bims-nocaut Biomed News
on Non-canonical autophagy
Issue of 2022‒07‒17
four papers selected by
Quentin Frenger
University of Strasbourg

  1. Autophagy. 2022 Jul 09.
      The conjugation of Atg8-family proteins with phospholipids on the double-membrane phagophore is one of the hallmarks of macroautopahgy/autophagy. However, in the past decades, Atg8-family proteins are also found on single-membrane structures, including the phagosome, endosome and lysosome. While the physiological importance of the non-canonical Atg8-family protein conjugation has been demonstrated, the mechanism of this process and the underlying regulation are still not very clear. In a recent paper, Hooper et al. found that during LC3-associated phagocytosis, reactive oxygen species are required for V-ATPase assembly, which is essential for the subsequent LC3 conjugation to the phagosome. Enhanced V-ATPase assembly and the direct engagement of ATG16L1 are also observed in a wide range of non-canonical Atg8-family protein conjugation processes, defining the V-ATPase and ATG16L1 as taking part in a common mechanism.
    Keywords:  ATG16L1; Atg8 conjugation; LAP; ROS; V-ATPase; non-canonical autophagy
  2. J Neuroinflammation. 2022 Jul 14. 19(1): 184
      BACKGROUND: Reactive oxygen species (ROS) often promote acute brain injury after stroke, but their roles in the recovery phase have not been well studied. We tested the hypothesis that ROS activity mediated by NADPH oxidase 2 (NOX2) contributes to acute brain injury but promotes functional recovery during the delayed phase, which is linked with neuroinflammation, autophagy, angiogenesis, and the PI3K/Akt signaling pathway.METHODS: We used the NOX2 inhibitor apocynin to study the role of NOX2 in brain injury and functional recovery in a middle cerebral artery occlusion (MCAO) stroke mouse model. Infarct size, neurological deficits and behavior were evaluated on days 3, 7, 10 and 14 after reperfusion. In addition, dynamic NOX2-induced ROS levels were measured by dihydroethidium (DHE) staining. Autophagy, inflammasomes, and angiogenesis were measured by immunofluorescence staining and western blotting. RNA sequencing was performed, and bioinformatics technology was used to analyze differentially expressed genes (DEGs), as well as the enrichment of biological functions and signaling pathways in ischemia penumbra at 7 days after reperfusion. Then, Akt pathway-related proteins were further evaluated by western blotting.
    RESULTS: Our results showed that apocynin injection attenuated infarct size and mortality 3 days after stroke but promoted mortality and blocked functional recovery from 5 to 14 days after stroke. DHE staining showed that ROS levels were increased at 3 days after reperfusion and then gradually declined in WT mice, and these levels were significantly reduced by the NOX2 inhibitor apocynin. RNA-Seq analysis indicated that apocynin activated the immune response under hypoxic conditions. The immunofluorescence and western blot results demonstrated that apocynin inhibited the NLRP3 inflammasome and promoted angiogenesis at 3 days but promoted the NLRP3 inflammasome and inhibited angiogenesis at 7 and 14 days after stroke, which was mediated by regulating autophagy activation. Furthermore, RNA-Seq and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis indicated that apocynin injection resulted in PI3K-Akt signaling pathway enrichment after 7 days of MCAO. We then used an animal model to show that apocynin decreased the protein levels of phosphorylated PI3K and Akt and NF-κB p65, confirming that the PI3K-Akt-NF-κB pathway is involved in apocynin-mediated activation of inflammation and inhibition of angiogenesis.
    CONCLUSIONS: NOX2-induced ROS production is a double-edged sword that exacerbates brain injury in the acute phase but promotes functional recovery. This effect appears to be achieved by inhibiting NLRP3 inflammasome activation and promoting angiogenesis via autophagy activation.
    Keywords:  Angiogenesis; Autophagy; Ischemic stroke; NADPH; NLRP3 inflammasome; ROS
  3. Front Immunol. 2022 ;13 892239
      MRGPRX2, the human member of the MAS-related G protein coupled receptors (Mrgprs), serves as the cellular target of human mast cells (MCs) for innate ligands, including neuropeptides and antimicrobial peptides. In addition, MRGPRX2 also functions as the receptor for multiple FDA-approved drugs. As such, MRGPRX2 is a mediator of MC responses in neurogenic inflammation, host defense and pseudoallergy. We analyzed the spatiotemporal patterns of MRGPRX2 following its binding of the neuropeptide substance P (SP). Herein, we show that MRGPRX2 internalizes via both endocytosis and macropinocytosis, followed by its distribution between a perinuclear region and the secretory granules (SGs). Further, we show that MRGPRX2-containing macropinosomes undergo resolution by a mechanism that involves dynamin and LC3, giving rise to the incorporation of both LC3 and MRGPRX2 into the SGs. SP then promotes the acidification of the LC3-associated SGs, presumably by stimulating their fusion with lysosomes. Taken together, our results reveal a unique mode of MRGPRX2 trafficking that complements endocytosis and involves macropinocytosis, autophagic machinery-assisted macropinosome resolution and receptor delivery to the SGs.
    Keywords:  MRGPRX2; autophagy; endocytosis; macropinocytosis; mast cell; secretory granules; substance P
  4. Biochim Biophys Acta Mol Basis Dis. 2022 Jul 07. pii: S0925-4439(22)00155-7. [Epub ahead of print] 166484
      Exosomal release pathway and autophagy together maintain homeostasis and survival of cells under stressful conditions. Autophagy is a catabolic process through which cell entities, such as malformed biomacromolecules and damaged organelles, are degraded and recycled via the lysosomal-dependent pathway. Exosomes, a sub-type of extracellular vesicles (EVs) formed by the inward budding of multivesicular bodies (MVBs), are mostly involved in mediating communication between cells. The unfolded protein response (UPR) is an adaptive response that is activated to sustain survival in the cells faced with the endoplasmic reticulum (ER) stress through a complex network that involves protein synthesis, exosomes secretion and autophagy. Disruption of the critical crosstalk between EVs, UPR and autophagy may be implicated in various human diseases, including cancers and neurodegenerative diseases, yet the molecular mechanism(s) behind the coordination of these communication pathways remains obscure. Here, we review the available information on the mechanisms that control autophagy, ER stress and EV pathways, with the view that a better understanding of their crosstalk and balance may improve our knowledge on the pathogenesis and treatment of human diseases, where these pathways are dysregulated.
    Keywords:  Autophagy; Cancer; Crosstalk; ER stress; Exosomes; Extracellular vesicles; Secretory autophagy; Unfolded protein response