Table Of ContentK. Nagata, K. Takeyasu (Eds.)
Nuclear Dynamics
Molecular Biology and Visualization of the Nucleus
K. Nagata, K. Takeyasu (Eds.)
Nuclear Dynamics
Molecular Biology and
Visualization of the Nucleus
With 48 Figures, Including 34 in Color
Springer
Kyosuke Nagata, Ph.D.
Professor
Department of Infection Biology
Graduate School of Comprehensive Human Sciences
University of Tsukuba
Tennohdai, Tsukuba, Ibaraki 305-8575, Japan
Kunio Takeyasu, Dr.Sci.
Professor
Laboratory of Plasma Membrane and Nuclear Signaling
Kyoto University Graduate School of Biostudies
Sakyo-ku, Kitashirakawa-Oiwake-cho, Kyoto 606-8502, Japan
Library of Congress Control Number: 2006938399
ISBN-10 4-431-30054-6 Springer Tokyo Berlin Heidelberg New York
ISBN-13 978-4-431-30054-0 Springer Tokyo Berlin Heidelberg New York
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Preface: Overview of Nuclear Organization and
Nuclear Dynamics
Kunio Takeyasu
Laboratory of Plasma Membrane and Nuclear Signaling, Kyoto University
Graduate School of Biostudies, Kitashirakawa Oiwake-cho, Sakyo-ku,
Kyoto 606-8502, Japan
In contrast to the fact that prokaryotes, which do not have cell nuclei, can
live only as unicellular organisms, eukaryotes bearing cell nuclei exist as a
huge variety of organisms from single cell to higher multicellular organ
isms having higher-level functions. The reason for this is that cell nuclei of
eukaryotes have a combination of mechanisms to store, retain, and transfer
the very long genome stably and to produce cells carrying different func
tions from the identical genome information. Such functions of cell nuclei
are supported by the intranuclear higher-order architecture. To understand
vital activities closely related to genome fiinction, such as cell prolifera
tion, environment responsiveness, reproduction, development, differentia
tion, and aging, it is necessary to know the fundamental structure and con
struction of the cell nucleus, "a container" of genome (shown above). In
this book, we investigate the dynamics of the nuclear structure as the basis
supporting genome function, thereby comprehensively understanding how
the higher-order structure of the nucleus is established and how it corre
lates with the expression of a variety of vital activities, including cell pro
liferation and ontogenesis, by combining biochemical and molecular ge
netic methods with the latest imaging techniques.
Nuclear Organization
A cell nucleus (hereafter referred to as nucleus) is an organelle that is
compartmentalized by a special membrane structure, the nuclear envelope.
A nucleus has a number of structural characteristics that other organelles
do not have. One of these is internal storage of genomic DNA, a chemi
cally single and physically very long supermolecule. In the case of human
cells, DNA approximately 2 m long is stored in a tiny nucleus with a dia
meter of ~5 // m. Accurate replication and distribution of such a long
VI K. Takeyasu
DNA molecule is required for the cell cycle regulation, and selective tran
scription of appropriate genes is necessary for transcriptional regulation.
These facts conjure up an image of a highly orderly structure being re
quired in a nucleus to keep and to express genome functions. The second
characteristic of a nucleus is that it provides "space" for molecules such as
proteins and RNAs to dissociate and distribute easily, since it is a giant
organelle and lacks an internal membrane structure as a partition. In fact,
many functional complexes (several kinds of nuclear bodies, such as nu
clear speckle, Cajal body, and promyelocytic leukemia oncoprotein [PML]
body, as well as nucleolus) exist in the nucleus (shown above), and its
structure and function show dynamic alteration depending on the physi
ological conditions. Such functional complexes are designated as intranu
clear compartments and/or intranuclear domains according to their func
tions and sizes. The third characteristic is a nuclear envelope structure,
which exists as a boundary between the nucleus and the cytoplasm. Nu-
clearpore complexes exist to link two lipid bilayers that constitute a nu
clear envelope, and regulate the bidirectional transport between the nucle
us and the cytoplasm. A mesh-like structure, known as the nuclear lamina,
in the inner layer of the nuclear envelope provide the scaffolding for
chromosome functions in animal cells. Thus, the nuclear envelope, includ
ing nuclear pores, supports genome function of a nucleus structurally and
in terms of substance transport.
