The histone chaperone ANP32B regulates chromatin incorporation of the atypical human histone variant macroH2A.

All vertebrate genomes encode for three large histone H2A variants that have an additional metabolite-binding globular macrodomain module, macroH2A. MacroH2A variants impact heterochromatin organization and transcription regulation and establish a barrier for cellular reprogramming. However, the mechanisms of how macroH2A is incorporated into chromatin and the identity of any chaperones required for histone deposition remain elusive. Here, we develop a split-GFP-based assay for chromatin incorporation and use it to conduct a genome-wide mutagenesis screen in haploid human cells to identify proteins that regulate macroH2A dynamics. We show that the histone chaperone ANP32B is a regulator of macroH2A deposition. ANP32B associates with macroH2A in cells and in vitro binds to histones with low nanomolar affinity. In vitro nucleosome assembly assays show that ANP32B stimulates deposition of macroH2A-H2B and not of H2A-H2B onto tetrasomes. In cells, depletion of ANP32B strongly affects global macroH2A chromatin incorporation, revealing ANP32B as a macroH2A histone chaperone. [Display omitted] • Split-GFP can be used as a high-throughput readout for chromatin incorporation • Genetic screen identified macroH2A regulators • ANP32B binds macroH2A-H2B dimers and deposits them onto tetrasomes in vitro • Loss of ANP32B affects macroH2A chromatin localization on a genome wide scale Mandemaker et al. establish a split-GFP based assay to measure chromatin incorporation and use it to perform a genetic screen to identify regulators of the histone variant macroH2A. They show that ANP32B can directly bind to macroH2A and affect its deposition both in vitro and in cells. [ABSTRACT FROM AUTHOR]