Structural and functional consequences of disease SNP's on the transcriptome

摘要 遗传关联研究确定了与特定表型相关的基因类型。很大一部分 与人类疾病表型相关的单核苷酸多态(SNPs) 基因的蛋白质编码区。在这些情况下，疾病病因学的确切分子机制 并不是立竿见影的。该提案特别关注映射到非编码和 基因的非翻译区(UTRs)。如果这些SNP改变了RNA的结构，它们被归类为 RiboSNitch。我们将通过实验验证8个新的、通过计算预测的与 人类疾病肌萎缩侧索硬化症、乳腺癌和结直肠癌、角化不良、先天性巨结肠 疾病、脂肪酶缺乏症、小头症侏儒症和精神分裂症。我们的工作将极大地利用 与NEXT相结合的化学结构探测技术在吞吐量和准确性方面的进展 世代排序。此外，这些技术现在使我们能够探测体内的RNA结构，从而 我们进一步了解细胞环境如何影响RNA折叠和核糖核酸链的功能。我们 还将进行荧光素酶定量分析和利用Tet-off诱导系统来研究 验证的核糖核酸链对翻译和RNA稳定性的功能影响以确定疾病 因果关系。总之，我们的发现将在多个新人类中建立SNP诱导的RNA结构变化 并扩大了对RNA结构在塑造人类表型方面的理解。

SUMMARY Genetic association studies identify the genotypes that correlate with specific phenotypes. A significant portion of the Single Nucleotide Polymorphisms (SNPs) that associate with human disease phenotypes map outside of the protein coding regions of genes. In these cases, the precise molecular mechanism of the disease etiology is not immediately apparent. This proposal focuses specifically on SNPs that map to non-coding and UnTranslated Regions (UTRs) of genes. If these SNPs alter the structure of the RNA, they are classified as a riboSNitch. We will experimentally validate eight novel, computationally predicted riboSNitches associated with the human diseases amyotrophic lateral sclerosis, breast and colorectal cancer, dyskeratosis, Hirschsprung's disease, lipase deficiency, microcephalic dwarfism, and schizophrenia. Our work will leverage significant advances in the throughput and accuracy of chemical structure probing techniques in combination with next generation sequencing. Furthermore, these techniques now enable us to probe RNA structure in vivo allowing us to further understand how the cellular environment affects RNA folding and the function of riboSNitches. We will also perform quantitative luciferase reporter assays and leverage Tet-off inducible systems to study the functional consequences of validated riboSNitches on translation and RNA stability to establish disease causality. Together, our findings will establish SNP-induced RNA structure change in multiple new human diseases and broaden understanding of RNA structure in shaping human phenotype.