A Structural View of Xenophagy, a Battle between Host and Microbes

  • Received : 2017.10.25
  • Accepted : 2017.11.10
  • Published : 2018.01.31


The cytoplasm in mammalian cells is a battlefield between the host and invading microbes. Both the living organisms have evolved unique strategies for their survival. The host utilizes a specialized autophagy system, xenophagy, for the clearance of invading pathogens, whereas bacteria secrete proteins to defend and escape from the host xenophagy. Several molecules have been identified and their structural investigation has enabled the comprehension of these mechanisms at the molecular level. In this review, we focus on one example of host autophagy and the other of bacterial defense: the autophagy receptor, NDP52, in conjunction with the sugar receptor, galectin-8, plays a critical role in targeting the autophagy machinery against Salmonella; and the cysteine protease, RavZ secreted by Legionella pneumophila cleaves the LC3-PE on the phagophore membrane. The structure-function relationships of these two examples and the directions of future research will be discussed.

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Fig. 1. Overview of xenophagy. Bacteria invade mammalian cells and the carbohydrates originally exposed to the outside the cells arenow towards the inside of the vacuoles (or phagosomes). Bacteria secrete effectors, such as Eis from Mycobacterium tuberculosis, ede-ma factor toxin from Bacillus anthracis, and cholera toxin from Vibrio cholerae, to modulate the induction of the host cell autophagysignaling. Some bacteria escape from the vacuoles and are ubiquitylated by host E3 Ub-ligases, such as LRSAM1, PARKIN, and theLUBAC complex. Most of the bacteria are restricted inside the vacuoles; however, bacterial division ultimately causes the rupture of thephagosomal membrane and, subsequently, the carbohydrates are now exposed to the cytoplasmic space. This acts as a danger signal tothe cells and the carbohydrates are recognized by the sugar receptor GAL8, which immediately recruits the autophagy receptor NDP52.This recognition step by the autophagy system is inhibited by bacterial proteins, such as RavZ from Legionella pneumophila, IcsB fromShigella flexneri, and ActA and internalin K (InlK) from Listeria monocytogenes. RavZ cleaves the LC3-PE molecule, leading to the com-plete inactivation and significant damage to host autophagy. The fusion step between the autophagosome and lysosome for autolyso-somes production is also blocked by VacA from Helicobacter pylori and ESAT-6 (early secreted antigenic target of 6 kDa) from Mycobac-terium tuberculosis. In practice, each xenophagy step is much more detailed and there are many different pathways involved in differentbacteria; these cannot be included in this simplified version.

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Fig. 2. Structure-function relationship of the interaction between galectin-8 and NDP52 for clearing invading Salmonella. (A) Domainstructure of NDP52 and the interacting proteins, LC3C, GAL8, and Ub. SKICH (skeletal muscle and kidney-enriched inositol phosphatasecarboxyl homology), CLIR (non-canonical LC3-interaction region), CC (coiled coil), GALBI (galectin-8 binding), and UBZ (ubiquitin-binding zinc finger) domains are colored light brown, orange, green, yellow, and purple, respectively. The interacting LC3C, GAL8, andUb are colored sky blue, salmon, and gray, respectively. (B) The structures of each domain complexed with the interacting partner. Thecolor scheme is the same as for panel (A). The atomic resolution structure of the central CC domain of NDP52 is not available, but thehomodimer forms parallel to CC were revealed by ACCORD and SAXS experiments (Kim et al., 2017). (C) A schematic model for Sal-monella clearance in collaboration between NDP52 and GAL8. The structures and colors for the molecules are the same as panels (A)and (B). The SCV (Salmonella-containing vacuole) is ruptured and the carbohydrates are now exposed to the cytoplasm in mammaliancells. The sugar receptor GAL8 recognizes the carbohydrate by using N-CRD and, simultaneously, the C-CRD of GAL8 binds to theGALBI region of NDP52. The parallel CC dimer of NDP52 is critical for the proper orientation of this bridging molecule. In homodimericNDP52, both GALBI regions point towards SCV and both LIR motifs orient towards LC3-anchored phagophore with proper spacingdefined by the length of CC. Then, the phagophore membrane engulfs the Salmonella to complete the autophagosome.

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Fig. 3. Proposed mechanisms of LC3 deconjugation by RavZ from Legionella pneumophila. (A) The domains and overall structure of RavZ.The N- and C-terminal regions containing LIR motifs (N-LIR1/2 colored light brown and C-LIR red) are invisible in electron density map.The catalytic (CAT: residues 49?325) and membrane targeting (MT: residues 326?423) domains are colored blue and orange, respec-tively. (B) The competition between RavZ and cysteine protease ATG4B for the same substrate, LC3B, although the cleavage sites aredifferent. RavZ cleaves the peptide bond between phenylalanine and glycine, whereas ATG4B cleaves the bond between C-terminalglycine and the lipid, phosphatidylethanolamine (PE). (C) The ‘Tethering and Cut’ model. RavZ is targeted to the phagophore membraneand interacts with the LC3-PE molecules anchored in the membrane. By using N- and C-terminal flexible LIR motifs, RavZ is tethered onthe membrane and subsequently cuts the specific peptide bond on the LC3-PE molecule. Currently, it is unclear whether RavZ cleavesone of the tethered LC3 molecules or other nearby LC3 molecules. (D) The ‘Lift and Cut’ model. The targeting of RavZ might be thesame as panel (C). The α3-helix (colored pink) of RavZ docks on the membrane and lifts a LC3-PE molecule via conformational change,after which the LC3-PE is cleaved at the active site of the catalytic domain of RavZ.


Supported by : National Research Foundation of Korea (NRF), Samsung Science & Technology Foundation


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