Protein-RNA interactions in cancer

项目摘要/摘要 蛋白质-RNA相互作用是基因调控的一个重要而未被充分研究的组成部分。RNA结合 蛋白质具有许多与mRNA的生产、剪接、加工和稳定性有关的功能 分子。几年前，我们发现肿瘤胎儿RNA结合蛋白IGF2BP3与3‘端结合。 并通过一种涉及RNA诱导的机制来调节mRNA的稳定性 沉默情结。在最近的工作中，我们发现IGF2BP3也在3‘-剪接位点附近结合，并可能 规范替代剪接。综上所述，我们的发现表明IGF2BP3在mRNA调控中具有双重作用。 IGF2BP3调节与增殖、迁移和信号传递有关的基因--这些基因对胎儿很重要 发展--但也与癌症有关。与这一基因调控功能一致，IGF2BP3在 广泛的恶性肿瘤，包括急性白血病，当高表达时预示着预后不良。 使用IGF2BP3缺乏症的新的小鼠遗传模型，我们现在发现IGF2BP3是必需的 在体内发展出一种完全穿透性的致命白血病。加在一起，我们广泛的前期工作，两者 已发布和未发布，为其功能提供了一个机械框架，并为 这种蛋白质在疾病中的重要性。为了充分了解这些蛋白质-RNA相互作用的性质，并 为了了解它们在癌症中的作用，我们提出了两个目标。在第一个目标中，我们将仔细描述机制 IGF2BP3的潜在功能通过一系列仔细执行的实验相结合(目标1)。在第二个目标中， 我们将使用一个集合来描述IGF2BP3在白血病的启动、增殖和维持中的重要性 精心构建的遗传模型和原代细胞的基因编辑。接下来，我们将描述两者是如何 已提出的基因调控机制-RNA稳定性和前mRNA剪接-在癌症的启动中发挥作用， 利用反向遗传学、miRNA调控和异构体特异性表达的组合。共同努力，这项工作 将导致对蛋白质-RNA相互作用的机制基础如何密切地进行分层、详细的理解 与基因表达失控和细胞恶性转化有关。重要的是，它将为 发展以巨噬细胞为特征的恶性肿瘤的新诊断和治疗方法 RNA结合蛋白的改变导致转录调控失调。

PROJECT SUMMARY/ ABSTRACT Protein-RNA interactions underlie an important and understudied component of gene regulation. RNA-binding proteins carry out numerous functions relating to the production, splicing, processing, and stability of mRNA molecules. A few years ago, we discovered that the oncofetal RNA binding protein, IGF2BP3, binds to the 3’ untranslated region of mRNA and regulates mRNA stability via a mechanism that involves the RNA-induced silencing complex. In more recent work, we found that IGF2BP3 also binds near 3’-splice sites, and may regulate alternative splicing. Together, our findings suggest dual roles in mRNA regulation for IGF2BP3. IGF2BP3 regulates genes that are related to proliferation, migration, and signaling- which are important in fetal development- but also in cancer. Concordant with this gene regulatory function, IGF2BP3 is overexpressed in a wide range of malignancies, including acute leukemia, and portends a poor prognosis when highly expressed. Using novel, murine genetic models of IGF2BP3 deficiency, we have now discovered that IGF2BP3 is required for the development of a fully-penetrant, lethal leukemia in vivo. Together, our extensive prior work, both published and unpublished, provides a mechanistic framework for its function, and a solid foundation for the importance of this protein in disease. To fully understand the nature of these protein-RNA interactions and to understand their role in cancer, we propose two aims. In the first aim, we will carefully delineate the mechanism underlying IGF2BP3 function by a combination of carefully executed experiments (Aim 1). In the second aim, we will characterize the importance of IGF2BP3 in leukemia initiation, propagation and maintenance using a set of carefully constructed genetic models and gene editing in primary cells. Next, we will characterize how the two proposed gene regulatory mechanisms- RNA stability and pre-mRNA splicing- play roles in cancer initiation, using a combination of reverse genetics, miRNA regulation, and isoform specific expression. Together, this work will lead to a layered, detailed understanding of how mechanistic basis of protein-RNA interactions is intimately connected to gene expression deregulation and the malignant transformation of cells. Importantly, it will pave the way to develop novel diagnostic and therapeutic approaches in malignancies characterized by massive transcriptomic dysregulation underpinned by alterations of RNA-binding proteins.