The garlic compound ajoene covalently binds vimentin, disrupts the vimentin network and exerts anti-metastatic activity in cancer cells
Date
2019
Journal Title
Journal ISSN
Volume Title
Publisher
BMC (part of Springer Nature)
Abstract
Background: Garlic has been used for centuries for its flavour and health promoting properties that include
protection against cancer. The vinyl disulfide-sulfoxide ajoene is one of the phytochemicals found in crushed
cloves, hypothesised to act by S-thiolating reactive cysteines in target proteins.
Methods: Using our fluorescently labelled ajoene analogue called dansyl-ajoene, ajoene’s protein targets in MDAMB-
231 breast cancer cells were tagged and separated by 2D electrophoresis. A predominant band was identified
by MALDI-TOF MS/MS to be vimentin. Target validation experiments were performed using pure recombinant
vimentin protein. Computational modelling of vimentin bound to ajoene was performed using Schrödinger and
pKa calculations by Epik software. Cytotoxicity of ajoene in MDA-MB-231 and HeLa cells was measured by the MTT
assay. The vimentin filament network was visualised in ajoene-treated and non-treated cells by
immunofluorescence and vimentin protein expression was determined by immunoblot. The invasion and migration
activity was measured by wound healing and transwell assays using wildtype cells and cells in which the vimentin
protein had been transiently knocked down by siRNA or overexpressed.
Results: The dominant protein tagged by dansyl-ajoene was identified to be the 57 kDa protein vimentin. The
vimentin target was validated to reveal that ajoene and dansyl-ajoene covalently bind to recombinant vimentin via
a disulfide linkage at Cys-328. Computational modelling showed Cys-328 to be exposed at the termini of the
vimentin tetramer. Treatment of MDA-MB-231 or HeLa cells with a non-cytotoxic concentration of ajoene caused
the vimentin filament network to condense; and to increase vimentin protein expression. Ajoene inhibited the
invasion and migration of both cancer cell lines which was found to be dependent on the presence of vimentin.
Vimentin overexpression caused cells to become more migratory, an effect that was completely rescued by ajoene.
Conclusions: The garlic-derived phytochemical ajoene targets and covalently modifies vimentin in cancer cells by
S-thiolating Cys-328. This interaction results in the disruption of the vimentin filament network and contributes to
the anti-metastatic activity of ajoene in cancer cells.
Description
CITATION: Kaschula, C. H., et al. 2019. The garlic compound ajoene covalently
binds vimentin, disrupts the vimentin
network and exerts anti-metastatic activity in cancer cells. BMC Cancer, 19:248, doi:10.1186/s12885-019-5388-8.
The original publication is available at https://bmccancer.biomedcentral.com
The original publication is available at https://bmccancer.biomedcentral.com
Keywords
Polysulfides, Allium sativum, Vimentin
Citation
Kaschula, C. H., et al. 2019. The garlic compound ajoene covalently
binds vimentin, disrupts the vimentin
network and exerts anti-metastatic activity in cancer cells. BMC Cancer, 19:248, doi:10.1186/s12885-019-5388-8