CELL CYCLE

Volgograd state medical university
Department of histology, embryology, cytology
Lecture:
CELL CYCLE
for the 1st year
English medium students
Volgograd,
Volgograd, 2013
2013
1
OBJECTIVES:
1. To describe how the nucleus controls the reproduction and
vital biochemical processes of the cell.
2. To explain the rationale for the need for two types of
chromatin, a nucleolus and a limiting membrane in the
nucleus.
3. To conceive how the nucleus acts as the staging area for the
formation of chromosomes which are formed from chromatin
containing coils of DNA and protein.
4. To display the role of the nucleus in mitosis and meiosis.
5. To demonstrate the need for the various types of RNA
synthesized in the nucleus.
6. To understand how chromosomal abnormalities cause certain
clinical disorders, e.g., Down syndrome.
2
DIAGRAM OF THE CELL
Most of the cell functions are under control of
the nucleus, the administrative area of the cell.
Nucleus is the largest single membrane-bound
compartment in the cell and contains nearly all of
the DNA possessed by the cell as well as the
mechanism for RNA synthesis.
3
The Main Constituents of the Cell and Their Distribution
The nucleus is limited by two nuclear membranes – nuclear envelope and is
filled with nucleoplasm that contains genetic information encoded in a DNA
4
molecules.
The nuclear envelope is composed of the inner and the outer nuclear
membrane with an 10- to 30- nm thick intervening perinuclear cistern
between them. The two concentric membranes of the nucleus are united at
the nuclear pores. They play two different roles.
The outer nuclear membrane is studded with ribosomes on its
cytoplasmic surface and it is continuous with the RER. It is also about 6 nm
thick. It is considered as a specialized region of the RER. Its cytoplasmic
surface usually possesses ribosomes actively synthesizing transmembrane
proteins that are destined for the inner & outer nuclear membranes.
NUCLEAR
ENVELOPE
NP
N
Ec
rER
NE
Hc
NUCLEUS
5
NUCLEAR ENVELOPE
The inner membrane is
about 6 nm thick and faces the
nuclear contents.
It is lined
by
the nuclear lamina, an
interwoven
meshwork
of
intermediate filaments, 80 to
100 nm thick, composed of
three proteins termed nuclear
lamins A, B &C. The filaments
form a regular square lattice as
a scaffold beneath the nuclear
membrane.
The
nuclear
lamina
network interacts with nuclear
membrane proteins & acts as a
nuclear cytoskeleton, possibly
interacting with chromatin in
the spatial organization of the
nucleus. It is involved in the
envelope breakdown at mitosis.
euchromatin
nuclear
envelope
nuclear
lamina
heterochromatin
nucleolus
nuclear pore
endoplasmatic
reticulum
6
The interruptions in the nuclear envelope - nuclear
pores provide regulated passageways for the exchange of material
between the cytoplasm and the nucleoplasm. They are involved in
RNA translocation into cytoplasm by active transport and protein
import into nucleus.
The number of nuclear pores ranges between a few dozen to
several thousand correlated directly with the metabolic activity of
7
the cell. The diameter ranges between 70-100 nm.
NUCLEAR PORE, TEM
In TEM the nuclear pores (P) perforating the nuclear membrane
appear as gaps between the segments of the double nuclear membrane
(NM) binding the perinuclear space (PNS). The pores form channels
which allow transport of small molecules, between the cytosol & the
8
nucleus.
High-resolution EM has revealed that the nuclear pore is
surrounded by non-membranous structures embedded in its rim. These
structures are called nuclear pore complex. It selectively guards
passage through the pore. The evidence suggests that each of the
nuclear pore complex is in communication with the other via the
nuclear lamina and certain pore-connecting fibers.
Particles of 17,000 D rapidly
enter the nucleus while larger
particles enter more slowly and
those of 60,000 D have difficulty
entering.
A variety of proteins must be
imported
from
the
cytoplasm
through the pores into nucleoplasm,
including DNA & RNA polymerases,
histones, gene regulatory proteins &
RNA-processing proteins. Some of
them interact with receptors at the
pore
margin
&
are
actively
transported through a pore channel.
Nuclear Pore Complex
9
Nuclear Pore Complex
Proteins that are actively
transported
into
the
nucleus contain short
peptide sequences called
nuclear import signals. In
addition to protein import
each nuclear pore in a
rapidly
growing
cell
transports approximately
3 ribosomes per minute
to the cytoplasm.
The pore complex contains granular and filamentous subunits.
Structurally pores are rimmed by 8 protein complexes to form the
nuclear pore complex. The central granules of the pore complex are
believed to be large proteins or components of ribosomes in transit
between different cell areas.
