c-Myc-regulated microRNAs in normal and pathologic cellular physiology

描述(由申请人提供)：该申请是一项高效的R01研究计划的竞争性更新，该计划的重点是阐明microRNAs(MiRNAs)在驱动肿瘤发生的关键信号通路中的作用。MiRNAs是18-24个核苷酸的RNA分子，调节部分互补的靶信使RNA的稳定性和翻译效率。在过去的十年里，包括我们在内的许多实验室已经证实，miRNAs在癌细胞中经常处于失调状态，并可以影响恶性肿瘤的方方面面，包括增加增殖、抗凋亡和转移。此外，我们由这笔资金资助的工作已经证明，miRNAs在经典癌基因和肿瘤抑制基因的下游提供关键功能，包括MYC、KRAS和P53。这些发现使我们证明了基于miRNA的治疗策略可以有效地抑制动物模型中的肿瘤发生。然而，许多重要的问题仍然没有得到回答。我们仍然不完全了解导致肿瘤中miRNA异常表达的机制。此外，我们对miRNAs功能的获得和丧失在体内驱动肿瘤发生的分子机制知之甚少。现有的绝大多数功能数据来自于改变细胞系中miRNA的表达，这种方法没有完全模拟miRNAs如何参与肿瘤的启动和进展。在这笔赠款的下一个资助期，我们建议通过检验以下中心假设来解决这些关键的知识缺口：第一，miRNA初级转录本的选择性剪接和序列特异性RNA结合蛋白的关联影响致癌和肿瘤抑制因子miRNAs的表达和活性；第二，抗肿瘤生成miRNAs的活性可以抑制体内早期和晚期的肝脏肿瘤发生和淋巴癌发生。为了检验这些假说，我们将追求三个具体目标。在目标1中，我们将使用细胞和动物模型来直接确定选择性剪接在发育和疾病环境中调节miRNA生产和活性的作用，包括肿瘤模型。在目标2中，将研究特定RNA结合蛋白在调节一组抗肿瘤形成miRNAs的生物发生中的作用。最后，在目标3中，我们将利用一个新开发的具有可调控miRNA表达的小鼠模型来研究miRNA介导的肿瘤抑制机制在小鼠肝癌和淋巴瘤模型中的作用。这些目标将利用我们和我们的合作者的丰富经验，在体外和体内评估miRNA的调节和功能。我们预计，这些研究揭示的原理将广泛适用于我们对miRNAs在癌症中的作用的理解，并可能发现新的机会来操纵miRNA的表达和功能用于治疗目的。

DESCRIPTION (provided by applicant): This application is for competitive renewal of a highly productive R01 research program focused on elucidating the roles of microRNAs (miRNAs) in key signaling pathways that drive tumorigenesis. miRNAs are 18-24 nucleotide RNA molecules that regulate the stability and translational efficiency of partially complementary target messenger RNAs. Over the last decade, many laboratories including ours have established that miRNAs are frequently dysregulated in cancer cells and can influence all aspects of malignancy including increased proliferation, resistance to apoptosis, and metastasis. Moreover, our work funded by this grant has documented that miRNAs provide crucial functions downstream of classic oncogenes and tumor suppressors including MYC, KRAS, and p53. These findings led to our demonstration that miRNA-based therapeutic strategies can potently inhibit tumorigenesis in animal models. Nevertheless, many important questions remain unanswered. We still do not fully understand the mechanisms that result in abnormal miRNA expression in tumors. Furthermore, we know very little about the molecular mechanisms through which gain- and loss-of-function of miRNAs drives tumorigenesis in vivo. The vast majority of existing functional data has been derived from altering miRNA expression in cell lines, an approach that does not fully model how miRNAs participate in tumor initiation and progression. During the next funding period of this grant, we propose to address these critical knowledge gaps by testing the following central hypotheses: First, that alternative splicing of miRNA primary transcripts and association of sequence-specific RNA binding proteins influences the expression and activity of oncogenic and tumor suppressor miRNAs; and second, that the activity of anti-tumorigenic miRNAs can suppress early- and late-stage liver tumorigenesis and lymphomagenesis in vivo. Three Specific Aims will be pursued in order to test these hypotheses. In Aim 1, we will use cellular and animal models to directly determine the role of alternative splicing in regulating miRNA production and activity in developmental and disease contexts, including tumor models. In Aim 2, the role of specific RNA binding proteins in regulating the biogenesis of a set of anti-tumorigenic miRNAs will be investigated. Finally, in Aim 3, we will utilize a newly-developed mouse model with regulatable miRNA expression to investigate the mechanisms underlying miRNA-mediated tumor suppression in mouse models of liver cancer and lymphoma. These aims will take advantage of our extensive experience, and that of our collaborators, in evaluating miRNA regulation and function in vitro and in vivo. We anticipate that the principles revealed by these studies will be broadly applicable to our understanding of the roles of miRNAs in cancer and may uncover new opportunities to manipulate miRNA expression and function for therapeutic purposes.