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NUCLEUS
Animal cells contain DNA in nucleus (contains ~ 98% of cell DNA) and mitochondrion. Both
compartments are surrounded by an envelope (double membrane). Nuclear DNA represents some
linear molecules and mitochondrial DNA represents circular molecules.
All eukaryotic cells contain at least one
nucleus. The contents of the nucleus are present
as a viscous, amorphous mass of material
enclosed by a complex nuclear envelope.
Nucleus consists of:
Chromosomes, which are present as
extended nucleoprotein fibers, called chromatin;
Nuclear matrix, which is a proteincontaining fibrillar network;
Nucleolus (or some nucleoli) that are
responsible for synthesis of rRNA and
assembling of ribosomes;
Nucleoplasm (or karyoplasm) – the fluid
substance in which the solutes of nucleus are
Fig. 1. Position of nucleus in the cell
dissolved.
CHROMATIN
The eukaryotic cell nucleus is typically ~1mm in diameter. It contains a large amount of DNA
(a total length of 1-2 meters), which must be efficiently packaged in such a way as to guarantee
access to genetic information. Thus, each DNA molecule is packed forming chromatin, a densely
staining material initially recognized in two different forms: highly condensed heterochromatin and
more diffuse euchromatin. Chemically chromatin is organized from 30% DNA + 40% histones +
25% non-histones + 5% RNA.
Chromatin fragments contain DNA (a negatively charged polymer) in complex with highly
positively charged (basic) proteins called histones, and much smaller amounts of other DNAbinding proteins, collectively referred to as
non-histone proteins. Human histone genes
are represented as a family of moderately
repeated sequences with variations in the
structure, organization, and regulation of the
different copies. The histones organize DNA
into a regular repeating structure, the basic
unit of which is the nucleosome, which wrap
and compact DNA into chromatin, limiting
DNA accessibility to the cellular machineries
which require DNA as a template. Histones
thereby play a central role in transcription
regulation, DNA repair, DNA replication and
chromosomal stability. There are 5 classes of
histones: H1, H2A, H2B, H3, H4. They
contain Leucine, Lysine, Arginine and other
basic amino acids. The quantity of histones is
the same in all tissues and they have no tissuespecificity. They induce the tertiary structure
Fig. 2. Various stages in the condensation of chromatin
of DNA (DNP). The non-histones are acid
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proteins, are very heterogeneous and comprise: enzymes (for replication, repair, transcription, RNA
processing, biogenesis of ribosomes), site-specific proteins, scaffold proteins. More active tissues
contain larger quantity and diversity of non-histones.
Euchromatin represents the active part of
chromatin; contain structural genes that are transcribed.
It is less compacted and replicates early in the S period
of interphase.
Heterochromatin is tightly condensed and
contains inactive sequences which are not transcribed.
It replicates late in the S period. There are two types of
heterochromatin:
constitutive
and
facultative.
Facultative heterochromatin contains genes that in
some condition, in some tissues may be active, so it
can be transformed into euchromatin. It contains
coding
but
inactive
sequences.
Facultative
heterochromatin assures cell differentiation, sexual
differentiation,
and
control
of
ontogenesis.
Constitutive heterochromatin contains repetitive
sequences, which always remain condensed. It is
represented by centromeres, telomeres, satellites,
spacers between genes. Constitutive heterochromatin is
the same in all cells.
Fig. 3. Structure of nucleus
LEVELS OF DNA CONDENSATION
I. Nucleosomal. Decondensed chromatin viewed in the electron microscope resembles “beads
on a string”. Each bead is a nucleosome containing 146 bp of DNA wrapped around the core
histone octamer, and sealed by a single molecule of linker histone (H1) bound at the point where
the DNA enters and exits. H1 also binds to the linker DNA (the string connecting one bead to the
next) (Fig. 2, 4). The histone octamer is composed of
two molecules each of the four core histones: 2xH2A,
2xH2B, 2xH3 and 2xH4. These are small, basic
proteins, which have been highly conserved during
evolution.
Histone H1, the linker histone, is not part of the
nucleosome core (Fig. 5). It is very rich in lysine
residues, less well conserved than the core histones,
and is larger. The length of DNA in the core particle
Fig. 4. Schematic diagram of nucleosome core
is invariable (146particle
bp), but the average length
of the linker DNA varies between species and
tissues, giving rise to a characteristic repeat
length (the average length of DNA in a
nucleosome).
