Zarrouk, Amira and Nury, Thomas and Dauphin, Aurélien and Frère, Perrine and Riedinger, Jean-Marc and Bachelet, Claude-Marie and Frouin, Frédérique and Moreau, Thibault and Hammami, Mohamed and Kahn, Edmond and Lizard, Gérard (2015) Impact of C24:0 on actin-microtubule interaction in human neuronal SK-N-BE cells: evaluation by FRET confocal spectral imaging microscopy after dual staining with rhodamine-phalloidin and tubulin tracker green. Functional Neurology, 30 (1). pp. 33-46. ISSN 1971-3274
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Abstract
Disorganization of the cytoskeleton of neurons has major consequences on the transport of neuro-transmitters via the microtubule network. The interaction of cytoskeleton proteins (actin and tubulin) was studied in neuronal SK-N-BE cells treated with tetracosanoic acid (C24:0), which is cytotoxic and increased in Alzheimer’s disease patients. When SK-N-BE cells were treated with C24:0, mitochondrial dysfunctions and a non-apoptotic mode of cell death were observed. Fluorescence microscopy revealed shrunken cells with perinuclear condensation of actin and tubulin. After staining with rhodamine-phalloidin and with an antibody raised against α-/β-tubulin, modifications of F-actin and α-/β-tubulin levels were detected by flow cytometry. Lower levels of α-tubulin were found by Western blotting. In C24:0-treated cells, spectral analysis and fluorescence recovery after photo-bleaching (FRAP) measured by confocal microscopy proved the existence of fluorescence resonance energy transfer (FRET) when actin and tubulin were stained with tubulin tracker and rhodamine-phalloidin demonstrating actin and tubulin colocalization/interaction. In control cells, no FRET was observed. Our data demonstrate quantitative changes in actin and tubulin, and modified interactions between actin and tubulin in SK-N-BE cells treated with C24:0. They also show that FRET confocal imaging microscopy is an interesting method for specifying the impact of cytotoxic compounds on cytoskeleton proteins.
| Item Type: | Article |
|---|---|
| Uncontrolled Keywords: | Actin, C24:0, FRET confocal spectral imaging microscopy, microtubule, rhodamine-phalloidin, tubulin tracker |
| Subjects: | 600 Tecnologia - Scienze applicate > 610 Medicina e salute (Classificare qui la tecnologia dei servizi medici) |
| Depositing User: | Marina Spanti |
| Date Deposited: | 20 Sep 2019 13:28 |
| Last Modified: | 20 Sep 2019 13:28 |
| URI: | http://eprints.bice.rm.cnr.it/id/eprint/12535 |
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