DUAL SCALE COMPUTING WITH RNA

RNA 双尺度计算

• 批准号: 7601404
• 负责人: David Mathews
• 金额: $ 0.03万
• 依托单位: CARNEGIE-MELLON UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2007
• 资助国家: 美国
• 起止时间: 2007-08-01 至 2008-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7601404
• 关键词: Acylation; Algorithms; Amber; Aspartic Acid-Specific tRNA; Awareness; Base Pairing; Benchmarking; Biodiversity; Bioinformatics; Biology; Cell physiology; Chemicals; Chemistry; Code; Collaborations; Computational Biology; Computer Retrieval of Information on Scientific Projects Database; Computer software; Dependence; Development; Dinucleoside Phosphates; Disease; Equilibrium; Escherichia coli; Free Energy; Frequencies; Functional RNA; Funding; GPR17 gene; Gene Expression; Gene Expression Regulation; Genes; Genetic Carriers; Genome; Genomics; Goals; Grant; Hour; Hydroxyl Radical; Immunity; In Vitro; Institution; Maps; Measurement; Mechanics; Messenger RNA; Methods; Modification; Molecular Models; Myotonic Dystrophy; N-methylisatoic anhydride; Nature; Nucleotides; Object Attachment; Pathway interactions; Peptides; Phase; Play; Primer Extension; Property; Proteins; RNA; RNA Editing; RNA Interference; RNA Sequences; RNA Splicing; Reaction; Reading; Research; Research Institute; Research Personnel; Resolution; Resource Allocation; Resources; Role; Salmonella typhi; Scanning; Science; Simulate; Singapore; Source; Structure; Techniques; Testing; Thalassemia; Thermodynamics; Time; Transcript; Transfer RNA; United States National Institutes of Health; Week; Work; base; computer program; improved; interest; molecular dynamics; novel; phosphodiester; programs; protein expression; quantum; reaction rate; simulation; tool

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Our understanding of the role of RNA in biology has expanded enormously over the last two decades. Originally, RNA was understood to participate in protein expression as a passive carrier of genetic information (mRNA) and as an adapter molecule (tRNA) for reading the code. Then RNA was discovered to catalyze reactions, first self-splicing, then phosphodiester bond cleavage, and, recently, peptide bond formation. RNA is now known to play important functions in many diverse cellular processes, such as development, immunity, RNA editing and modification, and post-transcriptional gene regulation. RNA is also an important player in many diseases, including Prader-Willi, b-thalassemia, and myotonic dystrophy. With the increasing awareness of RNAs biological diversity, the ability to harness RNA as a tool has increased. RNA sequences can be evolved in vitro to catalyze many reactions that are not part of the natural repertoire. Antisense and RNAi can be used to modulate gene expression. Our group is interested in developing new computational biology tools and applying these tools to understanding RNA structure and function. We are currently working on four projects with extensive computational requirements: 1) We have developed a method for finding novel non-coding RNA (ncRNA) genes in genomic sequence. These are RNA sequences that function directly, without coding for a protein. Our method relies on our Dynalign algorithm to determine the lowest free energy structure common to two, unaligned sequences (Mathews, 2005; Mathews & Turner, 2002). Folding free energies, as determined by nearest neighbor parameters (Mathews et al., 2004; Mathews et al., 1999), of two ncRNA sequences are significantly lower than folding free energies of random sequences of identical dinucleotide-frequency content. Dynalign requires no sequence identity to find the common structure. We have therefore found that we can discover homologous ncRNA genes in crudely aligned genome fragments with greater sensitivity than other methods, especially at low sequence identity. 