Dinucleosome DNA of human K562 cells: experimental and computational characterizations.
Dinucleosome formation is the first step in the organization of the higher order chromatin structure. With the ultimate aim of elucidating the dinucleosome structure, we constructed a library of human dinucleosome DNA. The library consists of PCR-amplifiable DNA fragments obtained by treatment of nuclei of erythroid K562 cells with micrococcal nuclease followed by extraction of DNA and adaptor ligation to the blunt-ended DNA fragments. The library was then cloned using a plasmid vector and the sequences of the clones were determined. The dominating clones containing the Alu elements were removed. A total of 1002 clones, which comprised a dinucleosome database, contained 84 and 918 clones from the clones before and after removing Alu elements, respectively. Approximately 70% of the clones were between 300 and 400 bp in size and they were distributed to various locations of all chromosomes except the Y chromosome. The clones containing A(2)N(8)A(2)N(8)A(2) or T(2)N(8)T(2)N(8)T(2) sequences were classified into three types, Type I (N shape), Type II (V shape) and Type III (M shape) according to DNA curvature plots. The locations of experimentally determined curved DNA segments matched well with the calculated ones though the clones of Types I and III showed additional curved DNA segments as revealed by the curvature plots. The distributions of complementary dinucleotides in the nucleosome DNA, at the ends of the dinucleosome DNA clones, allowed us to predict the positions of the nucleosome dyad axis, and estimate the size of the nucleosome core DNA, 125nt. The distributions of AA and TT dinucleotides, as well as other RR and YY dinucleotides, showed a periodicity with an average period of 10.4 bases, close to the values observed before. Mapping of nucleosome positions in the dinucleosome database based on the observed periodicity revealed that the nucleosomes were separated by a linker of 7.5+ approximately 10 x n nt. This indicates that the nucleosome-nucleosome orientations are, typically, halfway between parallel and antiparallel. Also an important finding is that the distributions of AA/TT and other RR/YY dinucleotides, apparently, reflect both DNA curvature and DNA bendability, cooperatively contributing to the nucleosome formation.
Kato
Megumi
M
Research Institute of Glycoscience, National Institute of Advanced Industrial Science and Technology, 1-1-1 Higashi, Tsukuba, Ibaraki 305-8566, Japan.
Onishi
Yoshiaki
Y
Wada-Kiyama
Yuko
Y
Abe
Takashi
T
Ikemura
Toshimichi
T
Kogan
Simon
S
Bolshoy
Alex
A
Trifonov
Edward N
EN
Kiyama
Ryoiti
R
D001483:Base Sequence; D002877:Chromosomes, Human; D004247:DNA / Q000235:genetics*; D015723:Gene Library; D006801:Humans; D020014:K562 Cells; D009707:Nucleosomes / Q000235:genetics*; D012091:Repetitive Sequences, Nucleic Acid
D009707:Nucleosomes; D004247:DNA