Alternative polyadenylation and non-stop mRNAs in Arabidopsis

智力优势：在真核生物中，基因调控的许多方式之一是通过一个被称为替代性多聚腺苷酸化的过程;这个过程需要沿着初级RNA转录物对信使RNA将终止的位点进行差异选择（并且将添加典型的poly（A）尾）。 之前NSF支持的研究发现了一类新的替代性多聚腺苷酸化位点，这些位点在与植物胁迫反应相关的基因中普遍存在。 这些位点被预测反映新的基因表达和调控机制。 为了验证这一预测，将研究终止于这些位点的mRNA的形成和性质。 在这个项目中要测试的假设是：在这些新的网站结束的mRNA是由典型的多聚腺苷酸化机制形成的;在这些网站结束的mRNA改变相关的mRNA的功能;和多聚腺苷酸化在这些网站是需要拥有这样的网站的基因的功能。 这项研究将涉及生物化学测定，高通量测序的DNA标签，查询mRNA-poly（A）连接以及RNA聚合酶II的位置沿着基因组，瞬时基因表达的研究，在烟草本塞姆亚纳，和实验利用拟南芥突变体和转基因植物。 更广泛的影响：拟议的研究解决了一种新的基因调控模式，涉及植物对胁迫的反应。 从这些研究中获得的知识将适用于大多数作物植物，这是合理的，预计新的战略管理疾病和非生物胁迫作物植物将即将到来。 研究涉及广泛的方法;学员将因此获得生物化学，基因组学和计算生物学以及分子遗传学的第一手经验。 研究方法和结果将融入肯塔基州大学的研究生和本科生课程。 此外，这些方法和结果将提供给一些以本科生为重点的机构的教师，作为开发高通量DNA测序和基因组学课程模块的推广工作的一部分。

Intellectual Merit: Among the many ways that genes are regulated in eukaryotes is via a process termed as alternative polyadenylation; this process entails the differential choice of sites along a primary RNA transcript at which the messenger RNAs will end (and to which the canonical poly(A) tail will be added). Previous NSF-supported research resulted in the discovery of a novel class of alternative polyadenylation sites that are prevalent in genes associated with stress responses in plants. These sites are predicted to reflect novel gene expression and regulatory mechanisms. To test this prediction, the formation and properties of mRNAs that end at these sites will be studied. The hypotheses to be tested in this project are: mRNAs that end at these novel sites are formed by canonical mechanisms of polyadenylation; mRNAs that end at these sites alter the functionalities of the associated mRNAs; and polyadenylation at these sites is required for the functions of genes that possess such sites. This research will involve biochemical assays, high throughput sequencing of DNA tags that query mRNA-poly(A) junctions as well as the positions of RNA polymerase II along the genome, transient gene expression studies in Nicotiana benthamiana, and experiments utilizing Arabidopsis mutants and transgenic plants. Broader Impacts: The proposed research addresses a new mode of regulation of genes involved in responses of plants to stress. The knowledge gained from these studies will apply to most crop plants, and it is reasonable to expect that new strategies for managing disease and abiotic stress in crop plants will be forthcoming. The research involves a wide range of approaches; trainees will thus gain first-hand experience in biochemistry, genomics and computational biology, and molecular genetics. The research methods and results will be integrated into graduate and undergraduate classes at the University of Kentucky. Additionally, the methods and results will be made available to faculty at a number of undergraduate-focused institutions as part of outreach efforts to develop curricular modules for high throughput DNA sequencing and genomics.