Mammalian protease-activated-receptor-1 and -2 (PAR
1 and PAR
2) are activated by proteases found in the flexible microenvironment of a tumor and play a central role in breast cancer. We propose in the present study that PAR
1 and PAR
2 act together as a functional unit during malignant and physiological invasion processes. This notion is supported by assessing pro-tumor functions in the presence of short hairpin;
shRNA knocked-down
hPar2 or by the use of a truncated PAR
2 devoid of the entire cytoplasmic tail. Silencing of
hPar2 by
shRNA-attenuated thrombin induced PAR
1 signaling as recapitulated by inhibiting the assembly of Etk/Bmx or Akt onto PAR
1-C-tail, by thrombin-instigated colony formation and invasion. Strikingly,
shRNA-
hPar2 also inhibited the TFLLRN selective PAR
1 pro-tumor functions. In addition, while evaluating the physiological invasion process of placenta extravillous trophoblast (EVT) organ culture, we observed inhibition of both thrombin or the selective PAR
1 ligand; TFLLRNPNDK induced EVT invasion by
shRNA-
hPar2 but not by scrambled
shRNA-
hPar2. In parallel, when a truncated PAR
2 was utilized in a xenograft mouse model, it inhibited PAR
1–PAR
2-driven tumor growth in vivo. Similarly, it also attenuated the interaction of Etk/Bmx with the PAR
1-C-tail in vitro and decreased markedly selective PAR
1-induced Matrigel invasion. Confocal images demonstrated co-localization of PAR
1 and PAR
2 in HEK293T cells over-expressing YFP-
hPar2 and HA-
hPar1. Co-immuno-precipitation analyses revealed PAR
1-PAR
2 complex formation but no PAR
1-CXCR4 complex was formed. Taken together, our observations show that PAR
1 and PAR
2 act as a functional unit in tumor development and placenta-uterus interactions. This conclusion may have significant consequences on future breast cancer therapeutic modalities and improved late pregnancy outcome.
相似文献