Functional role of genes and pathways in histone H3 mutated gliomas

组蛋白在进化上是高度保守的，它们的表观遗传修饰在真核细胞的分化和大脑发育中起着关键作用。在脑肿瘤中，组蛋白H3突变（K27M， G34R和G34V）被描述为恶性胶质瘤亚组的驱动突变，表现出侵略性的生物学行为和对治疗的不良反应。组蛋白H3突变已被证明可引起转录失调，但对功能相关基因和信号通路知之甚少。在拟议的DFG项目中，我们的目标是确定参与组蛋白h3突变胶质瘤生物学的基因和途径，这些基因和途径具有功能和临床相关性。特别是，我们希望更好地了解(1)组蛋白H3突变的过表达如何影响中枢神经系统发育，(2)研究哪些基因和信号通路参与了组蛋白H3突变的有害影响，(3)检查这些基因和信号通路在组蛋白H3突变肿瘤中的功能作用和临床相关性。为此，我们建立了果蝇模型，研究人类突变组蛋白H3.3的普遍存在和细胞类型特异性过表达的影响。在这个模型中，我们将采用双重方法。在一种无偏倚的方法中，我们将(1)通过在1099种不同的苍蝇株中杂交，表达在苍蝇神经系统中表达的基因siRNA结构，进行Gal4-UAS修饰剂筛选，并研究普遍存在的和细胞类型特异性敲低对过度表达突变组蛋白H3.3引起的表型的影响。同时，我们将(2)在该模型中研究K27M突变胶质瘤中发现的过表达基因的功能作用，以及人类基因过表达的果蝇同源基因的功能作用。结果将进一步验证通过研究药理抑制所确定的途径的影响。将分别在人类组蛋白h3突变的肿瘤细胞系和临床注释良好的组蛋白h3突变的肿瘤标本中检测已鉴定基因和信号通路的表达、功能作用（组蛋白三甲基化、增殖、迁移）以及临床相关性。该项目的结果将有助于更好地了解组蛋白h3突变胶质瘤的生物学基因和途径。从长远来看，它们有望帮助更好地治疗组蛋白h3突变胶质瘤患者。

Histones are evolutionarily highly conserved and their epigenetic modification plays a key role in the differentiation of eukaryotic cells and brain development. In brain tumors, histone H3 mutations (K27M, G34R and G34V) have been described as driver mutations in a subgroup of malignant gliomas showing aggressive biological behavior and poor response to therapy. Histone H3 mutations have been shown to cause transcriptional dysregulation, but little is known on functionally relevant genes and signaling pathways. Within the proposed DFG project, we aim at identifying genes and pathways involved in the biology of histone H3-mutated gliomas, which are of functional and clinical relevance. In particular, we want to better understand (1) how overexpression of mutated histone H3 affects central nervous system development, (2) investigate which genes and signaling pathways are involved in the detrimental effects of histone H3 mutations and (3) examine the functional role and clinical relevance of these genes and signaling pathways in histone H3 mutated tumors. To this end, we have established a fly model to study the effect of ubiquitous and cell-type specific over-expression of human mutated histone H3.3. In this model, we will employ a dual approach. In an unbiased approach we will (1) perform Gal4-UAS modifier screens by crossing in 1099 different fly strains expressing siRNA constructs of genes expressed in the fly nervous system and study the effect of ubiquitous and cell-type specific knockdown on the phenotype caused by overexpression of mutated histone H3.3. In parallel, we will (2) study the functional role of genes found to be over-expressed functional role of fly orthologues of human genes over-expressed in K27M mutated gliomas in this model. Results will be further validated by studying the effect of pharmacological inhibition of identified pathways. Expression, functional role (histone trimethylation, proliferation, migration) and clinical relevance of identified genes and signaling pathways will be examined in human histone H3-mutated tumor cell lines as well as clinically well-annotated histone H3-mutated tumor specimens, respectively. The results of this project will contribute to a better understanding of genes and pathways involved in the biology of histone H3-mutated gliomas. In the long term, they are expected to aid better treatments of patients harboring histone H3-mutated gliomas.