RUI: Studying B-bodies, a novel type of nuclear domains: from organization to practical exploitation.

这个项目研究管理细胞核组织的基本原则。细胞核对生物技术具有特殊的重要性，因为它存储着控制细胞和有机体特征的遗传信息。生物技术的持续成功在很大程度上取决于对核内部过程的更好理解。这个项目将专门专注于研究使某些蛋白质在细胞核内形成独特和孤立结构的分选机制。在另一组单独的实验中，从这个项目中学到的信息将被应用于改进目前的生物技术方法。期望的结果是使基因操作更有效、更安全。作为更广泛的影响，该项目将在肯纳索州立大学(KSU)提供一个教育平台，用当代分子生物学和基因工程的方法培训本科生和研究生。科罗拉多州立大学是一所动态发展的、以本科生为主的机构，致力于改善其研究环境和扩大研究生课程。在细胞核中没有内膜的情况下，核蛋白被组织成紧凑的、不同的区域，这些区域的边界被称为核域(NDS)。蛋白质分选到NDS中的调节和功能意义尚不清楚。该项目将使用一种新发现的名为B-Body的果蝇ND及其常驻RNA结合蛋白Bruno作为模型来研究ND的组装和功能。假设Bruno的蛋白质序列和非编码RNA的组合足以产生B-小体。分子生物学技术，包括诱变和原位RNA杂交，将被应用于鉴定重建B小体的必要成分。下一步，将测试B小体作为异位表达核蛋白的安全溢出部位的作用。包括DNA酶和转录因子在内的异源蛋白将被靶向B小体。生化分析和遗传记者将定量地表明，将蛋白质转移到B小体是否可以减少经常困扰过度表达实验的非特异性影响和毒性。这一奖项反映了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。

This project studies basic principles that govern organization of the cell nucleus. The nucleus has a special importance for biotechnology because it stores genetic information that controls traits of cells and organisms. Continuous success of biotechnology greatly depends on better understanding of the processes inside of the nucleus. This project will specifically focus on studying the sorting mechanism that enables certain proteins to form distinct and isolated structures within the nucleus. In a separate set of experiments, the information learned in this project will be applied to improve the current biotechnology methods. The desired outcome is to make genetic manipulations more efficient and safer. As a broader impact, this project will offer an educational platform at Kennesaw State University (KSU) to train undergraduate and graduate students with contemporary methods of molecular biology and genetic engineering. KSU is a dynamically developing, predominantly undergraduate institution that strives to enhance its research environment and expand graduate programs.In the absence of internal membranes in the nucleus, nuclear proteins are organized into compact, distinct areas with sharp boundaries that are called nuclear domains (NDs). The regulation and functional significance of protein sorting into NDs is not clear. This project will use a newly discovered Drosophila ND called a B-body and its resident RNA-binding protein called Bruno, as a model to study ND assembly and function. It is hypothesized that a combination of Bruno’s protein sequence and non-coding RNA is sufficient to create B-bodies. Molecular biology techniques, including mutagenesis and in situ RNA hybridization, will be applied to identify the essential components to reconstitute B-bodies. Next, B-bodies will be tested for their role as safe overflow sites for ectopically expressed nuclear proteins. Heterologous proteins including DNA enzymes and transcription factors will be targeted to B-bodies. Biochemical assays and genetic reporters will quantitatively indicate whether protein trafficking into B-bodies can reduce non-specific effects and toxicity that often plague overexpression experiments.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.