RNA-seq to determine PRC2 targets in the aging Drosophila brain.. RNA-seq to determine PRC2 targets in the aging Drosophila brain.

Aging is a prominent risk factor for neurodegenerative disease, therefore defining mechanisms critical for healthy brain aging should lead to insight into genes that modulate susceptibility to disease. To define such genes, we have pursued analysis of miR-34 mutants in Drosophila. The miR-34 mutant brain displays a gene profile of accelerated aging, and miR-34 upregulation is a potent suppressor of polyglutamine-induced neurodegeneration. We investigated targets of miR-34 to define those important for its functions in mitigating degeneration and impacting health of the brain with age. These studies show that miR-34 targets for silencing two components of polycomb repressive complex 2 (PRC2)—Pcl and Su(z)12—in the brain with age. PRC2 is a histone methyltransferase that confers the repressive H3K27me3 mark, suggesting that a critical role of miR-34 is to modulate the function of PRC2 to silence key genes in the brain with age. Remarkably, gene expression profiling of the brains of hypomorphic mutants in Enhancer of zeste (E(z)), the enzymatic methyltransferase component of PRC2, revealed a younger brain transcriptome profile and identified the small heat shock proteins as key modulated genes. These findings indicate that PRC2 epigenetic mechanisms impact the susceptibility of the brain to degenerative disease with age, and highlight the role of small heat shock proteins to protect the brain from age-associated decline and disease. Overall design: The design involves three factors: genotype, age, and cultivation temperature. Genotype can be either w[1118] (wild-type control) or w[1118]; E(z)[61] / E(z)[731] (E(z) hypomorphic mutant). Samples have an age of either 3d (3 days) or 20d (20 days). Animals are cultivated at either 25 degrees Celsius or 29 degrees Celsius. Samples are derived from Drosophila brain. We evaluate changes in gene expression with age. We also evaluate changes in gene expression in animals with hypomorphic mutations in E(z), with blocking on temperature.

衰老是神经退行性疾病的重要风险因素，因此阐明健康大脑衰老的关键机制，将有助于揭示调控疾病易感性的基因。为揭示此类基因，我们开展了果蝇（Drosophila）中miR-34（microRNA-34）突变体的相关分析。miR-34突变体的大脑呈现加速衰老的基因表达谱，且miR-34过表达是聚谷氨酰胺诱导的神经退行性病变的强效抑制剂。我们通过探究miR-34的靶基因，以明确其在缓解衰老相关大脑退化、维护大脑健康过程中发挥功能的关键靶点。本研究证实，miR-34在衰老大脑中靶向沉默多梳抑制复合体2（polycomb repressive complex 2, PRC2）的两个组分：Pcl与Su(z)12。PRC2是一种组蛋白甲基转移酶，可介导抑制性组蛋白修饰H3K27me3，这提示miR-34的关键功能之一是通过调控PRC2的活性，在衰老大脑中沉默关键基因。值得注意的是，对PRC2的酶活性甲基转移酶组分zeste增强子（Enhancer of zeste, E(z)）功能减退突变体的大脑进行基因表达谱分析，结果显示其大脑转录组谱更年轻化，并鉴定出小分子热激蛋白为关键调控靶点。上述研究结果表明，PRC2介导的表观遗传机制可影响大脑随衰老出现的退行性疾病易感性，并凸显了小分子热激蛋白在保护大脑抵御衰老相关功能衰退与疾病中的作用。实验整体设计：本实验包含三个变量因素：基因型、样本年龄与培养温度。基因型分为两类：w[1118]（野生型对照）以及w[1118]; E(z)[61]/E(z)[731]（E(z)功能减退突变体）。样本年龄分为3天（3d）与20天（20d）两组。果蝇培养温度设置为25℃与29℃两个梯度。实验样本均取自果蝇大脑。本研究旨在评估随衰老发生的基因表达变化，同时也将分析E(z)功能减退突变体在不同温度条件下的基因表达变化。