ironjustice
2013-01-18 23:39:19 UTC
Targeting the Metastasis Suppressor, NDRG1, Using Novel Iron
Chelators: Regulation of Stress Fiber-Mediated Tumor Cell Migration
via Modulation of the ROCK1/pMLC2 Signaling Pathway
Jing Sun, Daohai Zhang, Ying Zheng, Qian Zhao, Minhua Zheng, Zaklina
Kovacevic, and Des R. Richardson
Department of General Surgery, Ruijin Hospital,
Shanghai Jiao Tong University School of Medicine, Shanghai, P.R.China
(J.S., M.Z.); Department of Pathology, University of Sydney, New South
Wales, Australia (D.Z., Z.K., D.R.R.); and Department of
Pathophysiology, Key Laboratory of Cell Differentiation and Apoptosis
of Chinese Ministry of Education, Shanghai Jiao Tong University School
of Medicine, Shanghai, P.R. China (Y.Z., Q.Z.)
Address correspondence to:
Dr. D. R. Richardson, Iron Metabolism and Chelation Program,
Department of Pathology and Bosch Institute, University of Sydney,
Sydney, New South Wales, 2006 Australia.
E-mail: ***@med.usyd.edu.au
J.S. and D.Z. contributed equally as first authors. M.Z., Z.K. and
D.R.R. contributed equally as co-corresponding authors.
Abstract
The iron-regulated metastasis suppressor, N-myc downstream-regulated
gene 1 (NDRG1), is up-regulated by cellular iron depletion mediated by
iron chelators and can inhibit cancer cell migration.
However, the mechanism of how NDRG1 achieves this effect remains
unclear.
In this study, we implemented established and newly constructed NDRG1
overexpression and knockdown models using the DU145, HT29, and HCT116
cancer cell lines to investigate the molecular basis by which NDRG1
exerts its inhibitory effect on cell migration.
Using these models, we demonstrated that NDRG1 overexpression inhibits
cell migration by preventing actin-filament polymerization, stress
fiber assembly and formation. In contrast, NDRG1 knockdown had the
opposite effect. Moreover, we identified that NDRG1 inhibited an
important regulatory pathway mediated by the Rho-associated, coiled-
coil containing protein kinase 1 (ROCK1)/phosphorylated myosin light
chain 2 (pMLC2) pathway that modulates stress fiber assembly. The
phosphorylation of MLC2 is a key process in inducing stress fiber
contraction, and this was shown to be markedly decreased or increased
by NDRG1 overexpression or knockdown, respectively. The mechanism
involved in the inhibition of MLC2 phosphorylation by NDRG1 was
mediated by a significant (P < 0.001) decrease in ROCK1 expression
that is a key kinase involved in MLC2 phosphorylation. Considering
that NDRG1 is up-regulated after cellular iron depletion, novel
thiosemicarbazone iron chelators (e.g., di-2-pyridylketone 4,4-
dimethyl-3-thiosemicarbazone) were demonstrated to inhibit ROCK1/pMLC2-
modulated actin-filament polymerization, stress fiber assembly, and
formation via a mechanism involving NDRG1. These results highlight the
role of the ROCK1/pMLC2 pathway in the NDRG1-mediated antimetastatic
signaling network and the therapeutic potential of iron chelators at
inhibiting metastasis.
Footnotes
The work was supported by the National Health and Medical Research
Council of Australia [Project Grant 632778, Senior Principal Research
Fellowship 571123, Early Career Fellowship 1037323] and the National
Natural Science Foundation of China [Grants 30971488 and 81172521].
The authors declare no conflict of interest.
dx.doi.org/10.1124/mol.112.083097.
↵This article has supplemental material available at
molpharm.aspetjournals.org.
Received October 21, 2012.
Accepted November 27, 2012.
