The proto-oncogene c-Src and its downstream signaling pathways are inhibited by the metastasis suppressor, NDRG1

// Wensheng Liu 1, 2 , Fei Yue 1 , Minhua Zheng 1 , Angelica Merlot 2 , Dong-Hun Bae 2 , Michael Huang 2 , Darius Lane 2 , Patric Jansson 2 , Goldie Yuan Lam Liu 2 , Vera Richardson 2 , Sumit Sahni 2 , Danuta Kalinowski 2 , Zaklina Kovacevic 2 , Des. R. Richardson 2 1 Department of General Surgery, Ruijin Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai 200025, P.R.China 2 Molecular Pharmacology and Pathology Program, Department of Pathology and Bosch Institute, Blackburn Building (D06), University of Sydney, Sydney, New South Wales 2006, Australia Correspondence to: Des R. Richardson, e-mail: d.richardson@med.usyd.edu.au Zaklina Kovacevic, e-mail: zaklina.kovacevic@sydney.edu.au Minhua Zheng, e-mail: zmhtiger@yeah.net Keywords: NDRG1, metastasis suppressor, c-Src, cell migration Received: December 20, 2014      Accepted: February 08, 2015      Published: April 10, 2015 ABSTRACT N-myc downstream regulated gene-1 (NDRG1) is a potent metastasis suppressor that plays a key role in regulating signaling pathways involved in mediating cancer cell invasion and migration, including those derived from prostate, colon, etc . However, the mechanisms and molecular targets through which NDRG1 reduces cancer cell invasion and migration, leading to inhibition of cancer metastasis, are not fully elucidated. In this investigation, using NDRG1 over-expression models in three tumor cell-types (namely, DU145, PC3MM and HT29) and also NDRG1 silencing in DU145 and HT29 cells, we reveal that NDRG1 decreases phosphorylation of a key proto-oncogene, cellular Src (c-Src), at a well-characterized activating site (Tyr416). NDRG1-mediated down-regulation of EGFR expression and activation were responsible for the decreased phosphorylation of c-Src (Tyr416). Indeed, NDRG1 prevented recruitment of c-Src to EGFR and c-Src activation. Moreover, NDRG1 suppressed Rac1 activity by modulating phosphorylation of a c-Src downstream effector, p130Cas, and its association with CrkII, which acts as a “molecular switch” to activate Rac1. NDRG1 also affected another signaling molecule involved in modulating Rac1 signaling, c-Abl, which then inhibited CrkII phosphorylation. Silencing NDRG1 increased cell migration relative to the control and inhibition of c-Src signaling using siRNA, or a pharmacological inhibitor (SU6656), prevented this increase. Hence, the role of NDRG1 in decreasing cell migration is, in part, due to its inhibition of c-Src activation. In addition, novel pharmacological agents, which induce NDRG1 expression and are currently under development as anti-metastatic agents, markedly increase NDRG1 and decrease c-Src activation. This study leads to important insights into the mechanism involved in inhibiting metastasis by NDRG1 and how to target these pathways with novel therapeutics.