A Role for the Novel Poly(A) RNA-Binding Protein, ZC3H14, in Breast Cancer

描述(申请人提供)：仅今年一年，乳腺癌就将夺走大约39,000名美国女性的生命。因此，揭示乳腺癌发病机制的新分子机制对于填补我们对这种疾病的了解空白并挽救每年数千名妇女的生命至关重要。基因表达改变是癌症调控异常的一个关键点，越来越多的证据表明，转录后的mRNA处理在确保基因正确表达方面发挥着重要作用。转录后事件是由大量的RNA结合蛋白(RBPs)介导的，其中一类关键的蛋白是Poly(A)结合蛋白(PABS)。一种新的CCCH锌指多克隆抗体ZC3H14最近被认为与乳腺肿瘤的雌激素受体(ER)状态有关。虽然ZC3H14的分子功能尚不清楚，但芽生酵母对应物NaB2是适当控制Poly(A)尾长和从细胞核输出mRNA所必需的，这与关键的 ZC3H14在转录后调控中的作用。虽然ZC3H14是一种多克隆抗体，但我们的初步数据表明，在乳腺癌细胞中，ZC3H14调控着特定的mRNA转录。ZC3H14可能通过另一个识别特定mRNAs的RBP实现这种转录特异性，例如人类抗原R(Hur)，一种与乳腺癌密切相关的富含AU元素的结合蛋白。之前在酵母中的一项研究表明，NaB2和Pub1(酿酒酵母Hur Ortholog)之间的相互作用影响了靶mRNA转录本的稳定性。在对ZC3H14和HUR的全基因组分析中，我们发现有证据表明，这两个蛋白质的候选mRNA靶标的3‘UTR区存在共同的顺式元件，以及候选靶标之间存在显著重叠。这些数据表明，ZC3H14和HUR一样，都有特定的mRNA靶点，ZC3H14和HUR可能具有功能关系。在这个建议中，我们将使用多种方法来验证我们的假设，即ZC3H14与目标mRNA转录本3‘UTR中的特定顺式元件结合，以调节其稳定性和/或聚(A)尾长，可能与Hur一致。从长远来看，我们试图进一步深入了解限制性商业惯例在调节基因表达方面所起的作用。在我们的第一个目标中，我们将验证特定的ZC3H14靶标，并在这些靶标mRNAs中确定候选ZC3H14调控序列，以评估ZC3H14与靶标mRNAs结合的转录后后果。在我们的第二个目标中，我们将研究ZC3H14和HUR在调节靶mRNAs方面的功能关系。这项研究将深入了解聚(A)限制性商业惯例(如ZC3H14)作为转录后调控因子的作用及其在乳腺癌中的潜在参与。

DESCRIPTION (provided by applicant): Breast cancer will claim the lives of approximately 39,000 American women this year alone. Uncovering novel molecular mechanisms of breast cancer pathogenesis is therefore critical to fill gaps in our understanding of this disease and to save the lives of thousands of women every year. Altered gene expression is one critical point of dysregulation in cancer and there is increasing evidence that post-transcriptional processing of mRNA transcripts plays a major role in ensuring proper gene expression. Post-transcriptional events are mediated by a myriad of RNA-binding proteins (RBPs), one key class of which is comprised of poly (A) binding proteins (Pabs). A novel CCCH zinc finger Pab, ZC3H14, has recently been linked to the Estrogen Receptor (ER) status of breast tumors. Although the molecular function of ZC3H14 is unknown, the budding yeast counterpart, Nab2, is required for proper control of poly (A) tail length and mRNA export from the nucleus, consistent with a critical role for ZC3H14 in post-transcriptional regulation. Although ZC3H14 is a Pab, our preliminary data suggest that specific mRNA transcripts are modulated by ZC3H14 in breast cancer cells. ZC3H14 may achieve this transcript specificity via another RBP that recognizes specific mRNAs, such as Human Antigen R (HuR), an AU-rich element binding protein already strongly linked to breast cancer. A previous study in yeast demonstrated an interaction between Nab2 and Pub1 (S. cerevisiae HuR ortholog) that influenced the stability of target mRNA transcripts. In a genome-wide analysis of ZC3H14 and HuR, we identified evidence of consensus cis-elements located within the 3' UTR of candidate mRNA targets of the two proteins as well as a significant overlap between candidate targets. These data suggest that ZC3H14, like HuR, has specific mRNA targets, and that ZC3H14 and HuR may have a functional relationship. In this proposal we will employ multiple approaches to test our hypothesis that ZC3H14 binds to specific cis- elements within the 3' UTR of target mRNA transcripts to modulate their stability and/or poly (A) tail length, potentially in concert with HuR. In the long-term, we seek to provide further insight into the role that RBPs play in modulating gene expression. In our first aim, we will validate specific ZC3H14 targets and identify candidate ZC3H14 regulatory sequences in these target mRNAs to assess the post-transcriptional consequence of ZC3H14 binding to target mRNAs. In our second aim, we will examine the functional relationship between ZC3H14 and HuR in modulating target mRNAs. This study will provide insight into the role of poly (A) RBPs such as ZC3H14 as post-transcriptional regulatory players and their potential involvement in breast cancer.