Structural Co-evolution of the LARP Superfamily and its Role in Functional Plasticity

项目摘要/摘要 我们研究计划的长期目标是破译“非正则”蛋白质的分子机制-- RNA相互作用研究一个特别吸引人的与疾病相关的蛋白质家族，称为La相关蛋白质家族 蛋白质(LARP)超家族，在分子水平上。胞质LARPs在转录后调控中起关键作用 通过调节主动翻译、降解和储存mRNAs之间的微妙平衡来控制基因。 因此，许多LARP与导致它们的各种癌症和纤维增生性疾病密切相关。 一类重要的可用药靶点。然而，由于缺乏药物，治疗方法的发展一直受到阻碍。 详细的分子水平信息。我们的研究将阐明RNA识别的分子机制 尤其是LARPS展示的，以解释共性和个性的错综复杂，即它们是如何... MonRNA结合基序，称为La-模块，已经单独进化到允许特定的RNA识别和 从而实现其独特的功能。我们的研究基于我们在溶液和固态方面的独特优势 核磁共振波谱及其与其他生化、生物物理、计算和功能的紧密耦合 接近了。我们的初步努力将遵循两条线调查，同时集中在两名成员的 LAP超家族，hLARP6和hLARP1。在第一条线索中，我们将探索hLARP6的La-模块是如何 实现了对高度保守的5‘茎环(5’SL)基序的排他性识别，该基序在所有的垂直于 编码I型胶原蛋白的mRNAs。这条研究路线将提供详细的分子水平图，说明 La-模块的个别元件有助于5‘SL结合的特异性和亲和力。详细的见解 即将获得的数据，加上目前可用的生化和生物物理数据，将提供基本的见解 进入La-模块各个元素的分子共生，并弥合这一关键的知识鸿沟 制定针对纤维增生性疾病的治疗策略所需的。在第二行中 调查中，我们将类比地剖析hLARP1的La模块如何识别一种截然不同的RNA类型 与hLARP6相比。HLARP1被发现与增殖和细胞周期缺陷密切相关，并被认为是 在恶性细胞和组织中显著上调。最近的生化研究表明，La- HLARP1的模块依次与3‘聚(A)结合，然后与5ʹ末端的寡嘧啶(5ʹ顶部)基序结合 信使核糖核酸。值得注意的是，这种奇怪的两步行为尚未在任何其他LARP中观察到。我们会 通过研究hLARP1的结构和动态变化来揭示这一不寻常的分子机制 当Poly(A)和5‘顶端基序结合时。这项研究将首次揭示初始结合是如何 一个RNA与其La-模块结合，会引起与第二个RNA靶标结合所需的结构和动态变化。 总体而言，这两条线的比较研究将协同作用引导我们理解 将La模块的各个元件的共同进化与它们在RNA中的特定作用联系起来的原理 从而解释了结构塑性和动态塑性如何对功能塑性做出贡献。

PROJECT SUMMARY/ABSTRACT The long-term goal of our research program is to decipher the molecular mechanism of “non-canonical” protein- RNA interactions studying a particularly fascinating and disease-relevant family of proteins, called the La-related protein (LARP) superfamily, on a molecular level. Cytoplasmic LARPs play a pivotal role in post-transcriptional gene control by regulating the delicate balance between active translation, degradation, and storage of mRNAs. Hence, many LARPs are intimately implicated in various cancers and fibroproliferative diseases rendering them an important class of druggable targets. However, the development of therapies has been stunted by the lack of detailed molecular-level information. Our research will elucidate the molecular mechanism of RNA recognition exhibited by LARPs, in particular, to explain the intricacies of commonality and individuality, i.e. how their com- mon RNA-binding motif, called the La-module, has individually evolved to allow specific RNA recognition and thus achieve its distinct function. The investigation is based on our unique strength in solution and solid-state NMR spectroscopy and their close coupling with other biochemical, biophysical, computational, and functional approaches. Our initial efforts will follow two lines of inquiry, simultaneously focusing on two members of the LARP superfamily, hLARP6 and hLARP1. In the first line of inquiry, we will explore how the La-module of hLARP6 achieves the exclusive recognition of the highly conserved 5' stem-loop (5'SL) motif, which is found in all verte- brate mRNAs encoding type I collagens. This line of study will provide a detailed molecular-level map on how individual elements of the La-module contribute to the specificity and affinity of 5'SL binding. The detailed insights to be gained, together with currently available biochemical and biophysical data, will provide essential insights into the molecular “symbiosis” of the individual elements of the La-module and close this critical gap in knowledge required for the development of therapeutic strategies against fibroproliferative diseases. In the second line of inquiry, we will analogously dissect how the La-module of hLARP1 recognizes a distinctly different type of RNA compared to hLARP6. hLARP1 was found to be heavily involved in proliferation and cell cycle defects and to be significantly upregulated in malignant cells and tissues. Very recent biochemical studies revealed that the La- module of hLARP1 sequentially binds to the 3' poly(A) and then to the 5ʹ terminal oligopyrimidine (5ʹTOP) motifs of mRNAs. Notably, this peculiar two-step behavior has not yet been observed for any other LARP. We will uncover this unusual molecular mechanism by investigating the structural and dynamic changes of hLARP1 upon binding of the poly(A) and 5'TOP motifs. This study will for the first time reveal how the initial binding of one RNA to its La-module elicits structural and dynamic changes required for the binding of a second RNA target. Overall, these two lines of comparative investigations will synergistically lead us to understand the fundamental principles that connect the co-evolution of the individual elements of the La-module and their specific role in RNA recognition, thus explaining how structural and dynamic plasticity contribute to functional plasticity.