Preface: Overview of Nuclear Organization and Nuclear Dynamics VII
Fundamental Problems to be Solved
Intensive investigations of individual events in the nucleus, such as tran
scription of genes, DNA replication, repair, recombination, and RNA
processing, have been conducted to date. Biological factors involved in the
raw process of individual vital functions have been discovered by bio
chemical, molecular, biological, and genetic methods, and their molecular
bases and molecular mechanisms have been analyzed. Nevertheless, how
individual reactions are regulated temporally and spatially has not been
investigated so intensively in spite of its importance, since most of the
findings were based on in vitro reactions.
The amount of intranuclear proteins is huge (several hundred
mg proteins/ml, assuming that they are distributed evenly in the nucleus).
These proteins function without forming precipitations in the living cells
by forming highly orderly complexes with chromosomal DNAs and
RNAs. However, they have not been well-investigated biochemically,
since most nuclear proteins form highly insoluble precipitations as a result
of biochemical treatment, including DNA degradation. Although such in
soluble proteins, called nuclear matrix proteins, may potentially affect
gene expression as a nuclear architecture, the function of these has not
been clarified thus far due to difficulties in the biochemical analysis.
Trend of New Nucleus Research
Genomic sequences of a variety of living organisms have been read, and
genes coding for proteins, components of organisms, are being unraveled
comprehensively. The presence and function of novel proteins can be
speculated on and searched for using such genome information. In addi
tion, thanks to the development of proteomics technology, the protein pro
file of a number of organelles in a nucleus, including the nucleolus, is now
being clarified inclusively. Under the circumstances, single molecular dy
namics in living cells and visualization of intermolecular binding are now
enabled by great advances in the development of several fluorescent pro
teins and imaging techniques as independent technologies. By expressing
target proteins fused to fluorescent proteins in cells, membrane proteins as
well as nuclear proteins, which have thus far been difficult to handle bio
chemically, can be dynamically investigated in living cells. Based on the
genome information accumulated as mentioned above, it becomes possible
to map the dynamics of individual proteins in cells in a three-dimensional
VIII K. Takeyasu
structure, over time, by a combination of biochemical and molecular ge
netic methods with molecular imaging techniques.
Based on the dynamic viewpoint of time, unforeseen and unex
pected findings are being revealed, including the fact that importin j3,
which transports nuclear proteins into the nucleus in the intermittent phase,
is also involved in spindle formation, that components of the nuclear pore
complex are present in the centromere in the mitotic phase and are in
volved in chromosome disjunction, and that RNAs produced as transcrip
tional products transform DNA in their transcriptional regions into inactive
heterochromatin. In other words, spatiotemporal changes of cellular
molecules and structures in vital activity can now be captured.
In addition to the progress in the technologies investigating
molecular dynamics, techniques derived to analyze the genome are also
advancing markedly, and it has now been found that abnormalities of nu
clear proteins induce several diseases, thanks to identification of genes re
sponsible for these diseases. These include "nuclear membrane diseases,"
which are triggered by abnormalities in nuclear envelope proteins. For in
stance, a number of diseases, such as Emery-Dreifuss muscular dystrophy,
familial lipodystrophy, progeria syndrome, and Pelger-Huet malformation,
occur as a result of deletions and/or mutations of proteins of the inner layer
of the nuclear envelope as well as nuclear lamina. The fact that nuclear
envelope proteins are responsible for several diseases showing different
symptoms suggests that the nuclear envelope architecture can regulate the
expression of genome information dynamically, depending on the devel
opmental and environmental status.
