Chromatin Structure and DNA Processing in Euplotes Crassus

0078182 Jahlan-ciliated原生动物Euplotes crassus作为模型系统正在研究中，以揭示产生DNA重排的机制。 最终，研究这一系统应该有助于回答有关染色体和细胞核结构的问题，以及细胞如何操纵这些结构，使基因组保持功能稳定，同时提供基因组可塑性，这在进化和发育上都是有利的。 通过研究一个受控的重排过程，如发生在E。通过在大核形成过程中观察crassus的变化，可以确定涉及哪些染色体蛋白质，DNA序列和酶以及如何控制该过程。 E.像其他纤毛虫原生动物一样，crassus具有两种类型的细胞核，即大核和微核，它们的性质不同。 最值得注意的是，大核中不存在染色体分离，微核中不存在转录。 染色质和染色体结构和基因组组织在这两种类型的细胞核中不同。 此外，在从微核形成大核的发育过程中，发生广泛的DNA消除，包括位点特异性缺失和染色体断裂以及端粒添加。 这些重排的丰富性、它们的发育编程以及它们在单细胞生物中的发生可以在发育上同步化，为染色体结构和基因组重排机制的分析提供了独特的模型系统。 首席研究员已经确定了一个不寻常的染色质结构的高度丰富的Tec元件转座子，正在经历消除在大核发育。 此外，她的实验室已经鉴定了一种富含赖氨酸/精氨酸的组蛋白H1或HMG样“染色体支架”蛋白（p85），其具有以下特性：a）仅在发育中的大核中发现，B）与消除的DNA（包括Tec元件）相关，c）与拓扑异构酶II共定位，d）影响拓扑异构酶II的体外活性。 这表明拓扑异构酶II介导的染色体凝聚过程在纤毛虫基因组重排中的所有生物中都是保守的。 该项目旨在确定p85的序列，以便在其他生物中识别任何类似的蛋白质。 此外，p85在DNA消除中的作用将通过定义它与什么DNA序列结合以及它具有什么序列特异性以及通过确定这种蛋白质如何影响染色质结构来确定。 最后，将进一步分析p85与拓扑异构酶II的相互作用。

0078182 JahnThe ciliated protozoan Euplotes crassus is being studied as a model system in order to uncover the mechanisms producing rearrangements of DNA. Ultimately, investigating this system should help answer questions about the structure of chromosomes and the nucleus, and how cells manipulate these structures such that the genome remains functionally stable while providing genome plasticity, which can be evolutionarily and developmentally advantageous. By studying a controlled rearrangement process such as that occurring in the E. crassus during the formation of a macronucleus, it can be determined what chromosomal proteins, DNA sequences and enzymes are involved and how the process is controlled. E. crassus, like other ciliated protozoa, possesses two types of nuclei, macronuclei and micronuclei, that differ in their properties. Most notably, chromosome segregation is absent from the macronucleus and transcription is absent from the micronucleus. Chromatin and chromosome structure and genomic organization differ in these two types of nuclei. In addition, during the developmental process of forming macronuclei from micronuclei, extensive DNA elimination occurs that includes site-specific deletions and chromosome breakage with telomere addition. The abundance of these rearrangements, their developmental programming, and their occurrence in single-celled organisms that can be developmentally synchronized en masse, provide a unique model system for the analysis of chromosome structure and the mechanisms of genome rearrangements. The principal investigator has identified an unusual chromatin structure of the highly abundant Tec element transposons that are undergoing elimination during macronuclear development. In addition, her laboratory has identified a lysine/arginine-rich, histone H1 or HMG-like,'chromosome scaffold' protein (p85) that has the following properties: a) it is only found in the developing macronucleus, b) it is associated with eliminated DNA (including the Tec elements), c) it colocalizes with topoisomerase II, and d) it affects topoisomerase II activities in vitro. This points to a role for topoisomerase II-mediated, chromosome condensation processes that are conserved in all organisms in ciliate genomic rearrangements. This project aims to determine the sequence of p85 to allow identification of any similar proteins in other organisms. In addition, the role of p85 in DNA elimination will be determined by defining what DNA sequences it binds to and what sequence specificity it has, and by determining how this protein affects chromatin structure. Finally, the interaction of p85 with topoisomerase II will be further analyzed.