10
Gartner.p.56. The NPC spans the two nuclear
membranes. It is composed of three ring-like arrays of proteins
stacked on the top of each other, each displaying eight-fold
symmetry and interconnected by a series of spokes arranged in
a vertical fashion. In addition NPC has cytoplasmic fibers, a
transporter & a nuclear basket.
The cytoplasmic ring composed of 8 subunits is located
on the rim of the cytoplasmic aspect of the nuclear pore. Each
subunit possesses a filamentous fiber, believed to be a Ranbinding protein (a family of GTP binding proteins), that extends
into the cytoplasm. These fibers may mediate import into the
nucleus through the NPC by moving substrates along their
length toward the center of the pore.
The middle ring is composed of a set of 8
transmembrane proteins that project into the lumen of the
nuclear pore as well as into perinuclear cystern. These spokelike proteins appear to anchor the glycoprotein components of
the NPC into the rim of the NP.
11
NUCLEAR PORE COMPLEX
cytoplasmic
ring subunit
scaffold
nucleoplasmic
ring subunit
thick filament
transporter
unit
outer nuclear
membrane
inner nuclear
membrane
basket
The
nuclear pore is surrounded by non-membranous
structures embedded in its rim. These structures are called nuclear
pore complex (NPC). It selectively guards passage through the pore.
The pore complex contains granular and filamentous subunits.
Structurally pores are rimmed by 8 protein complexes to
12
form the nuclear pore complex.
The center of the middle ring is occupied by an hourglassshaped structure known as transporter, which is coupled to the
spoke-like proteins of the middle ring.
The central lumen of the middle ring is a gated channel that
restricts passive diffusion between the cytoplasm & nucleoplasm. It is
associated with additional protein complexes that facilitate the
regulated transport of materials across the nuclear pore complex.
A nucleoplasmic ring, analogous to the cytoplasmic ring, is
located on the rim of the nucleoplasmic aspect of the nuclear pore
and assists in the export of several types of RNA. A filamentous,
flexible, basket-like structure, the nuclear basket, appears to be
suspended from the nucleoplasmic ring and protruding into the
nucleoplasm. It becomes deformed during the process of nuclear
export.
Nuclear pore complex has a relative molecular mass of over 1
– megadaltons, but only a few of the molecules have been
characterized. A transmembrane glycoprotein called gr 210 extends
into the lumen of the nuclear pore. It has been suggested to play a
role in pore complex formation and may anchor the pore complex
material in the nuclear envelope.
13
Although the nuclear pore is relatively large, it is nearly filled
with the structures constituting the NPC. Because of the structural
conformation of those subunits, several 9- to 11-nm wide channels
are available for simple diffusion of ions & small molecules. However
molecules & particles larger than 11 nm cannot reach or leave the
nuclear compartment via simple diffusion; instead they are
selectively transported via a receptor-mediated transport process.
Signal sequences of molecules to be transported through the
nuclear pores must be recognized by one of the many receptor sites
of the NPC.
The bidirectional traffic between the nucleus & cytoplasm is
mediated by a certain group of proteins known as 1)exportins, which
transport macromolecules (RNA) from the nucleus to the cytoplasm
& importins which transport cargo (protein subunits of the
ribosomes) from the cytoplasm to the nucleus. Exportin & importin
function is regulated by a family of GTP-binding proteins known as
Ran. Transport across the NPC is usually but not necessarily always,
an energy-requiring process.
14
HIGHLIGHTS:
-the nuclear envelope is composed of two parallel unit
membranes (inner & outer nuclear membranes) that
enclose a perinuclear cystern,
- the membranes fuse with each other at certain regions to
form perforations known as nuclear pores;
-nuclear pores provide communication between the nucleus
and the cytoplasm;
-the nuclear pore complex is composed of the nuclear pore
and its associated glycoproteins;
-the nuclear pore functions in bidirectional nucleoplasmic
transport;
-the nuclear lamina is a scaffolding which maintains the
15
shape of the nucleus.
nucleus = nuclear envelope + nucleoplasm
16
The nucleoplasm:
-contains macromolecules & nuclear particles
involved in the maintenance of the cell,
-houses two major components:
-chromatin, the genetic material of the cell;
-the nucleolus, the center for ribosomal RNA (rRNA)
synthesis;
17
NUCLEUS
DNA, the cell’s genetic
material, resides in the nucleus
in the form of chromosomes,
which are clearly visible during
cell division. In the interval
between the cell divisions the
chromosomes are unwound in
the form of chromatin.
Nuclear DNA is tightly packed by association with special
proteins and forms the chromatin.
