We have previously reported that high extracellular Ca2+ stimulates
parathyroid hormone-related protein (
PTHrP) release from human prostate and
breast cancer cell lines as well as from H-500 rat Leydig
cancer cells, an action mediated by the
calcium-sensing receptor (CaR). Activating the CaR leads to phosphorylation of
mitogen-activated protein kinases (MAPKs) that participate in
PTHrP synthesis and secretion. Because the CaR is a
G protein-coupled receptor (GPCR), it is likely to transactivate the
epidermal growth factor receptor (EGFR) or the
platelet-derived growth factor receptor (PDGFR). In this study, we hypothesized that activation of the CaR transactivates the EGFR or PDGFR, and examined whether transactivation affects
PTHrP secretion in PC-3 human
prostate cancer cells. Using Western analysis, we observed that an increase in extracellular Ca2+ resulted in delayed activation of
extracellular signal-regulated kinase (ERK) in PC-3 cells. Pre-incubation with
AG1478 (an EGFR
kinase inhibitor) or an EGFR
neutralizing antibody inhibited the high Ca2+ -induced phosphorylation of ERK1/2.
GM6001, a pan
matrix metalloproteinase (
MMP) inhibitor, also partially suppressed the ERK activation, but
AG1296 (a PDGFR
kinase inhibitor) did not. High extracellular Ca2+ stimulates
PTHrP release during a 6-h incubation (1.5- to 2.5- and 3- to 4-fold increases in 3.0 and 7.5 mM Ca2+, respectively). When cells were preincubated with
AG1478,
GM6001, or an antihuman
heparin-binding
EGF (
HB-EGF) antibody,
PTHrP secretion was significantly inhibited under basal as well as high Ca2+ conditions, while
AG1296 had no effect on
PTHrP secretion. Taken together, these findings indicate that activation of the CaR transactivates the EGFR, but not the PDGFR, leading to phosphorylation of ERK1/2 and resultant
PTHrP secretion, although CaR-EGFR-ERK might not be the only signaling pathway for
PTHrP secretion. This transactivation is most likely mediated by activation of
MMP and cleavage of proheparin-binding
EGF (proHB-
EGF) to
HB-EGF.