Proteomics analysis of the bromodomain protein IBD2 reveals mechanistic insights into H2A.Z (Hv1) dynamics in Tetrahymena

INO80 and SWR1 are evolutionarily related ATP-dependent chromatin remodeling complexes that regulate the chromatin occupancy of the histone variant H2A.Z, playing critical roles in transcriptional regulation, genome replication, and DNA repair. While INO80 and SWR1's H2A.Z-related functions are well characterized in budding yeast and metazoans, much less is known about their composition and conserved roles outside the Opisthokonts. Here, we report that in the ciliate protozoan Tetrahymena thermophila, a distinct bromodomain-containing protein, IBD2, functions as a component of a putative INO80 complex. Chromatin immunoprecipitation followed by high-throughput sequencing (ChIP-seq) demonstrates that IBD2 is enriched near transcription start sites and promoter regions. Notably, the IBD2 genomic binding profile strongly correlates with that of H2A.Z (Hv1), supporting its association with the INO80 complex. Overall design: Examination of ChIP DNA for Ibd2-FZZ cells (biological duplicates) during vegetative growth. Examination of RNA of Wild type cells (biological duplicates) and Ibd1 KO cells (biological triplicates) during vegetative growth

INO80与SWR1属于进化上相关的ATP依赖型染色质重塑复合物（ATP-dependent chromatin remodeling complexes），可调控组蛋白变体H2A.Z的染色质占据状态，在转录调控、基因组复制与DNA修复过程中发挥关键作用。尽管INO80与SWR1的H2A.Z相关功能已在出芽酵母与后生动物（metazoans）中得到充分表征，但在后鞭毛生物（Opisthokonts）类群之外，人们对其复合物组成及保守功能的认知仍十分有限。本研究报道，在纤毛原生动物嗜热四膜虫（Tetrahymena thermophila）中，一种独特的含溴结构域蛋白IBD2可作为推定INO80复合物的组分之一发挥功能。通过染色质免疫沉淀联合高通量测序（ChIP-seq，chromatin immunoprecipitation followed by high-throughput sequencing）分析，证实IBD2在转录起始位点与启动子区域附近呈现富集。值得注意的是，IBD2的基因组结合谱与H2A.Z（Hv1）的结合谱高度相关，这进一步支持其与INO80复合物的关联。总体实验设计：对营养生长阶段的Ibd2-FZZ细胞进行染色质免疫沉淀DNA（ChIP DNA）检测（生物学重复2次）；对营养生长阶段的野生型细胞（生物学重复2次）与Ibd1基因敲除（KO）细胞（生物学重复3次）进行RNA检测。