Amphiregulin-EGFR signaling regulates PTHrP gene expression in breast cancer cells

Abstract

Parathyroid hormone-related protein (PTHrP) is an autocrine/paracrine factor produced by breast cancer cells that is speculated to play a major role in permitting breast cancer cells to grow into the bone microenvironment by stimulating the bone resorption axis. It has been previously shown that EGFR signaling induces the production of PTHrP in several primary and transformed epithelial cell types. Therefore, we investigated the relationship between EGFR and PTHrP gene expression in human breast cancer cells. Of a panel of 7 breast epithelial and cancer cell lines, the osteolytic, EGFRpositive lines (MDA-MB-231 and NS2T2A1) exhibited higher levels of PTHrP transcript expression. Amphiregulin mRNA levels in all lines were approximately 2 orders of magnitude higher than those of TGFα or HBEGF. In the EGFR bearing lines, the receptor was phosphorylated at tyrosine 992 under basal conditions, and the addition of 100 nM amphiregulin did not lead to the phosphorylation of other tyrosine residues typically phosphorylated by the prototypical ligand EGF. Treatment of the EGFR positive lines with the EGFR inhibitor PD153035 and amphiregulin-neutralizing antibodies reduced PTHrP mRNA levels by 50-70%. Stable EGFR expression in the MCF7 line failed to increase basal PTHrP mRNA levels; however, treatment of this cell line with exogenous EGF or amphiregulin increased PTHrP transcription 3-fold. Transient transfection analysis suggests that the MAPK pathway and ETS transcription factors mediate EGFR coupling to PTHrP gene expression. Taken together, it appears that autocrine stimulation of EGFR signaling by amphiregulin is coupled to PTHrP gene expression via EGFR Tyr992 and MAPK, and that this pathway may contribute to PTHrP expression by breast tumor cells.

Tissue remodeling induced by hypersecreted epidermal growth factor and amphiregulin in the airway after an acute asthma attack

“…22 Second, differences in the sites of ligand-induced EGFR tyrosine phosphorylation may underlie the divergent ligand-induced EGFR coupling to signaling effectors and biological responses. EGF abundantly stimulated EGFR phosphorylation at Tyr1045, whereas amphiregulin did not, which is in agreement with the observations of Gilmore et al 23 Phosphorylation of Tyr1045 creates a canonical binding site for the E3 ubiquitin ligase c-cbl, leading to EGFR ubiquitination and degradation by the 26S proteasome. 24,25 Also in our experiments, whereas EGF abundantly stimulated EGFR phosphorylation at Tyr992 in NHBEs, amphiregulin did not.…”

Differential Expression of Parathyroid Hormone–Related Protein in Adrenocortical Tumors: Autocrine/Paracrine Effects on the Growth and Signaling Pathways in H295R Cells

“…The experiments on H295R cells show that PTHrP is a key regulator of important biological activities, such as proliferation, cell cycle dysregulation, and apoptosis. PTHrP uses two signaling pathways, intracellular cAMP/PKA and calcium/PLC in H295R cells that can be involved in the mitogenic effects of PTHrP because they are largely dependent on mitogen-activated protein kinase, whose activity can be modulated by both PKA and protein kinase C (45,46). Further studies are needed to characterize the cascade of intracellular proteins activated by PTHrP in ACT cells leading to the activation of mitogen-activated protein kinase that in turn activates early genes involved in cell proliferation and differentiation (46,47).…”

EGFR-Ligand Signaling in Breast Cancer Metastasis: Recurring Developmental Themes

“…More recent studies have focused on the distinct downstream signaling and cellular behavior induced by AREG. Unlike exogenous EGF treatment, AREG stimulation of model cell lines and breast cancer cell lines is unable to induce efficient phosphorylation of many of the tyrosine residues in the C-terminal tail of the EGFR (22,43,51,52) and (Fig. 1).…”