Differential roles of ERK and Akt pathways in regulation of EGFR-mediated signaling and motility in prostate cancer cells

Gan, Yunhua; Shi, Changcheng; Inge, Landon J.; Hibner, M.; Balducci, James; Huang, Yao

doi:10.1038/onc.2010.240

Cited by 247 publications

(224 citation statements)

References 46 publications

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“…Other epithelial markers, ZO-1 and occluding, were not detected, whereas cytokeratin-18 was unchanged in either cell line, indicating that Ecad is the principal epithelial marker downregulated during EMT process in prostate cells. This finding is consistent with previous studies (45)(46)(47)(48).…”

Section: Discussionsupporting

confidence: 83%

Involvement of O-glycosylation defining oncofetal fibronectin in epithelial-mesenchymal transition process

Freire-de-Lima

Gelfenbeyn

Ding

et al. 2011

Proc. Natl. Acad. Sci. U.S.A.

112

126

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The process termed "epithelial-mesenchymal transition" (EMT) was originally discovered in ontogenic development, and has been shown to be one of the key steps in tumor cell progression and metastasis. Recently, we showed that the expression of some glycosphingolipids (GSLs) is down-regulated during EMT in human and mouse cell lines. Here, we demonstrate the involvement of GalNAc-type (or mucintype) O-glycosylation in EMT process, induced with transforming growth factor β (TGF-β) in human prostate epithelial cell lines. We found that: (i) TGF-β treatment caused up-regulation of oncofetal fibronectin (onfFN), which is defined by mAb FDC6, and expressed in cancer or fetal cells/tissues, but not in normal adult cells/tissues. The reactivity of mAb FDC6 requires the addition of an O-glycan at a specific threonine, inside the type III homology connective segment (IIICS) domain of FN. (ii) This change is associated with typical EMT characteristics; i.e., change from epithelial to fibroblastic morphology, enhanced cell motility, decreased expression of a typical epithelial cell marker, E-cadherin, and enhanced expression of mesenchymal markers. (iii) TGF-β treatment up-regulated mRNA level of FN containing the IIICS domain and GalNAc-T activity for the IIICS domain peptide substrate containing the FDC6 onfFN epitope. (iv) Knockdown of GalNAc-T6 and T3 inhibited TGF-β-induced up-regulation of onfFN and EMT process. (v) Involvement of GSLs was not detectable with the EMT process in these cell lines. These findings indicate the important functional role of expression of onfFN, defined by sitespecific O-glycosylation at IIICS domain, in the EMT process.O-glycosylated fibronectin | siRNA G lycoconjugates, such as glycosphingolipids (GSLs) and N-and O-linked glycoproteins, have been shown to play important roles in embryogenesis (1), tumorigenesis, and cancer progression (2). Specific types of glycosyl residues modulate particular signaling pathways and regulate cell phenotypes. For example, GM3 inhibits epithelial growth factor receptor (EGFR) activation (3, 4); GM2 associated with tetraspanine CD82 inhibits the activation of the hepatocyte growth factor receptor cMet (5); and O-Fucose glycans modulate Notch signaling, which controls the fate of many cell types (6-8).Epithelial-mesenchymal transition (EMT) was initially observed during early embryonic development and organ formation (9-11). Accumulating evidence has shown that the EMT process plays a key role in disease development, particularly in cancer progression to metastasis (10-14) and fibrosis (15). During EMT, cells lose their apical-basal polarity, change morphology to fibroblastic, display reduced expression of epithelial cell marker molecules such as E-cadherin (Ecad), and enhance expression of mesenchymal cell marker molecules such as fibronectin (FN), N-cadherin (Ncad), vimentin, and matrix-metalloproteinases (MMPs). When combined, these effects result in increased cell motility (10-15).Our recent studies found a reduction in specific GSLs (Gg4 and/ or GM2) in ...

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Section: Discussionsupporting

confidence: 83%

Involvement of O-glycosylation defining oncofetal fibronectin in epithelial-mesenchymal transition process

Freire-de-Lima

Gelfenbeyn

Ding

et al. 2011

Proc. Natl. Acad. Sci. U.S.A.

112

126

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“…Defective endocytosis of EGFR can be caused by altered ubiquitination (117,118), altered cytoskeletal interactions (119), and derailed endocytosis. All of these causes of EGFR dysfunction can be associated with oncogenic alterations (120).…”

Section: Outlook Of High-resolution 3d Tracking Of Egfrmentioning

confidence: 99%

Segmentation of 3D Trajectories Acquired by TSUNAMI Microscope: An Application to EGFR Trafficking

