Investigation Of The Differential Proteomics On Plasma Membrane With Metastasis Of Prostate Cancer Cell Mat-LyLu Mediated By Toxin Probes

Metastasis is the primary factors contributing to deaths in patients with cancer.According to statistics,more than 90%of all cancer patients died of tumor metastasis rather than primary tumor,and the regulation of tumor metastasis is influenced by many factors.Voltage-gate sodium channel subtype Nav1.7 has been found to be highly expressed in the prostate cancer cell line Mat-LyLu and modulate its matastasis,however,the mechanism underlying this process is not clarified-so far.Two peptide toxins JZTX-I and HNTX-?,which acting on Nav1.7,were isolated and identified from the venom of the Chinese spider Chilobrachys jingzhao and Selenocosmia hainana respectively.Previous studies showed that these two kinds of peptide toxins have an opposite effect on Nav1.7.JZTX-I enhanced the metastatic activity of Mat-LyLu cells by inhibiting the inactivation of Nav1.7 and then increasing sodium ions influx,while HNTX-? decreased the metastatic activity of Mat-LyLu cells by inhibiting the activation of Nav1.7 and then reducing sodium ions influx.Transwell migration and invasion assays indicated that the migratory capacity of Mat-LyLu cells increased by 50.2%and the invasive capacity increased by 19%after 5?M JZTX-I treatment for 24h.Whereas treated Mat-LyLu cells with 5?M HNTX-? for 24h,the migratory capacity decreased by 44%and the invasive capacity decreased by 60.1%.In this study,JZTX-? and HNTX-? were used as molecular probes to study the differential expression of plasma membrane proteins during the matastatic process of Mat-LyLu cells.2-DE was used to separate the purified plasma membrane proteins,and we found 108 differentially expressed proteins after ImageMaster analysis.Of these,29 was found in JZTX-I treated group and 79 in HNTX-? treated group.MALDI-TOF/TOF was then used to determine the identified proteins,and 64 proteins were obtain,including cell adhesion proteins(Muskelin),tumor associated proteins(Moesin,Fascin)and energy metabolism related proteins(Isocitrate dehydrogenase,Phosphoglycerate kinase,Glutamate dehydrogenase 1).We selected some tumor-matastasis-related differentially expressed proteins,such as Fascin,Muskelin,Annexin A2,Cofilin-1 and Nav1.7,to investigate their functional roles.The expression of these proteins were detected by Western blot,and the results were consistent with the observation from 2-DE analysis.The differentially expressed proteins we selected are directly or indirectly involved in the regulation of cytoskeleton and its associated proteins,and then change the motility of cells,which leads to the increase or decrease on cell metastatic potential.Rho GTPase signaling pathway plays a central regulatory role during this process,and inhibite the activity of Rac1 or RhoA will mitigate the metastatic activity of Mat-LyLu cells,accompanied by differential expression of corresponding proteins.