Understanding the Role of Nucleosome Turnover in the Malaria Parasite

描述（由申请人提供）：疟疾仍然是世界上最致命的传染病之一。为了确定新的治疗策略，我们必须全面了解疟原虫如何调节其复杂的生命周期。这个提议的目标是阐明协调染色质重组的机制。更具体地说，我们探讨是否核小体定位，占用和营业额直接发育程序的寄生虫感染周期的过程中。越来越多的证据表明，染色质的表观遗传状态在寄生虫基因表达和致病性中起作用。而组蛋白水平在大多数真核生物保持相当恒定的整个细胞周期，我们已经发现了“组蛋白崩溃”正是在最转录活跃阶段的寄生虫红细胞周期。因此，我们假设恶性疟原虫调节组蛋白周转和定位的独特方式可能以特定方式控制寄生虫基因表达。使用两种互补的方法，我们将测量整个寄生虫生命周期的无组蛋白和组蛋白结合的DNA含量。将使用下一代测序技术（Illumina（R））分析DNA富集区域。结果将生成与稳态mRNA模式相关的“开放”和“闭合”染色质区域的图谱。为了确定组蛋白翻译后修饰是否调节组蛋白周转，我们将使用质谱法。阶段特异性组蛋白标记将通过ChIP-seq实验进行验证。进一步的质谱和功能基因组学研究将用于确定参与调节组蛋白周转的染色质重塑复合物。使用这些独立的数据集，我们试图探索组蛋白定位，组蛋白翻译后修饰和寄生虫转录组之间的联系。揭示控制组蛋白周转的分子机制不仅可以让我们找到干扰转录表达波的新方法，还可以改变我们对在染色质水平控制寄生虫转录的分子机制的看法，并为新的抗疟疾疗法提供路标。 公共卫生相关性：该提案的目标是阐明协调染色质重组的机制。拟议的研究探讨是否核小体定位，占用和营业额直接发育程序在寄生虫感染周期的过程中。

DESCRIPTION (provided by applicant): Malaria remains one of the most deadly infectious diseases in the world. To identify new therapeutic strategies, we must gain a comprehensive understanding of how the malaria parasite regulates its complex life cycle. The goal of this proposal is to elucidate the mechanisms that orchestrate chromatin restructuring. More specifically, we explore whether nucleosome positioning, occupancy and turnover direct developmental programs over the course of the parasite infectious cycle. There is growing evidence that the epigenetic state of chromatin plays a role in parasite gene expression and pathogenicity. Whereas histone levels in most eukaryotes remain fairly constant across the cell cycle, we have found a "histone crash" precisely at the most transcriptionally active stage of the parasite erythrocytic cycle. Accordingly, we hypothesize that the unique manner in which P. falciparum regulates histone turnover and positioning may control the parasite gene expression in a specific manner. Using two complementary methods, we will measure histone-free and histone-bound DNA content across the parasite life cycle. DNA-enriched regions will be analyzed using next generation sequencing technology (Illumina(R)). Results will generate maps of "open" and "closed" chromatin regions that will be correlated with steady state mRNA patterns. To determine whether histone post-translational modifications regulate histone turnover, we will use a mass spectrometry approach. Stage-specific histone marks will be validated by ChIP-seq experiments. Further mass spectrometry and functional genomic studies will then be used to determine the chromatin-remodeling complexes that are involved in regulating histone turnover. Using these independent data sets, we seek to explore a connection between histone positioning, histone post-translational modifications and the parasite transcriptome. Revealing the molecular mechanisms that control histone turnover will not only allow us to find new ways to interfere with waves of transcriptional expression but will also transform our perceptions of the molecular mechanisms controlling the parasite transcription at the chromatin level and provide signposts for new anti-malarial therapies. PUBLIC HEALTH RELEVANCE: The goal of this proposal is to elucidate the mechanisms that orchestrate chromatin restructuring. The proposed research explores whether nucleosome positioning, occupancy and turnover direct developmental programs over the course of the parasite infectious cycle.