Depending on its transcriptional activity chromatin may be
condensed as heterochromatin (dense-staining) or extended as
18
euchromatin (light-staining).
Neutrophilic
Granulocyte, TEM
The distribution of chromatin is not uniform, and this reflects
varing degrees of unfolding according to whether genes are being
transcribed. Heterochromatin is often seen adjacent to the nuclear
membrane. The remaining of the chromatin, scattered throughout the
19
nucleus is euchromatin.
Hepatocyte,
x48,176.
TEM
Euchromatin
represents the active form
of chromatin where the
genetic material of the
DNA molecules are being
transcribed into RNA.
20
Nucleus
and
cytoplasm. Liver.
Mouse. Electron
microscopy.
X
20,318.
When euchromatin is examined with EM it is seen to
be composed of thread-like material 30 nm thick.
21
condensed section
of chromosome
30 nm
Gartner p.57 3-7
chromatin
fiber of
packed
nucleosome
“beads-on-astring”
form of
chromatin
11 nm
2 nm
300 nm
700 nm
1400 nm
metaphase
chromosome
extended section
of chromosome
DNA double helix
In chromosomes this is then wound again into a supercoiled
structure. These threads may be unwound resulting in 11-nm wide
structure resembling “beads on a string”. The beads are termed
nucleosomes, which are DNA molecules are organized around histones.
The nucleosomes are wound into a helix to form chromatin.
22
DNA
nucleosome
beads
nucleosome
packing
N
supercoiled
coiled
extended
chromosome chromo- metaphase
chromosome
Some
Stevens p.16 (2-10)
histone
DNA
2 nm
nucleosome
1.5 mcm
11 nm
700 nm
300 nm
FORMATION OF THE CHROMOSOMES
The nucleus contains DNA wound around proteins called histones to form
nucleosomes (N), which are regularly repeating globular structures similar to
beads on a string. The nucleosome string is then wound into filaments 30 nm
in diameter, which make up the structure of chromatin. Further condensation
into distinct chromosome is possible during cell replication when chromatin
forms large looped domains by attachment to DNA-binding proteins.
23
The nucleus houses chromosomes and is the location of RNA
synthesis. Both mRNA & tRNA are transcribed in the nucleus
whereas rRNA is transcribed in the nucleolus.
Process of Transcription
24
HIGHLIGHTS:
-Nuclear DNA is organized around histones into nucleosomes. The
nicleosomes are wound into a helix to form chromatin.
-Chromatin is a complex of DNA and proteins and represents relaxed
uncoiled chromosomes of the interphase nucleus.
-Nucleus chromatin is divided into two parts: heterochromatin (H) is
a dense- staining, while euchromatin is light-staining.
-Euchromatin represents actively transcibed cellular DNA.
-Heterochromatin is a highly condensed transcriptionally inactive
form of DNA.
-Further looping of the chromatin results in formation of the
supercoiled structures – chromosomes.
-Chromosomes are chromatin fibers that become so condenced and
tightly coiled during mitosis and meiosis that they are visible under
25
the light microscope.
The peripheral chromatin and
chromatin islands are clearly
evident, as is the nucleolusassociated chromatin (NC).
NC
Enterocyte
Lymphocyte
NC
26
Hepatocyte, TEM
The clear area within the nucleus is the nucleoplasm
representing the fluid component of the nucleus.
27
Ganglionar Cells
of the
Cerebellum,
TEM
The
nucleoplasm
consists
of
interchromatin
and
perichromatin
granules,
ribonuleoprotein
particles and the
nuclear matrix.
Interchromatin granules are which are 20 to 25 nm in diameter,
contain RNPs & several enzymes including ATPase, GTPase, betaglycerophasphatase & NAD-pyrophasphatase. They are located in
clusters scattered throughout the nucleus among the chromatin
material and appear to be connected to each other by thin fibrils. Their
28
function is unknown.
Schwann Cell, TEM
Perichromatin granules are 30 to 50 nm in diameter and are
located at the margins of heterochromatin. These electron-dense
particles are surrounded by a 250nm wide halo of a less dense region.
They are composed of densely packed fibrils of 4.7S low-molecularweight RNA complex to two peptides, resembling heterogenous nuclear
29
ribonucleoproteins.
Neurocytes
of the Retina,
TEM
Small nuclear ribonucleoprotein particles participate in splicing,
cleaving heterogenous RNPs. Although most of them are located in the
nucleus, some are limited to nucleoli. Several minor subgroups of these
particles have been discovered recently, but their function has not yet
30
been elucidated.
Nuclear matrix is defined both
in structural & biochemical terms.
