PAR1‐mediated RhoA activation facilitates CCL2‐induced chemotaxis in PC‐3 cells

Abstract

Patients with advanced prostate cancer often exhibit increased activation of the coagulation system. The key activator of the coagulation cascade is the serine protease thrombin which is capable of eliciting numerous cellular responses. We previously reported that the thrombin receptor PAR1 is overexpressed in prostate cancer. To investigate further the role of PAR1 in prostate cancer metastasis, we examined the effects of thrombin activation on cell adhesion and motility in PC-3 prostate cancer cells. Activation of PAR1-induced dynamic cytoskeletal reorganization and reduced PC-3 binding to collagen I, collagen IV, and laminin (P < 0.01) but not fibronectin. Expression of the cell surface integrin receptors did not change as assessed by flow cytometry. Immunofluorescence microscopy revealed that PAR1 stimulation caused reorganization of the focal adhesions, suggesting that PAR1 activation in PC-3 cells may be modulating cell adhesion through integrin function but not expression. Furthermore, RhoA was activated upon stimulation with thrombin with subsequent cell contraction, decreased cell adhesion, and induced migration towards monocyte chemoattractant protein 1 (MCP-1; CCL2). Thus, it appears that thrombin stimulation plays a role in prostate cancer metastasis by decreasing cell adhesion to the extracellular matrix and positioning the cell in a "ready state" for migration in response to a chemotactic signal. Further exploration is needed to determine whether PAR1 activation affects other signaling pathways involved in prostate cancer.

Prostatic trypsin-like kallikrein-related peptidases (KLKs) and other prostate-expressed tryptic proteinases as regulators of signalling via proteinase-activated receptors (PARs)

“…For example, Liu and colleagues have shown that thrombin increases DU145 adhesion to fibronectin and laminin via a mechanism involving PAR 1 (Liu et al, 2004). In contrast to DU145 cells, treatment of PC-3 cells with a similar concentration of thrombin and PAR 1 AP decreased cell adhesion to collagen I and IV and laminin and resulted in no change in adhesion to fibronectin (Loberg et al, 2007). In addition to cell migration and adhesion, examples of the co-opting of other normal cellular processes essential for cancer progression, such as tissue remodelling, cell growth, angiogenesis and mechanisms regulating apoptosis, have recently been shown to be mediated via PARs.…”

“…Consistently, PAR 1 and PAR 2 APs were able to initiate activation of the cytoskeleton reorganising GTPases RhoA, Rac1 and Cdc42 (Hall, 2005) in a manner similar to thrombin and trypsin in LNCaP cells (Greenberg et al, 2003). Loberg and colleagues have also shown that thrombin treatment of PC-3 cells causes retraction of the actin cytoskeleton and numerous microspike extensions from the cell body (Loberg et al, 2007). This treatment was accompanied by rapid activation of RhoA and Cdc42 with no change in the level of Rac-1 activation (Loberg et al, 2007).…”

“…Additionally, a low concentration of thrombin stimulated PC-3 and DU145 cell migration towards fibronectin (Shi et al, 2004). Interestingly, higher concentrations of thrombin resulted in unchanged cell migration of PC-3 cells on fibronectin and significantly decreased cell migration on plastic, collagen I and IV and laminin (Loberg et al, 2007), while synergistic treatment with PAR 1 and PAR 2 APs enhanced migration similar to thrombin in these cell lines. Interestingly, PAR 1 AP alone stimulated no migration, while PAR 2 AP resulted in only marginal migration.…”

“…Loberg and colleagues have also shown that thrombin treatment of PC-3 cells causes retraction of the actin cytoskeleton and numerous microspike extensions from the cell body (Loberg et al, 2007). This treatment was accompanied by rapid activation of RhoA and Cdc42 with no change in the level of Rac-1 activation (Loberg et al, 2007).…”

Curcumin blocks CCL2-induced adhesion, motility and invasion, in part, through down-regulation of CCL2 expression and proteolytic activity

“…CCL2 is a member of the CC subfamily of low molecular weight chemokines, a class of proteins that are essential in immune regulation, inflammation, and the healing response (7). Under aberrant pathological conditions, CCL2 has essential roles in the infiltration of tumor associated macrophages, which play a key role in increased tumorigenicity of PCa and other cancers (8). CCL2 acts as an autocrine and paracrine mitogenic and motogenic factor of the malignant human androgen-independent prostatic PC-3 cell line (6).…”