Mechanisms Governing DNA Replication Timing and Copy Number Control During Development

真核生物的基因组包含数百万到数十亿的DNA碱基对，每次细胞分裂时都必须精确地复制这些碱基对。不能准确复制DNA会导致改变细胞功能的突变，甚至导致细胞死亡。考虑到复制DNA的复杂性，这个过程可能需要很长时间才能完成。DNA不是随机复制的，而是在基因组复制的特定时间复制特定的DNA序列。该项目将利用跨学科的方法来了解关键因素如何控制特定DNA序列的复制。该项目还将为教师和学生提供教育和研究机会。首先，我们将为纳什维尔地区的高中教师提供直接和动手研究的机会，以帮助开发以实验为基础的课程。其次，我们将在课堂上开发新的方法，使本科生和研究生能够使用跨学科的方法来解决生物学中未解决的问题。最后，我们将为范德比尔特大学的本科生提供动手研究的机会。Rif1已被证明可以控制从酵母到人类的复制时间程序，但Rif1如何精确地执行这一功能仍不清楚。该项目旨在确定在发育过程中如何调节Rif1活性，从而帮助确定复制定时程序。使用果蝇作为模型系统，我们发现Rif1以发育编程的方式与复制机制相关。我们将结合果蝇强大的发育系统，结合生物化学和蛋白质组学方法来确定Rif1如何与复制机制相关联。然后，我们将使用遗传学和基因组学策略的组合来确定Rif1如何影响不同发育背景下的复制时间。最后，我们将应用染色体构象捕获技术来探究Rif1调节复制定时程序的分子机制。该奖项反映了美国国家科学基金会的法定使命，并通过使用基金会的知识价值和更广泛的影响审查标准进行评估，被认为值得支持。

Mechanisms governing DNA replication timing and copy number control during development The genomes of eukaryotic organisms contain millions to billions of base pairs of DNA that must be accurately copied every time a cell divides. Failure to accurately copy DNA can cause mutations that alter cell function or even lead to cell death. Given the complexity of copying DNA, this process can take many hours to complete. DNA is not copied randomly, but rather particular DNA sequences are replicated at specific times during the period of genome replication. This project will utilize interdisciplinary approaches to understand how key factors control when a specific DNA sequence is replicated. The project will also provide educational and research opportunities to benefit both teachers and students. First, we will provide direct and hands-on research opportunities for Nashville area high school teachers to aid in the development of an experimental-based curriculum. Second, we will develop new approaches in the classroom to expose undergraduate and graduate students to the power of using interdisciplinary approaches to solve unanswered questions in biology. Lastly, we will provide hands-on research opportunities to undergraduate students at Vanderbilt.Rif1 has been shown to control the replication timing program from yeast to humans, but precisely how Rif1 performs this function remains unknown. This project aims to identify how Rif1 activity is regulated during development and thus help determine the replication timing program. Using Drosophila melanogaster as a model system, we have found that Rif1 associates with the replication machinery in a developmentally programmed manner. We will combine the powerful developmental systems of Drosophila, with biochemical and proteomic approaches to determine how Rif1 associates with the replication machinery. We will then use a combination of genetics and genomics strategies to determine how Rif1 influences replication timing in different developmental contexts. Finally, we will apply chromosome conformation capture techniques to interrogate the molecular mechanisms that Rif1 utilizes to regulate the replication timing program.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.