A novel SNF2 ATPase complex in Trypanosoma brucei with a role in H2A.Z-mediated chromatin remodelling

A cascade of histone acetylation events with subsequent incorporation of a histone H2A variant plays an essential part in transcription regulation in various model organisms. A key player in this cascade is the chromatin remodellling complex SWR1, which replaces the canonical histone H2A with its variant H2A.Z. Transcriptional regulation of polycistronic transcription units in the unicellular parasite Trypanosoma brucei has been shown to be highly dependent on acetylation of H2A.Z, which is mediated by the histone-acetyltransferase HAT2. The chromatin remodellling complex, which mediates H2A.Z incorporation is not known and an SWR1 orthologue in trypanosomes has not yet been reported. In this study, we identified and characterised an SWR1-like remodelller complex in T. brucei that is responsible for Pol II-dependent transcriptional regulation. Bioinformatic analysis of potential SNF2 DEAD/Box helicases, the key component of SWR1 complexes, identified a 1211 amino acids-long protein that exhibits key structural characteristics of the SWR1 subfamily. Systematic protein-protein interaction analysis revealed the existence of a novel complex exhibiting key features of an SWR1-like chromatin remodelller. RNAi-mediated depletion of the ATPase subunit of this complex resulted in a significant reduction of H2A.Z incorporation at transcription start sites and a subsequent decrease of steady-state mRNA levels. Furthermore, depletion of SWR1 and RNA-polymerase II (Pol II) caused massive chromatin condensation. The potential function of several proteins associated with the SWR1-like complex and with HAT2, the key factor of H2A.Z incorporation, is discussed.

在多种模式生物中，一系列级联式的组蛋白乙酰化（histone acetylation）事件伴随后续组蛋白H2A变体（histone H2A variant）的整合，在转录调控过程中发挥不可或缺的核心作用。该级联反应的关键参与者是染色质重塑复合物SWR1（SWR1），其可将经典组蛋白H2A替换为其变体H2A.Z（histone H2A.Z）。已有研究证实，单细胞寄生虫布氏锥虫（Trypanosoma brucei）的多顺反子转录单位（polycistronic transcription units）的转录调控高度依赖H2A.Z的乙酰化修饰，该修饰由组蛋白乙酰转移酶HAT2（histone-acetyltransferase HAT2）介导。目前，介导H2A.Z整合的染色质重塑复合物尚未被阐明，且锥虫中尚未见SWR1同源物的相关报道。本研究中，我们鉴定并表征了布氏锥虫内一种类SWR1重塑复合物，该复合物负责依赖RNA聚合酶II（Pol II）的转录调控。通过对作为SWR1复合物核心组分的潜在SNF2 DEAD/Box解旋酶（SNF2 DEAD/Box helicases）进行生物信息学分析，我们鉴定出一段长度为1211个氨基酸的蛋白质，其具备SWR1亚家族的关键结构特征。系统性蛋白质相互作用分析显示，存在一种具备类SWR1染色质重塑复合物核心特征的新型复合物。通过RNA干扰（RNAi）介导该复合物ATP酶亚基的耗竭，会显著降低转录起始位点（transcription start sites）处的H2A.Z整合水平，并随后导致稳态mRNA（steady-state mRNA）水平的显著下降。此外，耗竭该类SWR1复合物以及RNA聚合酶II（Pol II）会引发大规模染色质凝聚（chromatin condensation）。本文还讨论了与该类SWR1复合物以及H2A.Z整合关键因子HAT2相关的多种蛋白质的潜在功能。