It contains 10% of total protein, 30%
of RNA, 1-3% of total DNA and 2-5%
of total nuclear phosphate. The
structural components include the
nuclear pore – nuclear lamina
complex, residual nucleoli, residual
RNP networks and fibrillar elements.
Functionally the nuclear matrix is
associated with DNA replication sites,
rRNA & mRNA transcription &
processing, steroid receptor binding,
heat shock proteins, carcinogen
binding, DNA viruses & viral proteins.
It has been suggested that the
nucleus may contain many interactive
subcompartments
that
function
spatially and temporally in a tightly
coordinated fashion to facilitate gene
expression.
Pituitary (Gonadotroph),
Rat, x8,936.
(asterisk – secretory granules).
31
NUCLEOLUS
The nucleolus presents a sponge-like appearance composed of
electron-lucent and electron-dense materials, suspended free in the
nucleoplasm. The electron-dense region is composed of pars
granulosa and pars fibrosa, while the electron-lucent region is the
32
nucleoplasm in which the nucleolus is suspended.
Macrophage,
TEM
The nucleus contains the chromatin and the nucleolus. The
nucleolus is clearly visible as an electron-dense circular area. It
measures 1-3 mcm in diameter, increasing in size with active gene
transcription and occupying up to 25% of the nuclear volume. Inactive
cells have indistict nucleoli while metabolically active cells have large
33
or multiple nucleoli.
Glandulocyte of
the Sebaceous
Gland
Parietal cell of the Stomach
34
Nucleolus
35
The nucleolus
is the deeply staining nonmembranous-bound structure within the nucleus that is
involved in rRNA synthesis and in the assembly of small
and large ribosomal subunits.
It is observed only during interphase because it
dissipates during cell division.
Densely staining regions are nucleolus-associated
chromatin which is being transcribed into rRNA.
NUCLEOLUS
NP
N
Ec
rER
NE
Hc
NUCLEUS
36
Adenohypophysis, TEM
Pars amorpha (pale
areas), so called nuclear
organizer
regions
with
specific
RNA-binding
proteins, correspond to
large loops of transcribing
DNA
containing
the
ribosomal RNA gene;
Pars fibrosa (densestaining region), contains
nucleolar
RNAs
being
transcribed;
Pars
granulosa
(granular regions) in which
maturing
ribosomal
subunits are assembled
EM showing the nucleolus from a cell actively producing protein. The
pars amorpha (A), pars fibrosa (F) and pars granulosa (G) are clearly
visible.
37
Also located in the pale-staining regions are the tips of
chromosomes 13-15, 21, 22 (in humans) containing the nucleolar
regions (NORs), where gene loci that encode rRNA are located.
NUCLEOLUS, TEM, x 80,000
38
CLINICAL CORRELATIONS:
In malignant cells the nucleolus may
become hypertrophic!
39
COMPARATIVE CHARACTERISTICS OF THE NUCLEAR CONSTITUENTS
nuclear
constituents
structure
function
nuclear
envelope
consists of inner and outer membranes
that become continuous around
nuclear
pores;
outer membrane
studded
with
ribosomes,
inner
membrane smooth; are separated by a
perinuclear cistern, nuclear pore
complex is associated with each pore
specialized segment of
endoplasmatic reticulum
that bounds nucleus,
nuclear
pores
permit
communication between
cytoplasm
and
nucleoplasm
nuclear
lamina
thin network of interwoven filaments
stabilizes inner nuclear
membrane;
attachment
site for components of
nucleoplasm
heterochromatin
dense staining, condensed chromatin
inactive, part of genome
not being expressed
euchromatin Light staining, dispersed chromatin
nucleolus
active, part of genome
being expressed
conspicious round body in nucleus, synthesis of ribosomes
nucleolonema consists of dense
granules in a matrix of filaments;
40
amorphous component may be present
Motor Neuron. Rhesus monkey, methylene blue and erythrosin, 612 x.
The fact that nuclei of
certain cells differ in appearance
and staining properties can be
directly related
to nuclear
function. This is a functionally
active nucleus of the motor
neuron located within the spinal
cord, but its elongated axon
leaves the cord to terminate on
striated skeletal muscle fibers.
A motor neuron is characterized by a multipolar perikaryon, cytoplasm
rich in Nissl bodies (ribonucleoproteins), and a stout dendrite, which
can be identified because it is structurally similar to the cell body from
which it arises. The nucleus, centrally placed within the cell body, is
spherical and unstained (contains euchromatin) and has a prominent
41
densely staining basophilic nucleolus rich in ribonucleic acid.
B1. Basophilic & B2. Orthochromatic
erythroblasts.
