Unravelling the functional role of the “epigenetic reader” MRG-1 in the spatial organization of the C. elegans genome

根据其转录状态，染色质主要分为两大类：活性常染色质和转录抑制和紧凑的异染色质。在物种间，常染色质和异染色质的空间分布不是随机的，并使基因组能够进行功能划分。特别是，沉默的异染色质主动地隔离在核的外围，而活性常染色质位于中央。尽管有如此惊人的进化保守，但异染色质的空间隔离在多大程度上对基因组功能的贡献尚不完全清楚。为了正式解决这个问题，需要确定产生核亚室的因素和机制，以及它们对基因表达的扰动的后果。在我最近的博士后工作中，我鉴定了MRG-1，它是活性染色质室的第一个组分，被证明是形成线虫肠道内异染色质空间分布的关键。尽管如此，关于这一规定是如何实施的，仍然存在根本的悬而未决的问题。特别的是，MRG-1携带一个能“读取”甲基化的组蛋白的染色质桶结构域。然而，决定MRG-1募集和功能的染色质信号尚不清楚。此外，染色质阅读器并不是单独行动的，通常是目标站点复合体的招聘人员。然而，负责异染色质组织的含有MRG-1的复合体是完全未知的。在这个提议中，我们的目标是确定i)参与MRG-1招募并有助于其在异染色质结构中的作用的组蛋白标记和相应的染色质修饰物，以及ii)在线虫肠道中MRG-1参与异染色质组织的复合体/蛋白质相互作用。为了实现我们的目标，我的团队将结合显微镜、遗传和生化方法。此外，最先进的分子生物学分析，如Emerin-DamID和RNAseq，将为我们提供相关突变体在肠道中染色质区划和基因表达的完整图景。通过这项工作，我们将识别在整个有机体的分化组织中塑造基因组3D组织的新玩家。此外，我们将能够在组织特异性水平上确定它们与基因表达的相关性。由于MRG-1和大多数染色质因子在蠕虫和哺乳动物之间高度保守，这项工作可能在表观遗传学和核结构领域产生广泛影响。

Largely based on its transcriptional status, chromatin falls into two main classes: active euchromatin and transcriptionally repressed and compacted heterochromatin. Across species, the spatial distribution of euchromatin and heterochromatin is not random and enables a functional compartmentalization of the genome. In particular, silent heterochromatin is actively sequestered at the nuclear periphery, while active euchromatin is centrally located. Despite such a striking evolutionary conservation, to which degree the spatial sequestration of heterochromatin contributes to genome function is not fully understood. To formally address this, the factors and mechanisms that generate nuclear sub-compartments need to be identified and the consequences of their perturbation for gene expression must be elucidated.During my recent postdoctoral work, I identified MRG-1, the first component of the active chromatin compartment shown to be a critical for shaping heterochromatin spatial distribution in the intestine of C. elegans. Nonetheless, fundamental open questions remain as to how this regulation takes place. In particular, MRG-1 bears a chromo barrel domain that can “read” methylated histones. Yet, the chromatin signals that determine MRG-1 recruitment and function are not known. Furthermore, chromatin readers do not act alone and typically serve as recruiters of complexes to target sites. However, the MRG-1-containing complex responsible for heterochromatin organization is completely unknown.In this proposal, we aim to identify i) the histone marks, and corresponding chromatin modifiers, that participate in MRG-1 recruitment and contribute to its role in heterochromatin architecture and ii) the complex/protein interactors through which MRG-1 functions in heterochromatin organization, in the C. elegans intestine.To achieve our goals, my team will employ a combination of microscopy, genetic and biochemical approaches. Moreover, state-of-the-art molecular biology assays such as Emerin-DamID and RNAseq will provide us with integrated landscapes of chromatin compartmentalization and gene expression in the intestine of relevant mutants. With this work, we will identify novel players that shape the 3D organization of the genome within a differentiated tissue of a whole organism. Additionally, we will be able to determine their relevance for gene expression, at the tissue-specific level.Because MRG-1 and most chromatin factors are highly conserved between worms and mammals, this work is likely to have a broad impact in the field of epigenetics and nuclear architecture.