Copper influx transporter 1 is required for FGF, PDGF and EGF-induced MAPK signaling

Abstract

Copper transporter 1 (CTR1) is the major copper (Cu) influx transporter in mammalian cells. We report here that CTR1 is required for the activation of signaling to the MAPK pathway by the ligands of three major receptor tyrosine kinases (RTK) including FGF, PDGF and EGF. Induction of Erk1/2 phosphorylation was compared in isogenic wild type CTR1+/+ and CTR1−/− cells. Whereas all three ligands increased pErk1/2 in the CTR1+/+ cells, they failed to do this in CTR1−/− cells. While FGF did not enhance the phosphorylation of AKT in the CTR1+/+ cells, both PDGF and EGF increased pAKT in the CTR1+/+ but not CTR1−/− cells. The deficit in Erk1/2 phosphorylation in the CTR1−/− cells was rescued by adding Cu to the medium, and it was induced in CTR1+/+ cells by treatment with a Cu chelator. Intracellular Cu availability was reduced in the CTR1−/− cells as reflected by increased expression of the Cu chaperone CCS. The failure of RTK-induced signaling to both Erk1/2 and AKT suggested the presence of a Cu-dependent step upstream of Ras. The Cu-dependent enzyme SOD1 is responsible for generating the hydrogen peroxide in response to RTK activation that serves to inhibit phosphatases that normally limit RTK signaling. SOD1 activity was reduced by a factor of 17-fold in the CTR1−/− cells, and addition of hydrogen peroxide restored signaling. We conclude that Cu acquired from CTR1 is required for signaling in pathways regulated by RTKs that play major roles in development and cancer.

Transplantation of Neural Stem Cells That Overexpress Sod1 Enhances Amelioration of Intracerebral Hemorrhage in Mice

“…Overexpression of Akt1 enhanced survival of grafted human NSCs transplanted into mouse brains with ICH induced by striatal collagenase injection, as demonstrated by Lee et al 10 Reactive oxygen species contributed to enhanced Akt activity in NSCs, 32,33 and an increase in Akt activity was induced by intracellular H 2 O 2 converted from superoxide by SOD1. 36 Although pAkt expression increased in the SOD1 Tg NSCs compared with the WT NSCs after exposure to H 2 O 2 , there was no difference in pAkt expression between SOD1 Tg NSCs and WT NSCs after exposure to Hb in this study. An earlier report by our group showed that cortical neurons that overexpress SOD1 had a tolerance against ischemic insults via upregulation of Akt signaling.…”

Molecular modulation of the copper and cisplatin transport function of CTR1 and its interaction with IRS-4

“…After immunoprecipitation of CTR1 from the HEK293 FLP-In T-Rex cells with anti-FLAG antibody, crosslinked binding partners were selectively released by reduction with β-mercaptoethanol which resulted in an extremely efficient recovery with exceptionally low background. Proteomic analysis of the released trypsin-digested proteins identified insulin receptor substrate 4 (IRS-4) and this was considered of interest because IRS proteins mediate a variety of biological effects downstream of the insulin and insulin-like growth factor 1 (IGF-1) receptors, and we had previously demonstrated that CTR1 plays an important role in regulating signaling from receptor tyrosine kinases [36]. The IRS-4 peptides identified by mass spectrometry are presented in Table 1.…”

“…However, this domain contains several motifs that are of interest to a potential scaffold role of CTR1 that facilitates the binding of other proteins. In Xenopus CTR1 is important for signaling during embryonic development [37] and recent studies have shown that it is essential for signaling from multiple receptor tyrosine kinases in mouse cells [36,38]. Molecular modeling suggests that the intracellular loop is a likely site for the docking of proteins that participate in this pathway [12].…”

A Re-Evaluation of the Role of hCTR1, the Human High-Affinity Copper Transporter, in Platinum-Drug Entry into Human Cells

“…Inhibition of proteasomal activity enhances delivery of cDDP in ovarian carcinoma cells, and promises significant clinical potential, but its basis is also not yet clear (Jandial et al, 2009). The recent observation that copper is involved in growth factor kinase cycling in tumor cells introduces a new complexity (Tsai et al, 2012), and recent studies suggest important interactions between ATP7A, hCTR1, Atox1, and the platelet-derived growth factor receptor in cell activation (Ashino et al, 2010). (A) cDDP-sensitive (A2780) cells were transiently transfected with siRNA duplexes against CTR1 or siRNA negative control in Opti-MEM medium.…”