Catalytic-dependent and independent functions of the histone acetyltransferase CBP promote pioneer factor-mediated zygotic genome activation [RNA-seq]. Catalytic-dependent and independent functions of the histone acetyltransferase CBP promote pioneer factor-mediated zygotic genome activation [RNA-seq]

Immediately after fertilization the genome is transcriptionally quiescent. Maternally encoded pioneer transcription factors reprogram the chromatin state and facilitate the transcription of the zygotic genome. In Drosophila, transcription is initiated by the pioneer factor Zelda. While Zelda-occupied sites are enriched with histone acetylation, a post-translational mark associated with active cis-regulatory regions, the functional relationship between Zelda and histone acetylation in zygotic genome activation remained unclear. We show that Zelda-mediated recruitment of the histone acetyltransferase CBP is essential for zygotic transcription and embryonic development. CBP catalytic activity is necessary for release of RNA Polymerase II (Pol II) into transcription elongation. However, CBP largely activates zygotic transcription independent of acetylation through Pol II recruitment. Neither acetylation nor CBP are required for the pioneering function of Zelda. Our data suggest that pioneer factor-mediated recruitment of CBP is a conserved mechanism required to activate zygotic transcription but that this role is separable from the function of pioneer factors in restructuring chromatin accessibility. Overall design: This data set contains single-embryo RNA-seq on embryos precisely staged 15 minutes into nuclear cycle 14 during the maternal-to-zygotic transition. GFP-CBP;his2AV-RFP and GFP-CBP;nos-deGrad/+;his2AV-RFP embryos were used to quantify the effect CBP knockdown has on transcription during zygotic genome activation. To assay the catalytic function of CBP we used an optogenetic line, CRY2-CBP, to inactivate CBP mediated histone acetyltransferase with blue light treatments. CRY2-CBP;his2av-RFP were treated with blue light to inactivate CBP catalytic activity via the CRY2 optogenetic tag or left in the dark as a control to assay the function of CBP mediated acetylation during ZGA. His2AV-RFP control embryos were treated similarly and used to compare to CRY2-CBP;His2avRFP embryos.

受精完成后即刻，基因组即处于转录静默状态。母源编码的先锋转录因子（pioneer transcription factor）可重编程染色质状态，并促进合子基因组（zygotic genome）的转录。在果蝇（Drosophila）中，转录起始由先锋转录因子泽尔达（Zelda）介导。尽管泽尔达结合位点富集组蛋白乙酰化修饰——这是与活跃顺式调控区域相关的翻译后修饰标记——但泽尔达与组蛋白乙酰化在合子基因组激活过程中的功能关联仍不明确。
本研究证实，泽尔达介导的组蛋白乙酰转移酶CBP（histone acetyltransferase CBP）招募过程，对于合子转录与胚胎发育至关重要。CBP的催化活性对于RNA聚合酶II（RNA Polymerase II，Pol II）释放进入转录延伸阶段不可或缺。然而，CBP主要通过招募RNA聚合酶II激活合子转录，这一过程并不依赖于乙酰化修饰。泽尔达的先锋功能既不依赖乙酰化修饰，也无需CBP的参与。本研究数据表明，先锋转录因子介导的CBP招募是激活合子转录的保守机制，但该功能与先锋转录因子重塑染色质可及性的功能相互独立。
实验设计：本数据集包含母源-合子转换时期核周期14阶段精准处于15分钟时相的单个胚胎RNA测序（RNA-seq）样本。我们采用GFP-CBP;his2AV-RFP与GFP-CBP;nos-deGrad/+;his2AV-RFP胚胎，量化CBP敲低对合子基因组激活过程中转录的影响。为检测CBP的催化功能，我们使用光遗传学品系CRY2-CBP，通过蓝光照射灭活CBP介导的组蛋白乙酰转移酶活性。CRY2-CBP;his2av-RFP胚胎经蓝光照射，通过CRY2光遗传学标签灭活CBP的催化活性；同时设置黑暗培养的对照组，以检测CBP介导的乙酰化在合子基因组激活（ZGA）过程中的功能。同时设置经相同处理的His2AV-RFP对照胚胎，用于与CRY2-CBP;His2avRFP胚胎进行比对分析。