Hope for Fruitful Research on Nuclear Dynamics
Many authors of chapters in this book participated in the International
Workshop on Nuclear Dynamics: Approaches from Biochemistry,
Molecular Biology and Visual Biology at Yokohama, Japan on December
5, 2002. All of us agreed that, in order to understand vital activity closely
related to genome function, it is prerequisite to understand the inner struc
ture of the cell nucleus that supports the genome function. Since the struc
ture is not always stable but varies from hour to hour, it is necessary to
capture dynamic changes of the structure and function from the viewpoint
of molecular dynamics. Since dynamic structural changes might be related
to unexpected functions, researchers from a broad range of specialized ar
eas including nuclear envelope, nuclear transport, chromosome structure,
transcription, RNA processing, and nuclear domain structures should pro-
Preface: Overview of Nuclear Organization and Nuclear Dynamics IX
mote studies by collaborating organically with researchers specializing in
imaging and nanotechnology. From this point of view, this book was writ
ten. It took over 3 years to complete, with the tremendous efforts of indi
vidual authors.
We hope that the outcomes of this field are prerequisite for the
understanding of gene expression, DNA replication, and chromatin struc
ture, and are likely to open new avenues for the elucidation of genes re
sponsible for "nuclear membrane diseases" and abnormalities in chromo
somal distribution, as well as mechanisms involved in nuclear
reprogramming. Study results may also support research activities aiming
at applications in regenerative medicine, which include the development of
artificial chromosomes used in gene therapy as well as artificial cells that
have specialized nuclear functions. In this sense, nuclear dynamics re
search could open up an extremely important field in terms of understand
ing the basics of life.
Contents
Preface: Overview of Nuclear Organization and Nuclear Dynamics
Kunio Takeyasu V
1. Visual Biology of Nuclear Dynamics: From Micro- to
Nano-dynamics of Nuclear Components
Shige H. Yoshimura 1
2. Nuclear Envelope: Nanoarray Responsive to Aldosterone
Hans Oberleithner 38
3. Mitotic Chromosome Segregation Control
Yu Xue, Chuanhai Fu, Yong Miao, Jianhui Yao, Zhen Dou,
Jie Zhang, Larry Brako, and Xuebiao Yao 55
4. Breakdown and Reformation of the Nuclear Envelope
Tokuko Haraguchi and Yasushi Hiraoka 89
5. Functional Organization and Dynamic Aspects of Nucleoli
During the Cell Cycle
Takuya Saiwakiand Yoshihiro Yoneda 107
6. Dynamics, Roles, and Diseases of the Nuclear Membrane,
Lamins, and Lamin-binding Proteins
Tsuneyoshi Horigome, Yasuhiro Hirano, and
Kazuhiro Furukawa 123
7. Gene Selectors Consisting of DNA-Binding Proteins, Histones,
and Histone-Binding Proteins Regulate the Three Major Stages
of Gene Expression
Shinsuke Muto and Masami Horikoshi 145
8. Dynamic Chromatin Loops and the Regulation of Gene
Expression
Hiroshi Kimura and Peter R. Cook 177
XII Contents
9. Nuclear Architecture: Topology and Function of Chromatin-
and Non-Chromatin Nuclear Domains
Satoshi Tashiro, Marion Cremer, Irina Solovei,
and Thomas Cremer 197
10-1. Regulation of Chromatin Structure by Curved DNA: How
Activator Binding Sites Become Accessible
Takashi Ohyama 227
10-2. Actin-Related Proteins Involved in Nuclear and Chromatin
Dynamics
Masahiko Harata 239
10-3. Effects of 5-Bromodeoxyuridine on Chromatin Structure
Dai Ayusawa 249
10-4. Transcriptional Modulation by Nuclear Matrix Protein
P130/MAT3 Associated with MAR/SAR
Yasuhide Hibino, Tatsuhiro Usui, and Koichi Hiraga 255
10-5. Breaking and Tessellating the Contiguous Nuclear Genome
Kojiro Ishii 263
Perspective—toward understanding the in situ genome function
Kyosuke Nagata 271
Index 277