Copyright © 2013 by The American Society for Pharmacology and
Experimental Therapeutics
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Chelators: Regulation of Stress Fiber-Mediated Tumor Cell Migration
via Modulation of the ROCK1/pMLC2 Signaling Pathway
Jing Sun, Daohai Zhang, Ying Zheng, Qian Zhao, Minhua Zheng, Zaklina
Kovacevic, and Des R. Richardson
Department of General Surgery, Ruijin Hospital,
Shanghai Jiao Tong University School of Medicine, Shanghai, P.R.China
(J.S., M.Z.); Department of Pathology, University of Sydney, New South
Wales, Australia (D.Z., Z.K., D.R.R.); and Department of
Pathophysiology, Key Laboratory of Cell Differentiation and Apoptosis
of Chinese Ministry of Education, Shanghai Jiao Tong University School
of Medicine, Shanghai, P.R. China (Y.Z., Q.Z.)
Address correspondence to:
Dr. D. R. Richardson, Iron Metabolism and Chelation Program,
Department of Pathology and Bosch Institute, University of Sydney,
Sydney, New South Wales, 2006 Australia.
E-mail: ***@med.usyd.edu.au
J.S. and D.Z. contributed equally as first authors. M.Z., Z.K. and
D.R.R. contributed equally as co-corresponding authors.
Abstract
The iron-regulated metastasis suppressor, N-myc downstream-regulated
gene 1 (NDRG1), is up-regulated by cellular iron depletion mediated by
iron chelators and can inhibit cancer cell migration.
However, the mechanism of how NDRG1 achieves this effect remains
unclear.
In this study, we implemented established and newly constructed NDRG1
overexpression and knockdown models using the DU145, HT29, and HCT116
cancer cell lines to investigate the molecular basis by which NDRG1
exerts its inhibitory effect on cell migration.
Using these models, we demonstrated that NDRG1 overexpression inhibits
cell migration by preventing actin-filament polymerization, stress
fiber assembly and formation. In contrast, NDRG1 knockdown had the
opposite effect. Moreover, we identified that NDRG1 inhibited an
important regulatory pathway mediated by the Rho-associated, coiled-
coil containing protein kinase 1 (ROCK1)/phosphorylated myosin light
chain 2 (pMLC2) pathway that modulates stress fiber assembly. The
phosphorylation of MLC2 is a key process in inducing stress fiber
contraction, and this was shown to be markedly decreased or increased
by NDRG1 overexpression or knockdown, respectively. The mechanism
involved in the inhibition of MLC2 phosphorylation by NDRG1 was
mediated by a significant (P < 0.001) decrease in ROCK1 expression
that is a key kinase involved in MLC2 phosphorylation. Considering
that NDRG1 is up-regulated after cellular iron depletion, novel
thiosemicarbazone iron chelators (e.g., di-2-pyridylketone 4,4-
dimethyl-3-thiosemicarbazone) were demonstrated to inhibit ROCK1/pMLC2-
modulated actin-filament polymerization, stress fiber assembly, and
formation via a mechanism involving NDRG1. These results highlight the
role of the ROCK1/pMLC2 pathway in the NDRG1-mediated antimetastatic
signaling network and the therapeutic potential of iron chelators at
inhibiting metastasis.
Footnotes
The work was supported by the National Health and Medical Research
Council of Australia [Project Grant 632778, Senior Principal Research
Fellowship 571123, Early Career Fellowship 1037323] and the National
Natural Science Foundation of China [Grants 30971488 and 81172521].
The authors declare no conflict of interest.
dx.doi.org/10.1124/mol.112.083097.
↵This article has supplemental material available at
molpharm.aspetjournals.org.
Received October 21, 2012.
Accepted November 27, 2012.
Copyright © 2013 by The American Society for Pharmacology and
Experimental Therapeutics
Who loves ya.
Tom
Jesus Was A Vegetarian!
http://tinyurl.com/2r2nkh
Man Is A Herbivore!
http://tinyurl.com/4rq595
DEAD PEOPLE WALKING
http://tinyurl.com/zk9fk