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目的探讨PES1与雌激素受体(ER)的相互作用及其对ER转录活性的影响。方法将体外翻译的PES1与纯化的GST-ERα和GST-ERβ蛋白分别混合,用GST pull-down验证在体外PES1与ERα和ERβ是否存在相互作用。将HA-PES1与FLAG-ERα或FLAGC-ERβ共转染293T细胞后进行免疫共沉淀,以验证PES1与ER是否在体内有相互作用。用含雌激素受体作用元件的荧光素酶报告基因检测PES1对ERα和ERβ转录活性的影响。结果 PES1与ERα、ERβ在体内外均存在相互作用,而且PES1与ERα的结合比与ERβ的强。在体内,在雌激素(E2)存在下,E2可以增强PES1与ERα的结合,而对PES1与ERβ的结合没有明显影响。PES1对ER转录活性的影响是E2依赖性的,PES1能升高ERα的转录活性而降低ERβ的转录活性(P<0.01)。结论 PES1是一种新的ER共调节因子,能反向调节ERα和ERβ的转录活性,需要进一步研究的是其在ER信号通路以及在E2诱发的肿瘤发生发展中的作用。
Objective To investigate the interaction between PES1 and estrogen receptor (ER) and its effect on ER transcriptional activity. METHODS: In vitro translated PES1 was mixed with purified GST-ERα and GST-ERβ proteins respectively and GST pull-down was used to verify the interaction between PES1 and ERα and ERβ in vitro. 293T cells were cotransfected with HA-PES1 and FLAG-ERα or FLAGC-ERβ and then co-immunoprecipitated to verify whether PES1 interacts with ER in vivo. The effect of PES1 on the transcriptional activity of ERα and ERβ was examined using a luciferase reporter gene containing estrogen receptor-responsive elements. Results PES1 interacted with ERα and ERβ in vitro and in vivo, and the binding of PES1 to ERα was stronger than that of ERβ. In vivo, E2 enhances the binding of PES1 to ERα in the presence of estrogen (E2), with no apparent effect on the binding of PES1 to ERβ. The effect of PES1 on ER transcriptional activity was E2-dependent. PES1 increased the transcriptional activity of ERα and decreased the transcriptional activity of ERβ (P <0.01). Conclusions PES1 is a new ER co-regulator that reversely regulates the transcriptional activity of ERα and ERβ. Further studies are needed to elucidate its role in ER signaling pathway and E2-induced tumorigenesis.