Global effect of H2A.Z hypervariant knockdown on basal expression of neuronal genes

Variant histone H2A.Z is an essential and conserved regulator of eukaryotic gene transcription. However, the exact role of this histone in the process remains perplexing. In vertebrates, H2A.Z has two hypervariants – H2A.Z.1 and H2A.Z.2 – that have almost identical amino-acid sequences excepting three residues. Due to such similarity, functional specificity of these hypervariants in neurobiological processes, if any, remain largely unknown. In this study with dissociated mature cortical neurons, we asked if H2A.Z hypervariants have functional specificity in regulating basal and activity-induced gene transcription. Hypervariant-specific RNAi and microarray analyses revealed that H2A.Z.1 and H2A.Z.2 regulate basal expression of largely non-overlapping gene-sets, including genes that code for several synaptic proteins. In response to neuronal activity, depletion of H2A.Z hypervariants impairs transcription of our model gene Arc differentially. As part of the +1 nucleosome, H2A.Z.2, not H2A.Z.1, is necessary for rapid immediate-early transcription by facilitating RNA Pol II promoter proximal pausing. In contrast, H2A.Z.1 plays complicated and perhaps indirect roles that affect Arc transcription kinetics context-dependently. Context-dependent roles of H2A.Z hypervariants are also evident amongst several other immediate early genes where their regulatory roles vary from gene to gene under different conditions. Together, our data suggest that H2A.Z hypervariants have context-specific roles that complement each other to mediate activity-induced neuronal gene transcription.变异型组蛋白H2A.Z（histone H2A.Z）是真核基因转录过程中不可或缺且高度保守的调控因子。然而，该组蛋白在该过程中的具体作用仍有待厘清。 在脊椎动物体内，H2A.Z存在两种超变体——H2A.Z.1与H2A.Z.2——二者氨基酸序列几乎完全一致，仅存在三处残基差异。由于序列相似性极高，这两种超变体在神经生物学过程中的功能特异性（若存在的话）在很大程度上仍未被揭示。 本研究以解离培养的成熟皮层神经元为实验模型，探究H2A.Z超变体在调控基础转录与活性诱导型基因转录过程中是否具备功能特异性。 针对超变体的特异性RNA干扰（RNA interference, RNAi）与基因芯片（microarray）分析结果显示，H2A.Z.1与H2A.Z.2分别调控两大几乎无重叠的基因集的基础表达，其中包含若干编码突触蛋白的基因。 在神经元活性刺激条件下，敲低两种H2A.Z超变体的表达将差异化地损伤模式基因Arc的转录进程。 作为+1核小体（+1 nucleosome）的组成组分，H2A.Z.2而非H2A.Z.1，可通过促进RNA聚合酶II（RNA polymerase II, RNA Pol II）在启动子近端的暂停，介导快速即刻早期基因（immediate early genes）的转录。 与之相对，H2A.Z.1则通过复杂且可能间接的途径，以情境依赖的方式影响Arc的转录动力学特征。 在其他数种即刻早期基因中，H2A.Z超变体的调控作用同样呈现情境依赖性，其功能随基因种类与实验条件的不同而存在差异。 综上，本研究数据表明，H2A.Z超变体具备情境特异性的功能角色，二者相互协同，共同介导活性诱导的神经元基因转录过程。