Screening The Molecular Chaperone Of Histone Variant MacroH2A

Eukaryotic genome is hierarchically organized into chromatin in the nucleus. Nucleosome, the basic unit of chromatin, consists of 147 base pairs(bp) of DNA wrapped around a histone octamer. As a important epigenetic regulator, histone variant can regulate the genome-associated biological processes.MacroH2A, which has a unique macro domain, is a variant of canonical histone H2A.The molecular weight of macroH2A is the threefold of other histones. The functions of macroH2A are usually related to gene silence. MacroH2A play an important role in X chromosome inactivation, embryonic development, tumorgenesis. Histone chaperones are defined mainly as factors that associate with histones and stimulate a reaction involving histone transfer without being part of the final product. The participation of histones chaperone in genesis, transfer, storage, metabolism is a prerequisite for histones. Formerly, the most studies about macroH2A were concentrated on functions, ignoring chaperones.In nuclear extracts (NE) the interaction between speculative histone chaperone Y and H2A-H2B dimmers was validated by trandem affinity purification (TAP) which is the classic technology for screening histone chaperones.We applied TAP technology to screen the molecular chaperone of histone variant macroH2A either. The NE of stable HEK293T cells that express macroH2A was contaminated by chromatins. We turn to HeLa S3 to execute the experiment, but it turn out the same. Probably the phenomenon is due to the macro domain. We used macroH2A deletion mutants as baits which can be deposited onto chromatins without macro domain. But the NE was also contaminated by chromatins. Several possible macroH2A chaperons were screened out from cytoplasmic matrix.They probably take a part in processing activity and entry into nucleus events of macroH2A. In summary, we screened out several macroH2A speculative chaperones from cytosolic extracts by valid TAP technology. We improved the TAP technology during the process of screening macroH2A chaperone from NE.