Abstract A31: Roles of Nogo-B receptor (NgBR) in regulating Ras translocation and modulating breast cancer cell sensitivity to chemotherapeutic drug and irradiation

Abstract

Abstract Ras, a critical oncogene, is one of small GTPase family members. It is active when bound to GTP and inactive when bound to GDP. The activation process needs membrane translocation of Ras because its activators are recruited by membrane receptors, such as EGF receptors. In addition, appropriate localization of activated Ras also is critical for interacting with the downstream effectors. Therefore, regulation of Ras membrane translocation is very important for Ras function. Nogo-B receptor (NgBR) is a type I receptor with a single transmembrane domain. Our previous work has shown that axon guidance gene family Nogo-B and its receptor (NgBR) are essential for chemotaxis and morphogenesis of endothelial cells in vitro and blood vessel formation in Zebrafish. Here, we demonstrate that NgBR can bind and recruit farnesylated Ras to plasma membrane, which results in the activation of PI3K-Akt and Raf-MEK-ERK signaling pathways in human breast cancer cells. Direct interaction between NgBR and farnesylated Ras has been demonstrated by co-immunoprecipitation of MBP-NgBR from incubation of purified recombinant farnesylated Ras and MBP-NgBR protein. Knockdown of NgBR in MDA-MB-231 human breast cancer cells remarkably reduces the amount of membrane-associated Ras. These results suggest that NgBR is a potential modulator for Ras translocation. Knockdown of NgBR expression in MDA-MB-231 breast cancer cells decreases cancer cell colony formation, and abolishes EGF-stimulated Ras activation and phosphorylation of Akt in MDA-MB-231 cells. In addition, overexpression of NgBR caused the transformation of NIH-3T3 cells via increasing Ras plasma membrane translocation and enhancing EGF-stimulated Ras signaling. Knockdown of NgBR decreases the survival and colony formation of MDA-MB-231 cells after irradiation and increases the cisplatin-induced apoptosis and Caspase-3 activity of MDA-MB-231 Cells. These results suggest that NgBR can increase MDA-MB-231 breast cancer cell sensitivity to chemotherapeutic drug and irradiation. Consequently, our preliminary results further demonstrated that inducible knockdown of NgBR in tumor xenograft of MDA-MB-231 cells in nude mice significantly retards tumor growth. Thus, these data demonstrate that NgBR is a unique receptor that recruits farnesylated Ras to the plasma membrane via its hydrophobic cytoplasmic domain and then activates both PI3K-Akt and Raf-MEK-ERK pathways in human breast cancer cells. Collectively, it is the first study to demonstrate that NgBR is a potential therapeutic target for inhibiting Ras-dependent growth of breast cancer and decreasing the sensitivity of breast cancer cells to chemotherapeutic drug and irradiation.