NSF-BSF: Deciphering the factors that govern the temperature-dependence of ADAR reactivity and dsRNA substrate recognition

RNA编辑是在包括人类在内的许多生物体中发现的一种过程，能够拦截遗传信息并改变编码信息。这使得一系列不同的蛋白质可以由单个基因合成。不同生物体通过编辑调节RNA中信息含量的能力可能会对生物体如何应对它们遇到的压力(例如温度变化、接触有害物质或病原体等)产生重大影响。这项合作工作涉及美国的两个实验室和以色列的两个实验室，他们将研究来自不同生物体的名为ADAR(作用于RNA的腺苷脱氨酶)的RNA编辑酶。这些努力将增进我们对有机体如何利用编辑在其独特的环境中茁壮成长的理解。该项目为跨学科培养分子生物学、核酸化学、酶学和结构生物学的学生提供了一个独特的机会。每个实验室将贡献不同的学科努力，以实现对ADAR作为体温函数的RNA编辑的深入理解，这在不断变化的气候中至关重要。该团队将通过将我们的成果传播给感兴趣的社区，包括RNA编辑科学界、研究生、本科生、K-12学生和普通公众，最大限度地扩大研究发现的更广泛影响。拟议的新工作是NSF和BSF的联合拨款；它将由Peter Beal教授和Andrew Fisher教授领导的美国团队以及Shay Ben-Aroya教授和Erez Levanon教授领导的以色列团队进行。Ben-Aroya/Levanon团队已经建立了面包师酵母S.cerevisiae作为一个通用的筛选平台，以评估来自不同生物的ADAR的编辑活性。利用这一平台，他们确定了鸟类ADAR是酵母中高效的腺苷到肌苷(A到I)编辑者，并提出了这是鸟类体温过高的结果的假说。在这个项目中，所有四个实验室将共同确定导致高度编辑活动的鸟类ADAR的关键特征，并确定其分子基础。这将通过结构域交换和随机突变以及在酵母中生长效应的额外筛选来实现。纯化的禽类ADAR蛋白将进行RNA结合、脱胺速率测量和ADAR-RNA复合体的结构研究。最后，来自生活在不同环境中的其他生物体的ADAR将被描述为定义所观察到的现象的一般性。该奖项反映了NSF的法定使命，并通过使用基金会的智力优势和更广泛的影响审查标准进行评估，被认为值得支持。

RNA editing is a process found in many organisms, including humans, capable of intercepting genetic messages and changing the encoded information. This allows for a range of different proteins to be made from a single gene. The ability of different organisms to modulate the information content in RNA through editing is likely to have a substantial impact on how organisms’ respond to stresses they encounter (e.g. changes in temperature, exposure to harmful substances or pathogens, etc). This collaborative effort involving two labs in the U.S. and two labs in Israel will investigate RNA editing enzymes called ADARs (adenosine deaminases that act on RNA) from different organisms. These efforts will advance our understanding of how editing can be used by an organism to thrive in its unique environment. This project provides a unique opportunity for interdisciplinary training of students spanning molecular biology, nucleic acid chemistry, enzymology, and structural biology. Each lab will contribute a different disciplinary effort to achieve in-depth understanding of RNA editing by ADARs as a function of body temperature, which is critically important in a changing climate. The team will maximize the broader impact of the research discoveries by disseminating our results to interested communities including the RNA editing scientific community, graduate students, undergraduate students, K-12 students and the general public. The proposed new work is a joint NSF-BSF grant; it will be carried out by an American team directed by Professors Peter Beal and Andrew Fisher and an Israeli effort led by Professors Shay Ben-Aroya and Erez Levanon. The Ben-Aroya/Levanon team have established the baker’s yeast, S. cerevisiae, as a versatile screening platform to evaluate the editing activity of ADARs from different organisms. Using this platform, they identified bird ADARs as highly efficient adenosine-to-inosine (A to I) editors in yeast and have advanced the hypothesis that this is a consequence of the high body temperature of birds. In this project, all four labs will work together to define the critical features of bird ADARs that lead to the high editing activity and define its molecular basis. This will be accomplished using domain swapping and random mutagenesis coupled with additional screens for growth effects in yeast. Purified bird ADAR proteins will be subjected to RNA binding, deamination rate measurements and structural studies of ADAR-RNA complexes. Finally, ADARs from other organisms, that inhabit different environmental niches, will be characterized to define the generality of the observed phenomenon.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.