Investigating the coordinated interplay of mRNA and tRNA modifications mediated by the FTO-TRMT10A complex

项目总结/摘要 RNA修饰几乎同时存在于所有的RNA种类中，并且可以显著改变RNA的性质。 和RNA的表达。mRNA的修饰可以通过几种机制影响基因表达， 本地化、稳定性和翻译效率。已知对tRNA的修饰会影响其稳定性， 丰富度以及翻译过程中的解码和延伸动态。许多失调 酶介导的RNA修饰与受损的神经发育有因果关系， 智力残疾和行为异常这个领域已经做了很大的努力来了解 单个修饰对分离RNA的功能影响，但mRNA和tRNA的相互作用 修改从未被探索过。修饰和RNA种类之间的潜在串扰 反映了在翻译后修饰中看到的组合控制。我们最近发现， 不同的RNA修饰酶，tRNA甲基转移酶10A（TRMT 10A）和mRNA去甲基化酶脂肪 Mass和肥胖蛋白（FTO）相互作用并共同控制基因表达。这些发现可能 代表了一种新的协同调节模式的基因调控完成的协调RNA修饰 TRMT 10A和FTO的活性。为了更好地了解这种酶复合物的功能作用，我们将 (1)定义这种RNA修饰复合物的生物化学和结构参数，以及（2）阐明 这些机制是FTO-TRMT 10A活性如何调节基因表达的基础。第一个目标将 利用生物物理工具和专业知识的阵列提供给生物化学系的优势， 生物物理学以获得野生型和突变型FTO-TRMT 10A的详细分子和结构特征 配合物第二个目标将通过采用分子生物学谱来实现， 生物化学技术在相关细胞系中分析RNA的亚细胞定位、稳定性和修饰 档案改变了宾大是一个特殊的环境，以支持我的培训和这项研究，培养一个 特别是跨多个知识和技术领域的研究人员的协作校园。无数 学习技术、使用专业设备、形成合作和分享想法的机会将 继续丰富我的研究和训练。总体而言，工具、培训和专业知识都已到位， 成功地完成了在mRNA和tRNA的新的协同调节的研究中设定的目标 修改.

Project Summary/Abstract RNA modifications are concurrently found on nearly all RNA species and can dramatically alter the properties and expression of RNA. Modifications to mRNA can affect gene expression through several mechanisms such as localization, stability, and translation efficiency. Modifications to tRNAs are known to affect their stability and abundance as well as their decoding and elongation dynamics during translation. Dysregulation of many enzyme-mediated RNA modifications have been causally linked to impaired neurological development, intellectual disabilities, and behavioral abnormalities. There has been a great effort by the field to understand the functional impact of individual modifications in isolated RNAs, but the interplay of both mRNA and tRNA modifications has never been explored. The potential for cross-talk between modifications and RNA species echoes the combinatorial control seen in post-translational modifications. We have recently discovered that two distinct RNA-modifying enzymes, tRNA methyltransferase 10A (TRMT10A) and the mRNA demethylase Fat Mass and Obesity protein (FTO), interact and function together to control gene expression. These findings may represent a novel coregulatory mode of gene regulation accomplished by the coordinated RNA-modifying activity of TRMT10A and FTO. In order to better understand the functional role of this enzyme complex, we will (1) define the biochemical and structural parameters of this RNA-modifying complex and (2) elucidate which mechanism(s) underlie how gene expression is modulated by the activity of FTO-TRMT10A. The first goal will take advantage of the array of biophysical tools and expertise available to the Department of Biochemistry and Biophysics to obtain detailed molecular and structural characteristics of wild-type and mutant FTO-TRMT10A complexes. The second goal will be accomplished by employing a spectrum of molecular biology and biochemistry techniques in relevant cell lines to analyze RNA subcellular localization, stability, and modification profile changes. UPenn is an exceptional environment to support my training and this study, fostering a particularly collaborative campus of researchers that cross many intellectual and technical fields. The countless opportunities to learn techniques, use specialized equipment, form collaborations, and share ideas will continue to enrich my research and training. Overall, the tools, training, and expertise are all in place to successfully accomplish the goals set in this proposed study of the novel coregulation of mRNA and tRNA modifications.