Alternative RNA splicing and protein products in leukemia outcome (PQ11)

描述(由申请人提供)：本申请将回答NCI的挑衅性问题11：RNA处理的变化如何有助于肿瘤的发展？肿瘤和白血病极大地增加了异常RNA剪接水平，这会产生大量转录本，可以编码不同的蛋白质。RNA产物复杂性的增加可能是由于转化细胞中的“嘈杂”剪接增加了，但没有功能后果。或者，变异的RNA及其编码的蛋白质可以积极地促进转化的表型。肿瘤中选择性剪接水平的增加导致了转录本的表达，这些转录本太多，无法单独测试，例如通过过度表达或击倒实验。此外，如果变异蛋白质的重要性是由于群体作用或组合效应，那么单独评估它们的重要性是不够的。我们的假设是，如果肿瘤或白血病表现出的选择性剪接增加有助于肿瘤的发展，那么确定由选择性剪接RNA编码的蛋白质变体的预测表达水平将更有信息量，并且与结果的相关性将比简单地测量RNA水平更好。如果选择性剪接的产物没有显著的生物学作用，那么确定选择性剪接的RNA产生了哪些蛋白质将不会在预测结果方面提供额外的优势。我们将应用一种创新的下一代RNA测序方法来分析一大组高危儿童B-祖细胞急性淋巴细胞白血病(ALL)样本中的选择性RNA剪接。尽管最近B-ALL的治疗方法有所改进，但这一高风险队列代表了一群几乎没有好的治疗选择的患者。我们的方法将产生超过99%的表达转录本的整个长度的结构信息，使我们能够分析基因表达、RNA剪接和预测在每个样本中产生的蛋白质变体的群体。比较这些数据集将使我们能够回答这个具有挑衅性的问题，并确定增加替代RNA剪接水平是否对B-祖细胞ALL的发展重要。选择性剪接机制包含许多特性不佳的酶和调节蛋白。如果发现更多的选择性剪接在肿瘤发展中发挥作用，这些蛋白质将成为开发新药或干预措施的新的潜在靶点。

DESCRIPTION (provided by applicant): This application will address NCI Provocative Question 11: How do changes in RNA processing contribute to tumor development? Tumors and leukemias have dramatically increased levels of aberrant RNA splicing, which generates a large population of transcripts that could encode variant proteins. This increased complexity of RNA products could be due to "noisy" splicing that is increased in transformed cells, but has no functional consequence. Alternatively, the variant RNAs and the proteins they encode could actively contribute to the transformed phenotype. Increased levels of alternative splicing in tumors leads to the expression of transcripts that are too numerous to test individually, for example by over-expression or knock down experiments. Furthermore, if the importance of the variant proteins is due to a mass action or combinatorial effect, then assessing their importance individually will not suffice. Our hypothesis is that if the increased alternative splicing exhibitd by tumors or leukemias contributes to tumor development, then determining the predicted expression levels of protein variants encoded by the alternatively spliced RNAs will be more informative, and will correlate with outcome better than simply measuring the RNA levels. If the products of alternative splicing play no significant biological role, then determining which proteins are produced by the alternatively spliced RNAs will offer no additional advantage in predicting outcome. We will apply an innovative next-generation RNA sequencing approach to the analysis of alternative RNA splicing in a large group of high risk childhood B- progenitor Acute Lymphocytic Leukemia (ALL) samples. Despite recent improvements in the treatments for B- ALL, this high risk cohort represents a group of patients for whom few good treatment options exist. Our approach will produce structural information over the entire length of more than 99% of expressed transcripts, allowing us to analyze gene expression, RNA splicing and the populations of protein variants that are predicted to be produced in each sample. Comparing these data sets will allow us to answer the Provocative Question and to determine whether increased levels of alternative RNA splicing are important in the development of B- progenitor ALL. The alternative splicing machinery contains many poorly characterized enzymes and regulatory proteins. If increased alternative splicing is found to play a role in tumor development these proteins will represent novel potential targets for the development of new drugs or interventions.