Fig. 5. Binding of histone H1 to the linker DNA
II. Chromatin folding (Solenoid). Most of the chromatin in the nucleus is in the form of a
highly condensed filament about 30-nm in
diameter (the 30-nm filament). Formation of
these highly condensed filaments is dependent
on H1 (one H1 molecule per nucleosome). The
condensed 30-nm filament is the result of
solenoidal (helical) folding of the beads-on-aFig. 6. The solenoidal model of chromatin
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string nucleosomal (10 nm) filament, having 6-12 nucleosomes per turn (Fig. 6).
III. Chromatin Loops. The interphase
chromosomes are organized into loops of chromatin
filament attached to the nuclear skeleton (nuclear
matrix) at their bases and projecting into the interior
of the nucleus (Fig. 7). Each loop may contain a gene
or related cluster of genes whose expression may in
principle be regulated at the level of loop structure.
The regions of DNA, which interact with matrix, are
Fig. 7. Attachment of solenoid fiber to scaffold
called MARs (matrix attachment regions) or SARs
(scaffold attachment regions), while . The loops contain an average of 40 000 - 80 000 bp.
IV. Metaphase Chromosomes. When the nuclear membrane breaks down during mitosis,
chromatin is reorganized to form metaphase chromosomes in which a chromosomal metaphase
scaffold is folded to form a quite regular helical coil, to which
chromatin loops are attached at their bases (Fig. 8). Sister
chromatids are usually of opposite helical handedness. The
organization of DNA into the organelle, which is a chromosome,
is most obvious at metaphase when the chromosome is
condensed into a highly structured body. At this point in the cell
cycle important chromosomal elements are visible: two arms
(short – p and long – q), centromere and telomere.
The telomeres are special sequences responsible for
preventing the shortening of chromosomes during replication,
protect linear molecules of DNA against actions of exonucleases
and prevent joining of different chromosomes. Structurally, the
telomeres are rich in tandemly repeated G/C sequences with a
total length of 3 – 20 kb.
The centromere is the genetic element (formed by
constitutive heterochromatin) responsible for chromosome
segregation during cell division. It is visible at metaphase as a
Fig. 8. Structure of a chromosome
constriction in mammalian chromosomes and is the site of
attachment of spindle microtubules to the proteins that make up the kinetochore. Centromere
contains A/T rich repetitive sequences. Within centromeres, H3 histone is substituted by CENP-A
histone.
Chromatine activation:
Gene activity depends on: the stage of ontogenetic period, type of cell, environment. It also
depends on the level of DNA condensation. Transcription is associated with nucleosomal level
only.
Transcriptionally active chromatin regions have core histones undergoing high rates of
acetylation and deacetylation. Histone acetylation (which is a type of post-translational modification
of histones) is generally linked to gene activation. The enzymes that acetylate conserved lysine
amino acids on histone proteins by transferring an acetyl group (functional group COCH3) are
called histone acetyltransferases (HAT), which can also acetylate non-histone proteins, such as
transcription factors and nuclear receptors to facilitate gene expression. Removal of the acetyl group
by histone deacetylases (HDACs) condenses DNA structure, thereby preventing transcription.
Another common way to block DNA and inhibit gene transcription is DNA methylation, which
involves the addition of a methyl group (CH3) to cytosine or adenine.
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So, the post-translational modifications of histones include:
 Methylation of H3 (Lys4) – active expression of a gene;
 Methylation of H3 (Lys9) – diminishing of transcription;
 Histone acetylation – assisting the transcription, gene activation;
 Histone deacetylation – chromatin condensation, inactivation of transcription;
 Phosphorylation of H1 – chromatin supercoiling
 Dephosphorylation of H1 – chromatin decondensation.
NUCLEAR ENVELOPE
Nuclear envelope is a double membrane that
surrounds the nucleus during most of the cell's
lifecycle. The outer nuclear membrane is continuous
with the membrane of the rough endoplasmic
reticulum (ER), having numerous ribosomes attached
to the surface. The outer membrane is also continuous
with the inner nuclear membrane since the two
layers are fused together at numerous tiny holes called
nuclear pores that perforate the nuclear envelope.
These pores regulate the selective passage of
molecules between the nucleus and cytoplasm, The
space between the outer and inner membranes is
termed the perinuclear space and is connected with
the lumen of the rough ER.
