Epigenetic control of transcription termination

染色质结构塑造真核生物基因组的表观遗传景观，并在基因转录调控中发挥重要作用。染色质如何控制RNA聚合酶II（RNAPII）的转录终止还知之甚少。本项目的目的是描述染色质在转录终止调控中的作用和功能。终止转录是一个复杂的过程，涉及转录RNAPII的分解和新生RNA的同时3'加工。转录终止位点（TTS）下游的转录必须被有效地终止，以避免下游基因的入侵和异常表达。转录终止也通过基因环反馈到转录起始。这些离散的通路之间的协调是如何完成的，人们还知之甚少。真核基因组被包装并组织成染色质核蛋白结构。染色质调节因子的协同作用改变了基因组的核小体组织，指挥决定转录输出的表观遗传信息。染色质畸变导致有缺陷的转录终止。然而，染色质在终止中的作用仍然知之甚少。ATP依赖的染色质重塑酶复合物INO 80在真核生物中高度保守。在植物中，INO 80的缺失导致严重的发育缺陷，如矮化和开花晚，并且其活性是大量基因的适当表达所必需的，包括FLC（FLOWERLOCUS C），其是控制开花时间的主要调节因子。这表明INO 80在植物发育中的重要但尚不清楚的作用。MPC最近在酿酒酵母中证明INO 80促进RNAPII从染色质中的去除（Lafon等人，Mol.Cell 2015）并促进转录终止和mRNA稳定化（Luzzi et al.，提交）。然而，INO 80介导的终止机制尚不清楚，我们的工作揭示了一个核小体调控网络作为转录终止过程的新参与者。该项目将测试以下假设：TTS处潜在染色质结构的动态调节定义了一种未表征的途径，该途径协调RNA加工与RNAPII的释放，确保功能性mRNA的有效终止和表达。我们会问：-INO 80如何调节转录终止并将其与mRNA稳定性结合？-染色质对转录终止的调控如何与植物可塑性的时空调控相结合？我们的策略采用了一个强大的组合基因组学，高通量遗传学和染色质生化分析的模式生物酿酒酵母和拟南芥。这种方法将揭示染色质和终止机制之间的物理和功能串扰，并阐明这种新的转录调控途径的分子基础。

Chromatin structure shapes the epigenetic landscape of the eukaryotic genome and plays an essential role in gene transcriptional regulation. How chromatin controls termination of transcription by RNA Polymerase II (RNAPII) is poorly understood. The objective of this project is to characterize the role and function of chromatin in regulation of transcriptional termination. Terminating transcription is an intricate process, involving disassembly of the transcribing RNAPII and simultaneous 3' processing of the nascent RNA. Transcription downstream the transcription termination site (TTS) has to be efficiently terminated to avoid invasion and aberrant expression of downstream genes. Transcriptional termination also feeds back to transcriptional initiation, through gene looping. How coordination between these discrete pathways is accomplished is poorly understood. The eukaryotic genome is packaged and organized into the chromatin nucleoprotein structure. The concerted actions of chromatin regulators alters the nucleosomal organization of the genome, commanding epigenetic information that dictates transcriptional output. Chromatin aberrations lead to defective transcription termination. However, the role of chromatin in termination remains poorly understood.The ATP-dependent chromatin remodelling enzymatic complex INO80 is highly conserved across eukaryotes. In plants, INO80 loss causes severe developmental defects such as dwarfisms and late flowering, and its activity is required for the proper expression of a large number of genes, including FLC (FLOWERING LOCUS C) a master regulator of flowering time control. This indicates an important, yet unclear role for INO80 in plant development. MPC recently demonstrated in S.cerevisiae that INO80 facilitates removal of RNAPII from chromatin (Lafon et al., Mol.Cell 2015) and promotes transcription termination and mRNA stabilization (Luzzi et al., submitted). However, the INO80-mediated mechanism in termination is not known.Our work revealed a network of nucleosome regulators as novel players of the transcriptional termination process. This project will test the hypothesis that dynamic regulation of the underlying chromatin structure at the TTS defines an uncharacterized pathway that coordinates RNA processing with release of RNAPII, ensuring efficient termination and expression of functional mRNAs. We will ask:- How does INO80 regulate transcription termination and couples it with mRNA stability?- How does chromatin regulation of transcription termination integrate with the spatio-temporal regulation of plant plasticity?Our strategy employs a powerful combination of genomics, high-throughput genetics and chromatin biochemical assays in the model organisms S.cerevisiae and Arabidopsis thaliana. This approach will uncover the physical and functional crosstalk between chromatin and the termination machinery and elucidate the molecular underpinnings of this novel transcriptional regulatory pathway.