Uncovering the Asymmetry and ATP-Dependent Phases of Nucleosome Editing

All genomic processes in eukaryotes occur in the context of chromatin – genomic DNA punctuated by DNA-wrapped histone octamers, called nucleosomes. Consequently, gene regulation is governed by the crosstalk between posttranslational histone modifications, histone variants, and ATP-dependent chromatin remodeling. The yeast chromatin remodeler SWR1C deposits the histone variant H2A.Z on nucleosomes near the transcription start site to regulate gene expression. However, much is currently unknown about how SWR1C initiates the deposition reaction and what intermediate steps are involved. In this body of work, I show that SWR1C deposits two copies of H2A.Z in an asymmetric manner on a promoter proximal-like nucleosome substrate, with a preference for H2A.Z deposition on the side of nucleosome facing away from the nucleosome-free region. Furthermore, using site-directed labeling and single-molecule techniques, I uncover three distinct phases of the deposition reaction by SWR1C: a priming step that uses ATP hydrolysis to unwrap the nucleosomal DNA, a rapid histone exchange step to deposit H2A.Z, and a release step that also uses ATP hydrolysis to detach the evicted H2A from the product nucleosome. Altogether, these findings highlight the complexity and intricate details of the nucleosome editing reaction by SWR1C