Genetic Characterization of Phytochrome Nuclear Bodies in Plant Light Signaling

描述（由申请人提供）：核小体（NB）是动物和植物细胞中的亚核巨蛋白或蛋白/RNA复合物，为基因表达、DNA复制和DNA修复的调节提供微环境。对动物“转录工厂”、PML小体、Cajal小体和polycomb小体的大量研究表明，某些NB是与靶基因直接相关的转录调控中心。然而，核小体和染色质之间相互作用的基本原理，包括染色质位点如何被募集到核小体以及核小体如何控制转录的分子机制，仍然是个谜。本实验室的长期目标是利用拟南芥中的光敏色素NB作为遗传模型来研究NB在真核细胞信号转导中功能的一般原理。光敏色素是一类红色和远红色光感受器，通过转录调控调控植物光依赖性发育。光敏色素的光活化和去活化触发快速NB组装和拆卸以及转录组的戏剧性重编程。然而，光敏色素NBs和基因调控之间的关系是完全未知的。我们的初步研究表明，光敏色素NBs与光敏色素抑制基因直接相关。因此，我们假设光敏色素NB是光介导的基因沉默的中心枢纽。为了进一步检验这一假设，我们提出使用染色质沉淀随后深度测序（ChIP-seq）来鉴定全局光敏色素NB相关的染色体基因座，以确定基因定位到光敏色素NB和转录活性之间的关系，并鉴定将基因基因座募集到光敏色素NB所需的顺式元件。这些实验将为光敏色素NB在基因表达中的功能提供直接证据，并开始揭示光敏色素NB招募基因的机制。更好地了解光敏色素NBs的功能不仅可以揭示植物光信号的新机制，而且可以为进化保守的核组织和转录调控机制提供更好的见解。

DESCRIPTION (provided by applicant): Nuclear bodies (NBs) are subnuclear mega-protein or -protein/RNA complexes in both animal and plant cells providing microenvironments for the regulation of gene expression, DNA replication, and DNA repair. An accumulating body of evidence from the studies on animal "transcription factories", PML bodies, Cajal bodies, and polycomb bodies suggests that some NBs are transcriptional regulatory hubs directly associated with target genes. However, the basic principles of the interaction between nuclear bodies and chromatin, including molecular mechanisms of how chromatin loci are recruited to nuclear bodies and how nuclear bodies control transcription, remain enigmatic. The long- term goal of our laboratory is to use phytochrome NBs in Arabidopsis as a genetic model to investigate general principles of NB function in eukaryotic cell signaling. Phytochromes are red and far-red photoreceptors mediating light-dependent plant development through transcriptional regulation. Photo-activation and -deactivation of phytochromes triggers rapid NB assembly and disassembly and dramatic reprogramming of the transcriptome. However, the relationship between phytochrome NBs and gene regulation is completely unknown. Our preliminary studies demonstrated that phytochrome NBs are directly associated with phytochrome-repressed genes. Therefore, we hypothesize that phytochrome NBs are central hubs for light-mediated gene silencing. To further test this hypothesis, we propose to identify global phytochrome NB associated chromosome loci using chromatin- precipitation followed by deep sequencing (ChIP-seq), to determine the relationship between gene positioning to phytochrome NBs and transcriptional activity, and to identify cis-elements required for recruiting gene loci to phytochrome NBs. These experiments will provide direct evidence for the function of phytochrome NBs in gene expression and begin to uncover mechanisms involved in recruiting genes to phytochrome NBs. A better understanding of the function of phytochrome NBs will not only uncover novel mechanisms of light signaling in plants, but also provide great insight into evolutionarily conserved mechanisms of nuclear organization and transcriptional regulation.