Autophagy is a highly conserved intracellular degrading system and its dysfunction is considered related to the cause of neurodegenerative disorders. A previous study showed that the inhibition of ...endocytosis transport attenuates soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) protein transport to lysosomes and block autophagy. The other studies demonstrated oxidative stress, one of the inducers of neurodegenerative diseases inhibits endocytosis transport. Thus, we hypothesized that oxidative stress-induced endocytosis inhibition causes alteration of SNARE protein transport to lysosomes and impairs autophagy. Here, we demonstrated that oxidative stress inhibits endocytosis and decreased the lysosomal localization of VAMP8, one of the autophagy-related SNARE proteins in a human neuroblastoma cell line. Moreover, this oxidative stress induction blocked the autophagosome-lysosome fusion step. Since we also observed decreased lysosomal localization of VAMP8 and inhibition of autophagosome-lysosome fusion in endocytosis inhibitor-treated cells, oxidative stress may inhibit VAMP8 trafficking by suppressing endocytosis and impair autophagy. Our findings suggest that oxidative stress-induced inhibition of VAMP8 trafficking to lysosomes is associated with the development of neurodegenerative diseases due to the blocked autophagosome-lysosome fusion, and may provide a new therapeutic target for restoring the autophagic activity.
Human Neuraminidase 1 and Related Diseases Tsukimoto, Jun; Itoh, Kohji
Trends in Glycoscience and Glycotechnology,
2023/07/25, Letnik:
35, Številka:
206
Journal Article
Recenzirano
Neuraminidase 1 (NEU1) is a lysosomal exo-glycosidase and cleaves glycoside bonds of non-reducing terminal sialic acid. The association of NEU1 with cathepsin A activates NEU1. Overexpression of NEU1 ...in mammalian cells causes self-association and crystallization of NEU1 in rough endoplasmic reticulum (ER). NEU1 deficiency, called sialidosis, is a type of lysosomal storage disease. There is no fundamental treatment for NEU1 deficiency. Gene therapy seems effective, but it is assumed to be dangerous because NEU1 crystallizes intracellularly and damages cells by rupturing biological membranes. In this mini-review, we summarize about functions of NEU1 and CTSA. We also describe NEU1 or CTSA deficiencies.
Neuraminidase 1 (NEU1) is a lysosomal exo-glycosidase and cleaves glycoside bonds of non-reducing terminal sialic acid. The association of NEU1 with cathepsin A activates NEU1. Overexpression of NEU1 ...in mammalian cells causes self-association and crystallization of NEU1 in rough endoplasmic reticulum (ER). NEU1 deficiency, called sialidosis, is a type of lysosomal storage disease. There is no fundamental treatment for NEU1 deficiency. Gene therapy seems effective, but it is assumed to be dangerous because NEU1 crystallizes intracellularly and damages cells by rupturing biological membranes. In this mini-review, we summarize about functions of NEU1 and CTSA. We also describe NEU1 or CTSA deficiencies.
Cation-dependent mannose 6-phosphate receptor (CD-MPR) and cation-independent MPR (CI-MPR) belong to the P-type lectin family. Both intracellular and cell surface MPRs can recognize and bind with the ...terminal mannose 6-phospahte (M6P) residues of N-glycans attached to the mammalian lysosomal enzymes and the related co-factors. Domain9 (Dom9), which is one of the extracytoplasmic region of the CI-MPR, has relatively higher affinity for M6P residues. Here we describe the production of recombinant Dom9-His protein by Pichia pastris, purification, and application as a probe for lectin blotting.
Abstract
Many lysosomal enzymes contain N-glycans carrying mannose 6-phosphate (M6P) residues. Modifying lysosomal enzymes by M6P residues requires a two-step process in the Golgi apparatus. Then the ...lysosomal enzymes with M6P residues are transported from the trans-Golgi network to endosomes and lysosomes by M6P receptors. In insect cells, M6P residues are not added to N-glycans. Therefore, many insect lysosomal enzymes are transported to lysosomes by the M6P-independent pathway. The expression and subcellular distribution of M6P-modifying enzymes were examined by amplifying DNA fragments of M6P-modifying enzymes, generating the corresponding plasmid constructs, and transfection each construct into Sf9 cells, an insect cell line. The human GlcNac-1-phosphotransferase α/β subunit, one of the M6P-modifying enzymes, was found to differ in maturation and localization between mammalian and insect cells. In mammalian cells, newly biosynthesized α/β subunit localized in the cis-Golgi. In Sf9 cells, most of the α/β subunit was localized in the endoplasmic reticulum, and few mature forms of α/β subunit were observed. However, by the co-expression of the human site-1 protease, the mature forms were observed significantly and co-localization with each protein. Our study indicates new insights into regulating the intracellular distribution of the human GlcNac-1-phosphotransferase α/β subunit in insect cells.
