| PC-1 was identified to be up-regulated in androgen-refractory prostate cancer cell line C4-2, compared with androgen sensitive parent cell line LNCaP, which was first cloned by professor Zhou Jian-guang. PC-1 belongs to Tumor Protein D52 family, many members of which are associated with a series of cancer and proliferation. The PC-1 protein contains 224 amino acids, the N-terminal 46 amino acids of which is unique, while the C-terminal 180 amino acids is highly homologous with D52 protein. PC-1 gene expression can be induced by androgen and predominantly expresses in prostate tissue.Previous studies reveal that PC-1 gene is located in 8q21.1 region of chromosome, which is frequently amplified in prostate cancer tissues. Immunohistochemical experiments reveal that PC-1 gene expression is predominantly up-regulated in prostate cancer tissue samples and prostate intraepithelial neoplasma (PIN), whereas remaining a low level in normal prostate tissue samples control. Moreover, immortalized NIH3T3 cells stably transfected with PC-1 expressing plasmid is transformed and display characteristics of malignant cancer cells. These results indicate that PC-1 possess some characteristics of oncogene and may play a particular role in prostate cancer progression.In this article, further investigation is performed to elucidate the biological functions of PC-1 gene in prostate cancer progression and the molecular mechanism mediating the functions of this gene. Firstly, gain-of-function analysis was implemented in C4-2B prostste cancer cell lines, cell clones constitutively expressing PC-1, and their mock-transfected counterparts were generated. MTT and colony anchor-independent growth in soft agar were performed to evaluate the effects of PC-1 gene expression on prostate cancer progression. RT-PCR and realtime PCR were used to analysis the expression of differential genes between C4-2B-PC-1 and C4-2B-neo cell lines. A series of experimental data demonstrated that that stable expression PC-1 was in positive correlation with prostate cancer malignancy, which enhanced the growth ability, changed growth and differentiate related gene expression of C4-2B cells. In order to explore the molecular mechanism of PC-1, the PC-1 regulated genes in LNCaP cell were further analyzed. There are Receptor Tyrosine protein Kinase signal pathway members EphA3 and SGEF/CSGEF among those responding genes, whose function were explored in order to identify the mechanism PC-1 gene worked.Our results showed EphA3 interacted with SGEF/CSGEF, which indicated that PC-1 may play effect on prostate cancer development by regulating RTK signal pathway.Both the results of GST pull down and co-immunoprecipitation showed that SGEF interacted with androgen receptor (AR). Moreover, SGEF interacted with AR mostly depending on its PH domain, and AR interacted with SGEF mostly depending on its Ligand Binding Domain according to the experiment of GST pull down and mammalian two-hybrid respectively. In fact, PC-1 interacted with AR by other independent research. Androgen receptor(AR) triggered by androgen actions plays an important role in prostate development, maintenance of prostate organs and prostate cancer. Our research indicated that there was some relationship between RTK signal pathway and AR signal pathway, and that the regulating system PC-1 participated was a complicate network.In order to make further studies on SGEF/CSGEF, rabbit polyclonal antibody was prepared. Good specificity and potency polyclonal antiserum was gained. PSA-Luc and AR-Luc reporting genes were used to analyze AR transcription activity in LNCaP cells, but the effects produced by SGEF/CSGEF is little.In conclusion, our data provide cellular and molecular evidence to PC-1 function in prostate cancer progression and its regulating signal pathway initiatorily, which gave molecular biologic grouding for PC-1 to be a molecular marker for prostate cancer diagnosis.
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