摘要
雌激素受体(ER)是核受体超级家族成员之一,它包括两种亚型:α和β。ER在乳腺癌的发生发展中起重要作用,目前认为ER是乳腺癌内分泌治疗的靶标和预后的指标之一。目前为止,乳腺癌内分泌的治疗主要是通过降低ER转录活性而实现。因此调节ER转录活性的蛋白因子的发现和相应的功能研究对于探讨乳腺癌的发病机制、预防乳腺癌的发生、有效开发治疗与雌激素相关疾病的药物具有重大意义。
本实验以ERαAF1为诱饵蛋白,通过酵母双杂交方法筛选得到了与ERαAF1结构域结合的蛋白collagen Ⅰ和FBLN1。collagen Ⅰ和FBLN1都是细胞外基质的组成成分,酵母双杂交实验证实了collagen Ⅰ、FBLN1与ERα的直接结合作用,可以推测,collagen Ⅰ、FBLN1可能参与ER信号途径并共同调节细胞的增殖,ECM除了是肿瘤细胞转移的机械屏障外,通过蛋白与蛋白间的相互作用在分子水平上对肿瘤的形成予以调节。
本实验以ERβ AF2为诱饵蛋白,筛选到了ERβ AF2的结构域结合的HPIP和一个新的未知蛋白。本文首次发现HPIP能改变ER的转录活性,HPIP是2000年Abramovich C等以PBX1作为诱饵,利用酵母双杂交技术从胎儿肝脏cDNA文库中筛选得到的。PBX是一组由Pbx基因编码的含有TALE高度保守结构的同源蛋白,
Estrogen receptor (ER) is one of the members of the nuclear receptor superfamily, and contains two subtypes: a and p. ER plays an important role in the development of breast cancer. Recently ER is thought to act as a target of the therapy of breast cancer and good index of prognosis. Until now, the endocrine therapy of mammary carcinoma is acquired mainly by decreasing ER transcriptional activity. Therefore, it is of great significance to discover and characterize the proteins that modulate ER activity in developing the drugs of diseases related to estrogen.With ERα AF1 as bait, collagen I and FBLN1 was identified by yeast two-hybrid. Collagen I and FBLN1 are component of extracellular matrix (ECM), yeast two-hybrid assasy confirmed that collagen I and FBLN1 combined with ER directly. The collagen I and FBLN1 may participate in the ER signaling pathway to combine to regulate the cell procreation. Besides as mechanical barrier, ECM regulate tumor formation on molecular level through interaction between proteins.With ERβ AF2 as bait, HPIP and a new agnoprotein was identified by yeast two-hybrid. A yeast two-hybrid screen of a fetal liver-hematopoietic cDNA library using PBX1 as bait led to the discovery of this novel protein that interacts with PBX1 by Abramovich C in 2000 . PBX is a homeodomain protein that functions in complexes with other homeodomain-containing proteins to regulate gene expression during developmental and differentiation processes,including PBX1 、 PBX2 、 PBX3 and PBX4 . This protein that we have termed hematopoietic PBX-interacting protein (HPIP) is mainly localized in the cytosol and in small amounts in the nucleus. A yeast two-hybrid simulate internal environment completeiy .Tentatively confirmed that the bait protein combined with HPIP directly.As ER can bind to ERE (estrogen response element), ERE-containing luciferase reporter assay is used to detect the effects of HPIP
引文
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