Structural support is provided to the nuclear envelope by two different networks of
intermediate filaments. Along the inner surface of the nucleus, one of these networks is organized
into the nuclear lamina (made of fibrous proteins – nuclear lamins and membrane associated
proteins), which binds to chromatin, integral membrane proteins, and other nuclear components.
The nuclear lamina is also thought to play a role in directing materials inside the nucleus toward the
nuclear pores for export and in the disintegration of the nuclear envelope during cell division and its
subsequent reformation at the end of the process. The other intermediate filament network is located
on the outside of the outer nuclear membrane and is not organized in such a systemic way as the
nuclear lamina.
The amount of traffic that must pass through the nuclear envelope on a continuous basis in
order for the eukaryotic cell to function properly is considerable. RNA and ribosomal subunits must
be constantly transferred from the nucleus where they are made to the cytoplasm, and histones, gene
regulatory proteins, DNA and RNA polymerases, and other substances required for nuclear
activities must be imported from the cytoplasm. An active mammalian cell can synthesize about
20,000 ribosome subunits per minute, and at certain points in the cell cycle, as many as 30,000
histones per minute are required by the nucleus. In order for such a tremendous number of
molecules to pass through the nuclear envelope in a timely manner, the nuclear pores must be
highly efficient at selectively allowing the passage of materials to and from the nucleus.
Nuclear Pores
It is generally thought that a protein structure called the nuclear pore complex (NPCs) that
surrounds each pore plays a key role in allowing the active transport of a selected set of large
molecules into and out of the nucleus. The nuclear pore proteins are called nucleoporins, which are
not only engaged in nucleocytoplasmic transport, but also in transcription regulation, kinetochore
organization and other cellular events. Given the various functions of nucleoporins, it is not
surprising to find them involved in a wide variety of human disease.
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In addition to their role in nuclear transport, nuclear pores are important as sites where the
outer membrane and inner membrane of the nuclear envelope are fused together. Due to this fusion,
the membranes can be considered continuous with one another although they have different
biochemical characteristics and can function in distinctive ways. Since the outer nuclear membrane
is also continuous with the membrane of the endoplasmic reticulum (ER), both it and the inner
nuclear membrane can exchange membranous materials with the ER. This capability enables the
nuclear envelope to grow bigger or smaller when necessary to accommodate the dynamic contents
of the nucleus.
KARYOPLASM
The karyoplasm or nucleoplasm or nuclear sap is a highly viscous liquid contained within the
nucleus that surrounds the chromosomes and other subnuclear organelles. A network of fibers
known as the nuclear matrix (scaffold, skeleton) can also be found in the nucleoplasm.
Nuclear Scaffold
The skeleton maintains the overall size and shape of the nucleus. The matrix acts as a
structural attachment site for the DNA loops during the interphase: evolutionary highly conserved
300-1000 bp long DNA sequences, referred to as SARs (Scaffold Associated Regions), have been
identified that define the base of DNA loops, anchoring them to specific proteins (SAPs - Scaffold
Associated Proteins). By means of such chromosomal attachment sites, the matrix might help to
organize chromosomes, localize genes, and regulate DNA transcription and replication within the
nucleus.
NUCLEOLUS
Nucleolus is a part of nucleus responsible for biogenesis of ribosomes. It is the place of:
transcription of ribosomal genes and synthesis of precursor rRNA (45S), followed by processing of
45S rRNA and formation of 3 types of rRNA: 5.8S + 18S + 28S. As a result of assembling the RNP
is formed: rRNA 18S + 33 ribosomal proteins = 40S RNP (small ribosomal subunit) and rRNA 28S
+ rRNA 5.8S +rRNA 5S + 49 ribosomal proteins =60S RNP (large ribosomal subunit).
Sequences of DNA containing ribosomal genes (tandem repeats of rRNA genes) form the
nucleolar organizer region (NOR). The human genome contains more than 200 clustered copies of
the rRNA genes on five different chromosomes (13, 14, 15, 21, 22). Transcription of rRNA genes is
executed by RNA-polymerase I (rRNA 45S) and III (for rRNA 5S). Further processing is needed to
generate the 18S RNA, 5.8S and 28S RNA molecules. This processing involves the function of a
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class of small nucleolar RNAs (snoRNAs) which are complexed with proteins and exist as smallnucleolar-ribonucleoproteins (snoRNPs). Once the rRNA subunits are processed, they are ready to
be assembled into larger ribosomal subunits.
Fig. 9. Scheme of ribosome biogenesis.
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