Growth regulated co-occupancy of Mediator and Lsm3 at intronic ribosomal protein genes [ChIP-seq]

Mediator is a well-known transcriptional co-regulator and serves as an adaptor between gene-specific regulatory proteins and RNA polymerase II. Studies on the chromatin-bound form of Mediator revealed interactions with additional protein complexes involved in various transcription-related processes, such as the Lsm2-8 complex which is part of the spliceosomal U6 snRNP complex. Here we employ ChIP-seq of chromatin associated with the Lsm3 protein and the Med1 or Med15 Mediator subunits. We identify 86 genes co-occupied by both Lsm3 and Mediator of which 73 were intron-containing ribosomal protein genes. In logarithmically growing cells, Mediator primarily binds to their promoter regions but also shows a second, less pronounced occupancy at their 3´-exons. During late exponential phase, we observe a near complete transition of Mediator from these promoters to a position in their 3´-ends, overlapping the Lsm3 binding sites approximately 250 bp downstream of their last intron-exon boundaries. Using an unbiased RNA-seq approach we show that transition of Mediator from promoters to the last exon of these genes correlates to reduction of both their mRNA levels and splicing ratios, indicating that the Mediator and Lsm complexes co-operate to control growth-regulated expression of intron-containing ribosomal protein genes at the levels of transcription initiation and splicing.中介体（Mediator）是一类经典的转录共调控因子，可作为基因特异性调控蛋白与RNA聚合酶II（RNA polymerase II）之间的衔接复合物。针对染色质结合型中介体的研究显示，其可与多种转录相关过程中的其他蛋白质复合物发生相互作用，例如属于剪接体U6 snRNP复合物组分的Lsm2-8复合物。本研究针对与Lsm3蛋白以及Med1或Med15中介体亚基结合的染色质开展染色质免疫共沉淀测序（ChIP-seq）实验，共鉴定得到86个同时被Lsm3与中介体共同占据的基因，其中73个为含内含子的核糖体蛋白基因。在对数生长期细胞中，中介体主要结合于这些基因的启动子区域，同时在其3'外显子区域也存在一处较弱的结合峰。在指数生长后期，我们观察到中介体几乎完全从这些启动子区域转移至基因3'末端区域，该区域与Lsm3的结合位点重合，且位于最后一个内含子-外显子边界下游约250 bp处。借助无偏倚RNA测序（RNA-seq）方法，我们证实中介体从启动子转移至这些基因最后一个外显子的过程，与其mRNA水平及剪接比率的下降显著相关，这表明中介体与Lsm复合物可协同在转录起始和剪接层面，调控含内含子核糖体蛋白基因的生长依赖性表达。