Graphical Abstract
Graphical Abstract
Three new farnesylated coumarins, communiferulins A–C (
1
–
3
), and a farnesylated chromone, ferchromone (
4
), were isolated from the roots of an Apiaceous plant
Ferula communis
. Their structures ...including the relative configurations were elucidated by a combination of spectroscopic analyses and calculations of the NMR data. Communiferulins A–C (
1
–
3
) had dihydrofuran rings fused to C-3 and C-4 of their coumarin moieties, while
3
possessed one additional furan ring. HPLC analyses using a chiral column showed
1
–
4
to be racemates, and the absolute configurations of (+)-
1
, (–)-
1
, (+)-
2
, and (–)-
2
were deduced by comparison of their ECD spectra with TDDFT-calculated spectra. Communiferulins A (
1
) and B (
2
), and ferchromone (
4
) showed inhibitory activities on IL-1β production from LPS-stimulated microglial cells.
Six dibenzo-1,4-dioxane derivatives (
1
–
6
) were isolated from the roots of a Hypericaceous plant
Hypericum ascyron
. Spectroscopic analyses revealed
2
and
4
–
6
to be new compounds. The partial ...racemic natures of
1
–
3
were concluded by chiral HPLC analyses, while
5
was confirmed to be a racemate. The absolute configurations
1
–
4
were deduced on the basis of ECD calculations. Biological activity evaluation of the dibenzo-1,4-dioxane derivatives along with two related compounds: hyperdioxanes A (
7
) and B (
8
), previously isolated from the same plant material by our group demonstrated that
7
exhibit an anti-HIV activity (IC
50
5.3 μM, TI 7.2) while
8
showed an inhibitory effect on IL-1β production (inhibition rate: 72.3% at 6.3 μM) from LPS-stimulated microglial cells.
The current work explores intermolecular interactions involved in the lateral propagation of cell‐signaling by epidermal growth factor receptors (EGFRs). Activation of EGFRs by binding an EGF ligand ...in the extracellular domain of the EGFR and subsequent dimerization of the EGFR initiates cell‐signaling. We investigated interactions between EGFRs in living cells by using single‐molecule microscopy, Förster resonance energy transfer (FRET), and atomic force microscopy. By analyzing time‐correlated intensity and propagation trajectories of quantum dot (QD)‐labeled EGFR single molecules, we found that signaling dimers of EGFR (EGF‐EGFR)2 are continuously formed in cell membrane through reversible association of heterodimers EGF(EGFR)2. Also, we found that the lateral propagation of EGFR activation takes place through transient association of a heterodimer with predimers (EGFR)2. We varified the transient association between activated EGFR and predimers using FRET from QD‐labeled heterodimers to Cy5‐labeled predimers and correlated topography and fluorescence imaging. Without extended single‐molecule fluorescence imaging and by using bio‐conjugated QDs, reversible receptor dimerization in the lateral activation of EGFR remained obscured.
Receptors hug repeatedly: Intermolecular interactions among receptors and ligands are essential for cell‐signaling and regular growth and functioning of cells. Single‐molecule fluorescence imaging and spectroscopy of EGFR in cells have shown that reversible dimerization among heterodimers and predimers is essential for the lateral propagation of cell signaling and continuous generation of signaling molecules.
Epidermal growth factor receptor (EGFR), an N-glycosylated transmembrane protein with an intracellular kinase domain, undergoes dimerization by ligand binding resulting in activation of the kinase ...domain and phosphorylation. Ganglioside GM3 containing sialyllactose inhibits the tyrosine kinase activity of EGFR through carbohydrate to carbohydrate interactions (CCI) between N-glycans with GlcNAc termini on EGFR and oligosaccharides on GM3. In this study, we provide further evidence for CCI between EGFR and GM3. (i) In vitro and in situ, the inhibitory effect of GM3 on EGFR tyrosine kinase was much higher in A431 cells upon exposure of the GlcNAc termini of the N-glycans to glycosidase treatment (neuraminidase and β-galactosidase) than in untreated A431 cells. Furthermore, the GM3-mediated inhibition was abrogated by co-incubation with N-glycan containing terminal GlcNAc. (ii) In situ, inhibition of EGFR phosphorylation by GM3 was not observed in α-mannosidase IB (ManIB)-knocked down A431 cells that accumulate high mannose-type N-glycans. (iii) EGFR binding to GM3 was enhanced in glycosidase-treated cells that accumulated GlcNAc termini, whereas GM3 did not bind to EGFR from ManIB-knocked down cells that accumulated high mannose-type N-glycans. These results indicate that GM3-mediated inhibition of EGFR phosphorylation is caused by interaction of GM3 with GlcNAc-terminated N-glycan on EGFR.
Six new limonoids, munropins A−F (1–6), were isolated from the aerial parts of a Chinese medicinal plant, Munronia pinnata (Meliaceae). The structures of 1–6 were assigned by detailed analyses of ...their spectroscopic data. Munropins A (1) and B (2) are limonoids possessing a prieurianin skeleton with α,β-unsaturated γ-lactam moieties at C-17. Munropins C (3), D (4), and E (5) are also prieurianin type limonoids with an α,β-unsaturated γ-lactone moiety, an acetyl group, and an acetoxyacetyl group at C-17, respectively. Munropin F (6) was assigned as a nimbolinin type limonoid with a γ-hydroxy-α,β-unsaturated γ-lactone moiety.
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•A Chinese traditional herbal medicine, Munronia pinnata, was studied.•Prieurianin and nimbolinin type limonoids were isolated from M. pinnata.•Their structures were elucidated by detailed spectroscopic analyses.•Munropins A and B are rare limonoids with γ-lactam moieties at C-17.
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