis factor; TRAF6: TNF receptor associated factor 6; Ub: ubiquitin; UBA: ubiquitin-associated; UBAN: ubiquitin-binding domain in ABIN proteins and NEMO; UBD: ubiquitin-binding domain; UBL: ubiquitin-like; ULK1: unc-51 like autophagy activating kinase 1.

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Recruitment of the autophagic machinery to endosomes during infection is mediated by ubiquitin.

Publications

Recruitment of the autophagic machinery to endosomes during infection is mediated by ubiquitin.

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