| One of the most effective comprehensive treatments for estrogen receptorpositive(ER+) breast cancer patients. However, about40%of the ER+breastcancer patients are primary or secondary resistant to endocrine therapy whichhas become a severe obstacle for the treatment of breast cancer.Estrogen-responsive finger protein(Efp) is one of the important downstreamtarget genes of ER. Being a E3ubiquitin ligase, Efp mediates the degradationof14-3-3σ through the ubiquitin proteasome pathway, which underlying thedrug resistance to endocrine treatment for ER-positive breast cancer patients.Newly study has been shown that the expression of Plk3is negativelyregulated to ER in breast cancer and it inhibit the proliferation of ER+breastcancer cells. Meanwhile, Plk3is easily degraded in cells and can be stabilizedby protease inhibitor MG132. Thus, we conjecture that the low expression ofPlk3in breast cancer cells may be caused by Efp through the ubiquitylationand degradation pathway. In this study, we investigate the expression and therelationship between Efp and Plk3in breast cancer cells and tissues, study therole and mechanism of Efp in breast cancer development, and provide somenew evidence and experiment data for breast cancer therapy.Methods:1Quantitative real-time PCR assay was carried out to test the mRNAexpression of Efp and Plk3in54breast cancer tissues, checkout therelationship between Efp, Plk3and relevant pathology index in clinical.2Gene transfection, RT-PCR assay were performed to evaluate theexpression of Efp and Plk3in ER+breast cancer cell MCF-7.3Gene transfection assay was performed to establishe human BreastCancer MCF-7transplanted nude mice model. Measure the tumor size ingrowth to make growth curve. Get the weight of transplanted tumor tissues after the mice were killed. Evaluate Efp and Plk3effects on tumordevelopment in vivo.4Immunohistochemical staining was used to detect the proteinexpression of Efp and Plk3in transplanted tumor tissue. Analyse therelationship between them.Results:1Relative quantitative analysis of Efp and Plk3mRNA expression of54breast cancer tissues indicates that the expression of Efp positively correlatedwith Plk3gene (r=0.554, P<0.01). No significant relationship was foundbetween Efp and Plk3mRNA with patient age, tumor size, menopausal status,Lymph node metastasis, pathological staging and pathological types. Theexpression of Efp gene is regulated with the expression level of K-i67protein(P=0.043). The expression of Plk3gene positively correlated with theexpression level of ER protein (r=0.307, P=0.025).2The results of RT-PCR shows that the transfection of Efp markedlyreduced the expression of Plk3mRNA, compared with the empty vectorsgroup (P<0.05). The mRNA expression levels of Plk3were significantly lowerafter co-transfected with Efp and Plk3than those transfeced with Plk3only(P<0.05).3Establish xenograft tumor in nude mice successfully. Transfected withempty vector, transfected with Efp plasmid alone, co-transfected with Efp andPlk3, transfected with Plk3alone were performed to MCF-7cells separately.Transfected cells were than treated with nude mice. The four groups ofdifferently treated xenograft tumor had no significant difference in tumorgrowth trend when analysed the tumor volume(F=0.009，P>0.05). Tumorweight of four groups was1.88±0.15g,2.13±0.19g,1.96±0.23g and1.87±0.12g, respectively. Transfection of Efp increased tumor weight comparedwith the empty vectors treated group (P<0.05). Transfected with Efp higherthe tumor weight compared with the Plk3transfection group (P<0.05).4The results of immunohistochemical staining shows that the proteinlevels of Plk3in differently treated xenograft tumor were8.26±1.15,2.48± 0.04,4.57±0.06and9.75±0.02, measured by integrated optical density(IOD). Compared with Efp transfection treated group, the expression of Plk3protein was markedly lower than Efp free group(P<0.05). The proteinexpression levels of Plk3were significantly lower after co-transfected withEfp and Plk3than those transfeced with Plk3only (P<0.05).Conclusion:Efp inhibits the transcriptional activity of Plk3, while further explorationneeds to be carried out to find the interaction between Efp and Plk3at genelevel. |
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