title: Clioquinol induces autophagy in cultured astrocytes and neurons by acting as a zinc ionophore.
abstract: Recent studies have demonstrated that clioquinol, an antibiotic with an anti-amyloid effect, acts as a zinc ionophore under physiological conditions. Because increases in labile zinc may induce autophagy, we examined whether clioquinol induces autophagy in cultured astrocytes in a zinc-dependent manner. Within 1h of exposure to 0.1-10 μM clioquinol, the levels of microtubule-associated protein 1 light chain 3 (LC3)-II, a marker of autophagy, began to increase in astrocytes. Confocal live-cell imaging of GFP-LC3-transfected astrocytes showed the formation of LC3(+) autophagic vacuoles (AVs), providing a further indication that clioquinol induced autophagy. Addition of 3-methyladenine or small-interfering RNA against autophagy-related gene 6 (ATG6/Beclin-1) blocked clioquinol-induced increases in LC3-II. FluoZin-3 fluorescence microscopy showed that, like the zinc ionophore pyrithione, clioquinol increased intracellular zinc levels in the cytosol and AVs in an extracellular zinc-dependent manner. Zinc chelation with N,N,N',N'-tetrakis-(2-pyridylmethyl) ethylenediamine (TPEN) reduced, and addition of zinc increased the levels of LC3-II and LC3(+) puncta, indicating that zinc influx plays a key role therein. Moreover, astrocytes and SH-SY5Y cells expressing mutant huntingtin (mHttQ74) accumulated less aggregates when treated with clioquinol, and this effect was reversed by TPEN. These results indicate that clioquinol-induced autophagy is likely to be physiologically functional. The present study demonstrates that clioquinol induces autophagy in a zinc-dependent manner and contributes to clearance of aggregated proteins in astrocytes and neurons. Hence, in addition to its metal-chelating effect in and around amyloid beta (Aβ) plaques, clioquinol may contribute to the reduction of Aβ loads by activating autophagy by increasing or normalizing intracellular zinc levels in brain cells.
authors.affiliation: Department of Biology, University of Yonsei, College of Life Science and Biotechnology, Seoul 120-749, Republic of Korea.
authors.forename: Mi-Ha
authors.initials: MH
authors.lastname: Park
authors.forename: Sook-Jeong
authors.initials: SJ
authors.lastname: Lee
authors.forename: Hyae-Ran
authors.initials: HR
authors.lastname: Byun
authors.forename: Yunha
authors.initials: Y
authors.lastname: Kim
authors.forename: Young J
authors.initials: YJ
authors.lastname: Oh
authors.forename: Jae-Young
authors.initials: JY
authors.lastname: Koh
authors.forename: Jung Jin
authors.initials: JJ
authors.lastname: Hwang
mesh_terms: D048868:Adaptor Proteins, Signal Transducing / Q000378:metabolism; D000818:Animals; D001253:Astrocytes / Q000187:drug effects* / Q000378:metabolism; D001343:Autophagy / Q000187:drug effects* / Q000502:physiology; D000071190:Autophagy-Related Protein 8 Family; D015153:Blotting, Western; D045744:Cell Line, Tumor; D007464:Clioquinol / Q000494:pharmacology*; D007150:Immunohistochemistry; D007476:Ionophores / Q000494:pharmacology*; D051379:Mice; D008840:Microfilament Proteins / Q000378:metabolism; D018613:Microscopy, Confocal; D009474:Neurons / Q000187:drug effects* / Q000378:metabolism; D015032:Zinc / Q000378:metabolism*
chemical_list: D048868:Adaptor Proteins, Signal Transducing; D000071190:Autophagy-Related Protein 8 Family; C498698:GABARAPL2 protein, human; D007476:Ionophores; D008840:Microfilament Proteins; D007464:Clioquinol; D015032:Zinc