2) We have incorporated nudged elastic band (NEB) (Jnsson et al., 1998) into the AMBER molecular dynamics package (Pearlman et al., 1995) in collaboration with Dr. David Case, The Scripps Research Institute. NEB provides a time-scale independent method for finding low-energy pathways for conformational changes. We are currently applying NEB to understanding strand invasion, by which intermolecular base pairs displace intramolecular base pairs. This is important for understanding RNAi and antisense mechanisms. 3) We are using free energy calculations to understand the nature of SHAPE mapping of RNA structure. SHAPE maps RNA structures using N-methylisatoic anhydride to acylate the 2 OH of flexible RNA nucleotides (Merino et al., 2005; Wilkinson et al., 2005). It is known that the acylation reaction rate is dependent on the pKa of the 2 OH, but it is unclear why flexible nucleotides have lower pKas. Our simulations with AMBER using tRNA structures will be used to determine the connection. 4) We are developing new methods based on the Jarzynski equality to determine unfolding free energies of RNA hairpins on reasonable timescales (Jarzynski, 1997). Our goal is to test whether the AMBER forcefield can reproduce unfolding free energies found experimentally by mechanical pulling (Liphardt et al., 2002; Liphardt et al., 2001). Ultimately, we will model the molecular-level details of mechanical unfolding of RNA. For this Development Application to Teragrid, we propose to use most of the allocation in finding non-coding RNA sequences (project 1 above). To start, we will scan an alignment of the E. coli genome to S. Typhi that we have constructed using MUMmer 2 (Kurtz et al., 2004). We estimate that this will require 6,000 CPU hours on a 3 GHz Intel Pentium 4. We also plan use the balance of the allocation to benchmark our methods outlined in projects 2-4 and to start the scan of another genome for ncRNA genes. This will allow us to prepare a Medium Resource Allocation for Teragrid resources in the next 4 to 6 months. References: Jarzynski, C. (1997). Nonequilibrium equality for free energy differences. Phys. Rev. Lett. 78, 2690-2693. Jnsson, H., Mills, G. & Jacobsen, K. W. (1998). Nudged elastic band method for finding minimum energy paths of transitions. In Classical and Quantum Dynamics in Condensed Phase Simulations (Berne, B. J., Ciccoti, G. & Coker, D. F., eds.), pp. 385-404. World Scientific, Singapore. Kurtz, S., Phillippy, A., Delcher, A. L., Smoot, M., Shumway, M., Antonescu, C., et al. (2004). Versatile and open software for comparing large genomes. Genome Biol 5, R12. Liphardt, J., Dumont, S., Smith, S. B., Tinoco, I., Jr. & Bustamante, C. (2002). Equilibrium information from nonequilibrium measurements in an experimental test of Jarzynski's equality. Science 296, 1832-5. Liphardt, J., Onoa, B., Smith, S. B., Tinoco, I. J. & Bustamante, C. (2001). Reversible unfolding of single RNA molecules by mechanical force. Science 292, 733-7. Mathews, D. H. (2005). Predicting a set of minimal free energy RNA secondary structures common to two sequences. Bioinformatics 21, 2246-2253. Mathews, D. H., Disney, M. D., Childs, J. L., Schroeder, S. J., Zuker, M. & Turner, D. H. (2004). Incorporating chemical modification constraints into a dynamic programming algorithm for prediction of RNA secondary structure. Proc. Natl. Acad. Sci. USA 101, 7287-7292. Mathews, D. H., Sabina, J., Zuker, M. & Turner, D. H. (1999). Expanded sequence dependence of thermodynamic parameters provides improved prediction of RNA Secondary Structure. J. Mol. Biol. 288, 911-940. Mathews, D. H. & Turner, D. H. (2002). Dynalign: An algorithm for finding the secondary structure common to two RNA sequences. J. Mol. Biol. 317, 191-203. Merino, E. J., Wilkinson, K. A., Coughlan, J. L. & Weeks, K. M. (2005). RNA structure analysis at single nucleotide resolution by selective 2'-hydroxyl acylation and primer extension (SHAPE). J Am Chem Soc 127, 4223-31. Pearlman, D. A., Case, D. A., Caldwell, J. W., Ross, W. S., Cheatham, T. E., III, DeBolt, S., et al. (1995). AMBER, a package of computer programs for applying molecular dynamics and free energy calculations to simulate the structural and energetic properties of molecules. Comp. Phys. Commun. 