The middle region of an HP1-binding protein, HP1-BP74, associates with linker DNA at the entry/exit site of nucleosomal DNA.
In higher eukaryotic cells, DNA molecules are present as chromatin fibers, complexes of DNA with various types of proteins; chromatin fibers are highly condensed in metaphase chromosomes during mitosis. Although the formation of the metaphase chromosome structure is essential for the equal segregation of replicated chromosomal DNA into the daughter cells, the mechanism involved in the organization of metaphase chromosomes is poorly understood. To identify proteins involved in the formation and/or maintenance of metaphase chromosomes, we examined proteins that dissociated from isolated human metaphase chromosomes by 0.4 m NaCl treatment; this treatment led to significant chromosome decondensation, but the structure retained the core histones. One of the proteins identified, HP1-BP74 (heterochromatin protein 1-binding protein 74), composed of 553 amino acid residues, was further characterized. HP1-BP74 middle region (BP74Md), composed of 178 amino acid residues (Lys(97)-Lys(274)), formed a chromatosome-like structure with reconstituted mononucleosomes and protected the linker DNA from micrococcal nuclease digestion by approximately 25 bp. The solution structure determined by NMR revealed that the globular domain (Met(153)-Thr(237)) located within BP74Md possesses a structure similar to that of the globular domain of linker histones, which underlies its nucleosome binding properties. Moreover, we confirmed that BP74Md and full-length HP1-BP74 directly binds to HP1 (heterochromatin protein 1) and identified the exact sites responsible for this interaction. Thus, we discovered that HP1-BP74 directly binds to HP1, and its middle region associates with linker DNA at the entry/exit site of nucleosomal DNA in vitro.
Hayashihara
Kayoko
K
Department of Biotechnology, Graduate School of Engineering, Osaka University, Suita, Osaka 565-0871, Japan.
Uchiyama
Susumu
S
Shimamoto
Shigeru
S
Kobayashi
Shouhei
S
Tomschik
Miroslav
M
Wakamatsu
Hidekazu
H
No
Daisuke
D
Sugahara
Hiroki
H
Hori
Naoto
N
Noda
Masanori
M
Ohkubo
Tadayasu
T
Zlatanova
Jordanka
J
Matsunaga
Sachihiro
S
Fukui
Kiichi
K
D001665:Binding Sites; D000090266:Chromobox Protein Homolog 5; D002868:Chromosomal Proteins, Non-Histone / Q000378:metabolism*; D002877:Chromosomes, Human; D004247:DNA / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000378:metabolism*; D006801:Humans; D008677:Metaphase; D009707:Nucleosomes / Q000378:metabolism*; D011485:Protein Binding; D011487:Protein Conformation
D002868:Chromosomal Proteins, Non-Histone; D004268:DNA-Binding Proteins; C548039:HP1BP3 protein, human; D009707:Nucleosomes; D000090266:Chromobox Protein Homolog 5; D004247:DNA