ErbB Receptor Control of Breast Cancer Growth

描述（由申请人提供）：我们最近克隆了一种新的蛋白， Ebp 1，与ErbB-3受体的胞质结构域相互作用。 用ErbB-3配体heregulin（HRG）处理细胞可诱导细胞凋亡。 Ebp 1从细胞质易位到细胞核。受管制的核 Ebp 1的积累表明它可能作为转录因子发挥作用， 一种在激活前被隔离在细胞质中的辅助调节因子，类似于 STAT或Smads。Ebp 1的过表达抑制增殖并诱导 人乳腺癌细胞的分化。EBP 1与肿瘤相互作用 抑制蛋白Rb和辅抑制因子Sin 3A，并抑制转录 与细胞周期相关的基因。本提案的总体目标是 进一步了解Ebp 1的转录抑制机制， 确定抑制转录的能力如何有助于Ebp 1的 生物效应。在具体目标1中，我们将证实Ebp 1诱导的 转录抑制介导其生物学效应。具体 实验包括1）确定Ebp 1调节活性的能力 2）Ebp 1转录因子的鉴定和定位 抑制结构域3）描绘Ebp 1与 组蛋白脱乙酰酶（HDAC）复合物4）评估Ebp 1结合的能力 DNA作为蛋白质复合物的一部分5）检查Ebpl诱导的 转录抑制其生物学效应6）鉴定Ebp 1 有约束力的伙伴在具体目标2中，我们将继续研究 Ebp 1通过磷酸化发挥作用。我们将a）确定规则， 不同生长条件下磷酸化Ebp 1亚细胞定位 条件B）使用SELDI技术绘制Ebp 1磷酸化位点图和用途 c）测试Ebp 1的能力 磷酸化突变体，以调节转录、细胞生长和 分化第三，我们将定义正常的生理功能， ebp 1在哺乳动物系统中通过基因敲除小鼠的建立和研究。这些 实验应该有助于理解Ebp 1的能力， 抑制转录有助于其对细胞生长的影响， 分化开发合理和特异性的治疗方法， 阻遏物复合物为癌症治疗提供了新的方向。一个 了解Ebp 1抑制细胞生长和诱导 分化可能有助于设计新的分子靶点， 癌症治疗

DESCRIPTION (provided by applicant): We have recently cloned a novel protein, Ebp1, which interacts with the cytoplasmic domain of the ErbB-3 receptor. Treatment of cells with the ErbB-3 ligand, heregulin (HRG) induces the translocation of Ebp1 from the cytoplasm to the nucleus. The regulated nuclear accumulation of Ebp1 suggests that it may function as a transcriptional coregulator that is sequestered in the cytoplasm before activation similar to STAT or Smads. Overexpression of Ebp1 inhibits proliferation and induces differentiation of human breast cancer cells. Ebp1 interacts with the tumor suppressor protein, Rb and the corepressor Sin3A, and represses transcription of cell cycle-related genes. The overall aim of the current proposal is to further understand the mechanisms of Ebp l's transcriptional repression and to determine how the ability to repress transcription contributes to Ebp1's biological effects. In Specific Aim 1 we will confirm the role of Ebp1-induced transcriptional repression in mediating its biological effects. Specific experiments include 1) determining the ability of Ebp1 to modulate the activity of endogenous promoters 2) identifying and mapping Ebp1 transcriptional repression domains 3) delineating the interactions of Ebp1 with components of histone deacetylase (HDAC) complexes 4) assessing the ability of Ebp1 to bind DNA as part of a protein complex 5) examining the contribution of Ebpl induced transcriptional repression to its biological effects 6) indentifying Ebp1 binding partners. In specific Aim 2, we will continue to study regulation of Ebp1 function by phosphorylation. We will a) determine the regulation and subcellular localization of phosphorylated Ebp1 under different growth conditions b) map Ebp1 phosphorylation sites using SELDI technology and use this knowledge to design phosphospecific antibodies c) test the ability of Ebp1 phosphorylation mutants to regulate transcription, cell growth, and differentiation. Third, we will define the normal physiological functions of Ebp1 in a mammalian system by the creation and study of knockout mice. These experiments should contribute to an understanding of how the ability of Ebp1 to repress transcription contributes to its effects on cell growth and differentiation. The development of rational and specific therapies that target repressor complexes is providing a new direction in cancer treatment. An understanding of the mechanisms by which Ebp1 inhibits cell growth and induces differentiation may contribute to the design of new molecular targets for cancer therapy.