Atomic Resolution Structure of Human Centromeric Nucleosomes Containing CENP-ATue, Jul 19, 2011
The Kurumizaka laboratory (Waseda University, Graduate School of Advanced Science and Engineering) has determined the crystal structure of the human centromeric nucleosome containing CENP-A (CENP-A nucleosome). The CENP-A nucleosome structure elucidated by the laboratory yielded physical evidence for an octameric CENP-A nucleosome and a shorter length of wrapped DNA, which had previously not been demonstrated explicitly. Therefore, the structure has provided important insights for understanding the formation and maintenance mechanisms of the centromeric chromatin architecture, containing the CENP-A nucleosome.
In eukaryotes, chromosomes are accurately segregated during mitosis and meiosis. Centromeres are the specific chromosomal regions for the formation of kinetochores, to which microtubules are attached. Centromeres are epigenetically marked by the assembly of nucleosomes containing CENP-A, which is the centromere-specific histone H3 variant. However, the underlying mechanisms of centromeric chromatin formation and maintenance have not been elucidated, although centromere inheritance is considered to be dictated by the CENP-A nucleosome.
To determine the architecture of the CENP-A nucleosome, we purified the bacterially expressed human histones H2A, H2B, H4, and CENP-A, and assembled the CENP-A nucleosome with a 147 base pair palindromic DNA, derived from a human α-satellite sequence. The CENP-A nucleosome was crystallized, and the structure was determined at 3.6 Å resolution. In the CENP-A crystal structure, the histone octamer, containing two each of histones H2A, H2B, H4, and CENP-A, wrapped the DNA in a left-handed orientation, in the same manner as nucleosomes containing other histone H3 variants. Biochemical experiments confirmed that the DNA was left-handedly wrapped within the CENP-A nucleosome.
Two major CENP-A nucleosome models, the octasome and hemisome models, have been proposed. The current crystal structure of the CENP-A nucleosome is consistent with the octasome model, in which two each of histones H2A, H2B, H4, and CENP-A form a histone octamer, with the DNA left-handedly wrapped around it. The hemisome model, in which one each of histones H2A, H2B, H4, and CENP-A form a heterotypic tetramer, with the DNA right-handedly wrapped around it, is unlikely.
Our structural and biochemical analyses revealed that the DNA regions at the entrance and exit of the CENP-A nucleosome are flexible. These observations are perfectly consistent with the previous reports. The specific DNA conformations at the entrance and exit of the CENP-A nucleosome may provide the binding sites for centromere-specific DNA-binding proteins, and may function to form the centromere-specific chromatin architecture.