Angiogenin-induced RNA cleavage in cancer

描述(申请人提供)：本提案的目的是确定血管生成素(Ang)诱导的转移RNA(TRNA)裂解如何促进肿瘤的生长和存活。我们的中心假设是，Ang诱导的tRNA裂解改变蛋白质翻译，增强血管生成、促生长和促生存蛋白的表达，从而促进肿瘤的发生。这是基于我们自己的初步数据，表明Ang选择性地切割tRNAs，产生抑制翻译起始的生物活性片段(即tiRNAs)。TE提出的研究的基本原理是，一旦我们知道tRNA裂解如何重新编程蛋白质翻译，我们将能够调节这一事件来治疗癌症。我们将通过完成三个特定的目标来验证我们的中心假设：目的1.确定YB-1如何与tiRNAs合作抑制癌细胞中的翻译启动。我们的工作假设是，tiRNAs与YB-1的核酸结合冷休克结构域结合，以促进与翻译起始复合体成分的相互作用。目的2.确定血管紧张素转换酶(Ang)诱导的tRNA裂解如何改变蛋白质翻译以促进癌细胞的增殖和存活。我们的工作假设是，tiRNAs“激活”YB-1，允许优先翻译内部核糖体进入位点(IRSE)和含有上游开放阅读框架(UORF)的转录本，编码促进肿瘤细胞生长和存活的蛋白质。目的3.确定血管紧张素转换酶(Ang)诱导的tRNA裂解在肿瘤发生中的作用。我们的工作假设是，tiRNAs作用于Ang下游，促进肿瘤生长。我们将确定YB-1癌基因是否与tRNA片段合作，重新编程癌细胞中的蛋白质翻译。YB-1是一种针对促进上皮向间充质转化的翻译机制的癌基因。我们将使用无偏见的基因阵列和候选基因方法来识别其翻译受tRNA切割影响的转录本。我们将使用siRNA敲除来确定Ang抑制剂RNH1是否具有肿瘤抑制蛋白的功能。我们将确定tRNA片段、YB-1和RNH1的表达如何影响小鼠异种移植瘤的发生。最后，我们将确定tRNA裂解是否可以作为前列腺癌的预后生物标记物。这项拟议的研究的贡献将是确定Ang的核糖核酸酶活性如何促进肿瘤细胞的生长和存活。这一贡献意义重大，因为它为开发预防血管紧张素转换酶介导的肿瘤生长的药物策略提供了分子基础。这项拟议的研究具有创新性，因为它专注于Ang的直接靶点，即其RNA底物，并试图确定tRNA裂解如何促进肿瘤发生。

DESCRIPTION (provided by applicant): The objective of this proposal is to determine how angiogenin (ANG)-induced transfer RNA (tRNA) cleavage facilitates tumor growth and survival. Our central hypothesis is that ANG-induced tRNA cleavage alters protein translation to enhance the expression of angiogenic, pro-growth and pro-survival proteins to promote tumorigenesis. This is based upon our own preliminary data showing that ANG selectively cleaves tRNAs to produce bioactive fragments (i.e., tiRNAs) that inhibit translation initiation. The rationale for te proposed research is that, once we know how tRNA cleavage re-programs protein translation, we will be able to modulate this event to treat cancer. We will test our central hypothesis by the completion of three specific aims: AIM 1. Determine how YB-1 cooperates with tiRNAs to inhibit translation initiation in cancer cells. Our working hypothesis is that tiRNAs bind to the nucleic acid-binding cold shock domain of YB-1 to promote interactions with components of the translation initiation complex. AIM 2. Determine how ANG-induced tRNA cleavage alters protein translation to augment the proliferation and survival of cancer cells. Our working hypothesis is that tiRNAs "activate" YB-1 to allow the preferential translation of internal ribosome entry site (IRSE) and upstream open reading frame (uORF)-containing transcripts encoding proteins that promote tumor cell growth and survival. AIM 3. Determine the role of ANG-induced tRNA cleavage in tumorigenesis. Our working hypothesis is that tiRNAs act downstream of ANG to promote tumor growth. We will determine whether YB-1, an oncogene that targets the translational machinery to promote the epithelial- mesenchymal transition, partners with tRNA fragments to re-program protein translation in cancer cells. We will use non-biased gene array and candidate gene approaches to identify transcripts whose translation is modulated in response to tRNA cleavage. We will use siRNA knockdown to determine whether the ANG inhibitor RNH1 functions as a tumor suppressor protein. We will determine how the expression of tRNA fragments, YB-1 and RNH1 influence tumorigenesis in murine xenografts. Finally, we will determine whether tRNA cleavage serves as a prognostic biomarker for prostate cancer. The contribution of the proposed research will be to determine how the ribonuclease activity of ANG promotes the growth and survival of tumor cells. This contribution is significant because it provides a molecular basis for the development of pharmacologic strategies to prevent ANG-mediated tumor growth. The proposed research is innovative because it focuses on the direct target of ANG, its RNA substrates, and attempts to determine how tRNA cleavage promotes tumorigenesis.