Liu

Perillo

Liu

et al. 2016

Biophysical Journal

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Whereas important discoveries made by single-particle tracking have changed our view of the plasma membrane organization and motor protein dynamics in the past three decades, experimental studies of intracellular processes using singleparticle tracking are rather scarce because of the lack of three-dimensional (3D) tracking capacity. In this study we use a newly developed 3D single-particle tracking method termed TSUNAMI (Tracking of Single particles Using Nonlinear And Multiplexed Illumination) to investigate epidermal growth factor receptor (EGFR) trafficking dynamics in live cells at 16/43 nm (xy/z) spatial resolution, with track duration ranging from 2 to 10 min and vertical tracking depth up to tens of microns. To analyze the long 3D trajectories generated by the TSUNAMI microscope, we developed a trajectory analysis algorithm, which reaches 81% segment classification accuracy in control experiments (termed simulated movement experiments). When analyzing 95 EGF-stimulated EGFR trajectories acquired in live skin cancer cells, we find that these trajectories can be separated into three groups-immobilization (24.2%), membrane diffusion only (51.6%), and transport from membrane to cytoplasm (24.2%). When EGFRs are membrane-bound, they show an interchange of Brownian diffusion and confined diffusion. When EGFRs are internalized, transitions from confined diffusion to directed diffusion and from directed diffusion back to confined diffusion are clearly seen. This observation agrees well with the model of clathrin-mediated endocytosis.

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“…The phosphatidylinositol 3-kinase/AKT pathway is a downstream mediator of EGFR signaling (31). Protein immunoblotting of H460-AGR2KD and JH-EsoAd1-AGR2KD cell lysates revealed a 3.5-and 2.3-fold reduction, respectively, in AKT protein phosphorylation after AGR2 expression was reduced by RNA interference (Fig.…”

Section: Agr2 Expression Stimulates Egf Receptor Signaling-areg Protementioning

confidence: 99%

The Human Adenocarcinoma-associated Gene, AGR2, Induces Expression of Amphiregulin through Hippo Pathway Co-activator YAP1 Activation

Dong¹,

Gupta²,

Pai³

et al. 2011

Journal of Biological Chemistry

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Anterior Gradient Homolog 2 (AGR2) is expressed by the normal intestine and by most human adenocarcinomas, including those derived from the esophagus, pancreas, lung, breast, ovary, and prostate. Xenografts of human adenocarcinoma cell lines in nude mice previously demonstrated that AGR2 supports tumor growth. In addition, AGR2 is able to induce in vitro a transformed phenotype in fibroblast and epithelial cell lines. The mechanism underlying the growth promoting effects of AGR2 is unknown. The present study shows that AGR2 induces expression of amphiregulin (AREG), a growth promoting EGFR ligand. Induced AREG expression in adenocarcinoma cells is able to rescue the transformed phenotype that is lost when AGR2 expression is reduced. Additional experiments demonstrate that AGR2 induction of AREG is mediated by activation of the Hippo signaling pathway co-activator, YAP1. Thus AGR2 promotes growth by regulating the Hippo and EGF receptor signaling pathways. Anterior Gradient Homolog 2 (AGR2)2 encodes a 17 kDa protein that is highly conserved in vertebrates. AGR2 was first described in Xenopus laevis, where its expression is responsible for the development of a glandular organ called the cement gland (1). A significant role in tissue regeneration was established for AGR2 in salamanders where it functions in nerve-dependent limb regeneration (2). AGR2 is also expressed by secretory cells in the normal murine intestine (3). In humans, enhanced AGR2 expression was first described in breast cancer, which was followed by similar observations in most human adenocarcinomas, including those derived from the esophagus, pancreas, lung, ovary, and prostate (4 -11). Both in vitro and in vivo studies demonstrated that AGR2 promotes tumor growth and metastasis (3,6,12). In adenocarcinoma cell lines and nontransformed fibroblasts, AGR2 induces cell proliferation and anchorage-independent growth in soft agar. Human adenocarcinoma cell lines grown in vivo as mouse xenografts result in smaller tumors when AGR2 expression is reduced (3, 6). In vitro studies examining cell migration suggested that AGR2 may function in a non-cell autonomous fashion (3).The mechanisms responsible for AGR2 effects on growth and transformation are unknown. AGR2 expression in humans is restricted to epithelial cells, for which the EGF signaling pathway serves a regulatory role in controlling cell growth. The present study tested the hypothesis that AGR2 affects cell signaling, and potentially that of the EGFR pathway, which has established significance in epithelial cancers. EXPERIMENTAL PROCEDURESCell Lines-H460 lung adenocarcinoma cells obtained from Dr. David Beer (University of Michigan, Ann Arbor, MI) were previously known as SEG-1 and was the focus of a previous publication (3). A recent report revealed that SEG-1 cells are actually H460 lung adenocarcinoma cells (13). The H460 cells used in this study were reassessed by Winand Dinjens, Erasmus Medical Center, Rotterdam, the Netherlands using the Powerplex 16 TM system (Promega Corp., Madison, WI) to...

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Differential roles of ERK and Akt pathways in regulation of EGFR-mediated signaling and motility in prostate cancer cells

Cited by 247 publications

References 46 publications

Involvement of O-glycosylation defining oncofetal fibronectin in epithelial-mesenchymal transition process

Involvement of O-glycosylation defining oncofetal fibronectin in epithelial-mesenchymal transition process

Segmentation of 3D Trajectories Acquired by TSUNAMI Microscope: An Application to EGFR Trafficking

The Human Adenocarcinoma-associated Gene, AGR2, Induces Expression of Amphiregulin through Hippo Pathway Co-activator YAP1 Activation

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