91, 1-41. Wilkinson, K. A., Merino, E. J. & Weeks, K. M. (2005). RNA SHAPE chemistry reveals nonhierarchical interactions dominate equilibrium structural transitions in tRNA(Asp) transcripts. J Am Chem Soc 127, 4659-67.

该子项目是利用该技术的众多研究子项目之一 资源由 NIH/NCRR 资助的中心拨款提供。子项目和 研究者 (PI) 可能已从 NIH 的另一个来源获得主要资金， 因此可以在其他 CRISP 条目中表示。列出的机构是 对于中心来说，它不一定是研究者的机构。 在过去的二十年里，我们对 RNA 在生物学中的作用的理解有了极大的扩展。最初，RNA 被认为作为遗传信息 (mRNA) 的被动载体和用于读取代码的接头分子 (tRNA) 参与蛋白质表达。然后人们发现RNA可以催化反应，首先是自剪接，然后是磷酸二酯键断裂，最近是肽键形成。现在已知 RNA 在许多不同的细胞过程中发挥重要作用，例如发育、免疫、RNA 编辑和修饰以及转录后基因调控。 RNA 在许多疾病中也发挥着重要作用，包括 Prader-Willi、b 地中海贫血和强直性肌营养不良。随着人们对 RNA 生物多样性的认识不断增强，利用 RNA 作为工具的能力也随之增强。 RNA序列可以在体外进化来催化许多不属于自然反应的反应。反义和RNAi可用于调节基因表达。我们小组有兴趣开发新的计算生物学工具并应用这些工具来理解 RNA 结构和功能。我们目前正在开展四个具有广泛计算需求的项目：1) 我们开发了一种在基因组序列中寻找新型非编码 RNA (ncRNA) 基因的方法。这些是直接发挥作用的 RNA 序列，无需编码蛋白质。我们的方法依靠 Dynalign 算法来确定两个未对齐序列共有的最低自由能结构（Mathews，2005；Mathews & Turner，2002）。由最近邻参数确定的两个 ncRNA 序列的折叠自由能 (Mathews et al., 2004; Mathews et al., 1999) 明显低于具有相同二核苷酸频率含量的随机序列的折叠自由能。 Dynalign 无需序列同一性即可找到共同结构。因此，我们发现，与其他方法相比，我们可以在粗略比对的基因组片段中以更高的灵敏度发现同源 ncRNA 基因，特别是在序列同一性较低的情况下。 2) 我们与 Scripps 研究所的 David Case 博士合作，将轻推弹性带 (NEB)（Jnsson 等人，1998）纳入 AMBER 分子动力学软件包（Pearlman 等人，1995）。 NEB 提供了一种独立于时间尺度的方法来寻找构象变化的低能量途径。我们目前正在应用 NEB 来理解链入侵，即分子间碱基对取代分子内碱基对。这对于理解 RNAi 和反义机制很重要。 3) 我们使用自由能计算来理解 RNA 结构的 SHAPE 映射的本质。 SHAPE 使用 N-甲基靛红酸酐酰化柔性 RNA 核苷酸的 2 个 OH 来绘制 RNA 结构图（Merino 等人，2005 年；Wilkinson 等人，2005 年）。已知酰化反​​应速率取决于2 OH的pKa，但尚不清楚为什么柔性核苷酸具有较低的pKa。我们使用 tRNA 结构对 AMBER 进行的模拟将用于确定连接。 4）我们正在开发基于 Jarzynski 等式的新方法，以确定合理时间尺度上 RNA 发夹的展开自由能（Jarzynski，1997）。我们的目标是测试 AMBER 力场是否可以重现通过机械拉力实验发现的展开自由能（Liphardt 等人，2002 年；Liphardt 等人，2001 年）。最终，我们将模拟 RNA 机械展开的分子水平细节。对于 Teragrid 的开发应用程序，我们建议将大部分分配用于查找非编码 RNA 序列（上面的项目 1）。首先，我们将扫描我们使用 MUMmer 2 构建的大肠杆菌基因组与伤寒沙门氏菌的比对（Kurtz 等，2004）。我们估计这将需要 3 GHz Intel Pentium 4 上的 6,000 个 CPU 小时。我们还计划使用分配的余额来对项目 2-4 中概述的方法进行基准测试，并开始扫描另一个基因组中的 ncRNA 基因。这将使我们能够在未来 4 到 6 个月内为 Teragrid 资源准备中等资源分配。参考文献：Jarzynski, C. (1997)。自由能差异的非平衡平等。物理。莱特牧师。 78、2690-2693。 Jnsson, H.、Mills, G. 和 Jacobsen, K. W. (1998)。用于寻找跃迁最小能量路径的微移弹性带方法。 《凝聚相模拟中的经典和量子动力学》（Berne, B. J.、Ciccoti, G. 和 Coker, D. F. 编辑），第 385-404 页。世界科学，新加坡。 Kurtz, S.、Phillippy, A.、Delcher, A. L.、Smoot, M.、Shumway, M.、Antonescu, C. 等。 （2004）。用于比较大型基因组的多功能开放软件。基因组生物学 5，R12。 Liphardt, J.、Dumont, S.、Smith, S. B.、Tinoco, I., Jr. 和 Bustamante, C. (2002)。 Jarzynski 等式实验测试中非平衡测量的平衡信息。科学 296, 1832-5。 Liphardt, J.、Onoa, B.、Smith, S. B.、Tinoco, I. J. 和 Bustamante, C. (2001)。通过机械力可逆地展开单个 RNA 分子。科学 292, 733-7。马修斯，D.H.（2005）。预测两个序列共有的一组最小自由能 RNA 二级结构。生物信息学 21, 2246-2253。 Mathews, D. H.、Disney, M.D.、Childs, J. L.、Schroeder, S.J.、Zuker, M. 和 Turner, D. H. (2004)。将化学修饰约束纳入动态编程算法中以预测 RNA 二级结构。过程。国家。阿卡德。科学。美国 101，7287-7292。 Mathews, D. H.、Sabina, J.、Zuker, M. 和 Turner, D. H. (1999)。热力学参数的扩展序列依赖性提供了对 RNA 二级结构的改进预测。 J.莫尔。生物。 288、911-940。马修斯，D. H. 和特纳，D. H. (2002)。 Dynalign：一种用于查找两个 RNA 序列共有的二级结构的算法。 J.莫尔。生物。 317、191-203。 Merino, E. J.、Wilkinson, K. A.、Coughlan, J. L. 和 Weeks, K. M. (2005)。通过选择性 2'-羟基酰化和引物延伸 (SHAPE) 以单核苷酸分辨率进行 RNA 结构分析。美国化学学会杂志 127, 4223-31。 Pearlman, D. A.、Case, D. A.、Caldwell, J. W.、Ross, W. S.、Cheatham, T. E., III、DeBolt, S. 等。 （1995）。 AMBER，一套计算机程序，用于应用分子动力学和自由能计算来模拟分子的结构和能量特性。比较。物理。交流。 91、1-41。 Wilkinson, K. A.、Merino, E. J. 和 Weeks, K. M. (2005)。 RNA SHAPE 化学揭示了 tRNA(Asp) 转录本中的平衡结构转变以非层次相互作用为主。美国化学学会杂志 127, 4659-67。