Nuclear organization of the Polycomb target FLC during the cold-induced epigenetic silencing of vernalization

表观遗传调控是指在缺乏初始调控因子的情况下，在许多细胞世代中，有时是通过转代方式，维持特定的基因表达状态。它对大多数生物体的正确生长、发育和环境反应至关重要。在许多情况下，表观遗传调控的分子基础与染色质的修饰、DNA的甲基化或相关组蛋白的翻译后修饰相关。例如，对控制细胞分化、形态发生和发育时间至关重要的表观遗传沉默机制涉及所谓的“多梳复合物”，这是一组蛋白复合物，其将甲基化基团添加到组蛋白3的氨基末端尾部的特定残基上。这个“标记”然后吸引其他沉默蛋白，使基因关闭。非编码RNA通常将Polycomb蛋白招募到其特定靶标，但决定其作用特异性的确切机制仍在积极研究中。每次细胞分裂时，染色质上的“标记”都需要复制到子染色质链上。对于维持表观遗传沉默状态可能非常重要的一个特征是将靶点“储存”在细胞核的某些位置，其中收集了所有类似调节的基因。在雌性哺乳动物中X染色体之一失活的情况下，表观遗传沉默后的这种核重新定位已被广泛表征，但许多其他系统中的新兴数据表明，这通常对表观遗传调控非常重要。核位置的变化如何与发生的所有其他变化联系起来仍然不清楚，因为通常很难在一个表观遗传沉默的目标上研究所有问题。在过去的15年里，Dean实验室研究了在一个称为春化的过程中，编码植物开花重要抑制因子的基因（称为FLC）的表观遗传沉默。冬季漫长的寒冷经历缓慢但肯定地关闭了这种基因的表达，这种关闭在随后的春季和夏季被记住，使植物在有利的条件下开花。与许多哺乳动物系统不同，植物系统具有广泛的遗传分析可以与分子，计算和细胞生物学技术相结合的优势。这导致FLC的调节被认为是环境介导的表观遗传调节的范例。我们希望建立在这一优势，并利用一种新的实验技术来观察在冷诱导的表观遗传沉默过程中参与春化的FLC的核组织的变化。在春化过程的不同阶段，关于核组织和染色质相互作用的新信息将通过突变体和特定转基因系的分析与其他机制信息相结合。这项工作的结果将使我们能够将细胞核组织变化的重要性与组蛋白“标记”积累、非编码RNA产生和春化作用下的细胞自主表观遗传转换的动态变化相结合。它也将提供重要的结论，更普遍地了解核组织是如何与表观遗传调控相互关联的。

Epigenetic regulation is the maintenance of specific gene expression states in the absence of the initial regulator over many cell generations, and sometimes transgenerationally. It is profoundly important for correct growth, development and environmental response in most organisms. In many cases the molecular basis of epigenetic regulation is associated with modification of chromatin, the methylation of the DNA or post-translational modification of the associated histones. For example, an epigenetic silencing mechanism central to control of cell differentiation, morphogenesis and developmental timing involves the so-called 'Polycomb complexes', a set of protein complexes that add methylation groups to a particular residue on the amino-terminal tail of histone 3. This 'mark' then attracts other silencing proteins that keep the gene switched off. Non-coding RNAs often recruit the Polycomb proteins to their specific targets but the exact mechanisms determining the specificity of their action is still being actively dissected. The 'marks' laid down on the chromatin need to be copied to the daughter chromatin strand every time a cell divides. A feature that may be very important for maintenance of the epigenetically silenced state is 'storage' of the targets in certain places in the nucleus, where all similarly regulated genes are collected. This nuclear re-positioning upon epigenetic silencing has been extensively characterized in the case of inactivation of one of the X-chromosomes in female mammals, but emerging data in many other systems suggests this is generally very important for epigenetic regulation. How a change in nuclear position ties in with all the other changes that occur is still not clear because it is often hard to study all the questions on one epigenetically silenced target. Over the last 15 years the Dean laboratory has studied the epigenetic silencing of the gene (called FLC) encoding an important repressor of flowering in plants in a process called vernalization. The very long period of cold experience during winter slowly but surely shuts off expression of this gene and this shut-down is remembered during the subsequent spring and summer, allowing the plants to flower in favourable conditions. Unlike many mammalian systems the plant system has the advantage that extensive genetic analysis can be combined with molecular, computational and cell biology techniques. This has resulted in the regulation of FLC being considered as the paradigm for environmentally-mediated epigenetic regulation. We want to build on this advantage and exploit a new experimental technique to observe changes in the nuclear organization of FLC during the cold-induced epigenetic silencing process involved in vernalization. The new information on nuclear organization and chromatin interactions, at different phases of the vernalization process, will be integrated with the other mechanistic information through analysis of mutants and specific transgenic lines. The results emerging from this work will allow us to integrate the importance of changes in nuclear organization with dynamic changes in histone 'mark' accumulation, non-coding RNA production and the cell autonomous epigenetic switching that underlies vernalization. It will also provide important conclusions for understanding more generally how nuclear organization is interconnected with epigenetic regulation.