Mechanism of heterochromatin regulation by STAT

STAT对异染色质的调控机制 异染色质是DNA的紧密堆积形式，对于染色体压缩和转录非常重要。 沉默以及基因组稳定性、动物寿命和肿瘤抑制。异染色质 动力学，即，它的建立、维持和损失是如何控制的，仍然不完全清楚。我们 先前已经证明了uSTAT在异染色质维持中的生理作用，但是， 机制尚不清楚。该项目的总体目标是在分子水平上研究 STAT在建立和维持异染色质中的作用，并了解STAT磷酸化如何 充当将基因沉默转换为活跃转录的分子开关。我们之前已经展示了 免疫染色显示STAT的一部分在氨基酸700附近的临界酪氨酸处未磷酸化 uSTAT（称为uSTAT）定位于细胞核中，与异染色质蛋白1（HP 1）相关。我们有 从遗传学上表明，STAT对异染色质的维持是必不可少的，STAT的激活（通过 该酪氨酸上的磷酸化）与异染色质破坏有关。我们进一步表明， 人uSTAT 5A能够促进异染色质形成并抑制肿瘤生长。在新 初步研究使用染色质免疫沉淀，然后进行深度测序（ChIP-seq），我们有 发现大多数染色质结合的果蝇STAT位于异染色质中， STAT导致异染色质的整体减少，其由lys 9处的三甲基化组蛋白3标记 （H3K9me3）。我们进一步发现，当uSTAT被迫与常染色质结合时， 基因以HP 1依赖的方式。这些结果表明，uSTAT可能发挥重要作用，不仅在 维持而且启动异染色质形成。HP 1和H3 K9 me 3是 异染色质，其中HP 1是中心组分。已经表明，将HP 1募集到常染色质 足以形成异染色质。然而，HP 1不直接结合DNA，仅具有弱的 H3 K9 me 3的亲和力，因此可能需要DNA结合蛋白因子用于其初始募集， 异染色质基因座，或用于加强其与H3 K9 me 3的结合。我们假设uSTAT在以下方面发挥作用： 由于uSTAT二聚体的分子确认，异染色质的建立和维持。 我们将使用基因组学、遗传学和生物化学工具的组合来研究导致 异染色质的建立，STAT和HP 1在异染色质维持中的各自作用， uSTAT结合和HP 1募集中对顺式元件的要求，以及uSTAT和pSTAT的性质 DNA二聚体和HP 1结合。了解非经典STAT功能的分子机制， 异染色质调控不仅可以阐明异染色质的基本细胞生物学过程， 动力学以及与肿瘤抑制和表观遗传人类相关的异染色质基因沉默 疾病

Mechanism of heterochromatin regulation by STAT Heterochromatin is a tightly packed form of DNA important for chromosomal compaction and transcriptional silencing as well as for genome stability, animal longevity, and tumor suppression. How heterochromatin dynamics, i.e., its establishment, maintenance, and loss, is controlled remains incompletely understood. We have previously demonstrated a physiological role of uSTAT in heterochromatin maintenance, but the mechanism remains unclear. The overall goal of this project is to investigate, at the molecular level, the role of STAT in establishing and maintaining heterochromatin, and to understand how STAT phosphorylation can serve as a molecular switch converting gene silencing to active transcription. We have previously shown by immunostaining that a fraction of STAT not phosphorylated at the critical tyrosine around amino acid 700 (termed uSTAT) is localized in the nucleus in association with Heterochromatin Protein 1 (HP1). We have shown genetically that STAT is essential for heterochromatin maintenance, and that STAT activation (by phosphorylation on this tyrosine) is associated with heterochromatin disruption. We have further shown that human uSTAT5A is capable of promoting heterochromatin formation and suppressing tumor growth. In new preliminary studies using chromatin immunoprecipitation followed by deep sequencing (ChIP-seq), we have found that the majority of chromatin-bound Drosophila STAT is localized in heterochromatin, and that loss of STAT leads to a global decrease in heterochromatin, which is marked by trimethylated histone 3 at lys9 (H3K9me3). We have further found that, when forced to bind to euchromatin, uSTAT can repress nearby genes in an HP1-dependent manner. These results suggest that uSTAT may play an important role not only in maintaining but also in initiating heterochromatin formation. HP1 and H3K9me3 are hallmarks of heterochromatin, with HP1 being the central component. It has been shown that recruiting HP1 to euchromatin is sufficient for heterochromatin formation. However, HP1 does not bind DNA directly and has only weak affinity for H3K9me3, and thus may require DNA-binding protein factors for its initial recruitment to heterochromatic loci, or for strengthening its binding to H3K9me3. We hypothesize that uSTAT plays roles in both establishment and maintenance of heterochromatin due to the molecular confirmation of uSTAT dimers. We will use a combination of genomic, genetic, and biochemical tools to investigate the initial events leading to heterochromatin establishment, the respective roles of STAT and HP1 in heterochromatin maintenance, the requirement for cis-element in uSTAT binding and HP1 recruitment, and the properties of uSTAT and pSTAT dimers in DNA and HP1 binding. Understanding the molecular mechanism of noncanonical STAT function in heterochromatin regulation should shed light on not only the basic cell biological process of heterochromatin dynamics but also heterochromatic gene silencing relevant to tumor suppression and epigenetic human diseases.