Human, air-dried smear, Wright's*
stain, 1416 x.
Compare the appearance of
the functionally active nucleus of the
motor neuron with that of the
functionally inactive nucleus of the
maturing red blood cell.
B2: The small pyknotic nucleus of an
orthochromatic erythroblast (normoblast).
The chromatin (heterochromatin) is very
compact & densely stained. Such
a
nucleus is functionless and is ultimately
discarded, resulting in a mature nonnucleated erythrocyte.
B1: A basophilic erythroblast, one of the early cells in the development of the red blood corpuscle. The nucleus occupies a significant
portion of the cell and that its stainable inactive chromatin is coarse. A
nucleolus is present but obscured by stainable heterochromatin. The
cytoplasm of this cell contains ribonucleic acid, giving it a lavender
42
hue. Compare this cell with a mature non-nucleated RBC.
Monocytes,
human, airdried blood
smear,
Wright's stain,
4416 x.
Monocytes are the largest cells found in normal blood. The
nucleus is centrally or peripherally located, indented, and ovoid or
horse-shoe- shaped; the nuclear chromatin is not as dense as that of
lympho-cytes. Cytoplasm is abundant and contains azurophilic
granules. Monocytes are voracious phagocytes. The monocyte seen on
the extreme right shows pseudopodia extending from the cell body and
contains a phago-cytized red cell nucleus. Note the comparative size of
43
erythrocytes and monocytes.
NUCLEUS
Female sex chromatin
Peripheral blood smear,
Human, air-dried smear,
Wright's stain, 1416 x.
Sex
chromatin
(Barr*
body), composed of one of the X
chromosomes
that
remains
condensed in interphase, is
usually seen as a discrete
structure in the nucleus. Sex
chromatin is presumably present
in all cells in the female,
but, in the majority of cells, it is
obscured within the nucleus.
Portion of a peripheral blood smear showing abundant red blood
cells and a single neutrophilic granulocyte. The deeply stained chromatin
of the mature neutrophil nucleus is characteristically segmented or
multilobed. The lobes are attached by thin chromatin threads. Attached to
one of the lobes is a small "drumstick" appendage, the sex chromatin.
44
This appendage is found in about 3 percent of the neutrophils in females.
NUCLEUS
Female sex chromatin
B. buccal epithelium, human,
buccal epithelium scraping,
aceto-orcein stain, 500 x.
C. corpus luteum.
H. & E., 1416 x.
B: A buccal smear was prepared by scraping the stratified squamous
epithelium of the oral cavity. The stratified squamous cell is poorly
delineated, but the nucleus of the cell is stained and shows a sex
chromatin body (reddish-brown coloration) on the nuclear membrane.
The sex chromatin body may appear convex or triangular in shape. It
may also lie near the center of the nucleus.
C: A nucleus of the granulosa lutein cell with a sex chromatin body
45
adhering to the nuclear membrane.
CHROMOSOMES
Metaphase chromosomes, peripheral lymphocyte culture,
air-dried trypsin-banded, Giemsa* stain, 2142 x.
68-hour
peripheral
lymphocyte
culture,
phytohemagglutininstimulated to
induce
lymphocyte
transformatio
n and mitosis.
Mitosis has been arrested in metaphase (when they are
maximally condensed) with colchicine, and the chromosomes have
been dispersed with hypotonic potassium chloride permitting pairing
and numbering of chromosomes via a conventional system of
karyotyping – an analysis of chromosome number. Interphase nuclei
46
are seen adjacent to the dispersed chromosomes.
CHROMOSOMES. Karyotype Human, peripheral lymphocyte culture
68-hour peripheral
lymphocyte
culture,
phytohemagglutininstimulated to induce
lymphocyte transformation and mitosis.
Analysis of male chromosomes seen in the previous slide. This
arrangement of the chromosomes into groups is known as a karyotyping. Note the dark and light areas (bands) that characterize each
chromosome. Cells containing the full complement of chromosomes
(46) are said to be diploid (2n). Germ cells are haploid (1n).
Note the 22 pairs of autosomes (Nos. 1 to 22) and the pair of sex
chromosomes (XY in this case). The 22 pairs of chromosomes have
been classified on morphological grounds into 7 groups (A to G).
The chromosomes seen here are judged to be normal. Abnormal,
duplicated, or missing chromosomes can be related to defective
47
somatic and mental development in man.
CLINICAL CORRELATIONS:
-One item that may be observed from the karyotype is aneuploidy,
an abnormal chromosome number.
-People with Down syndrome have an extra chromosome 21
(trisomy 21), they exhibit mental retardation, stubby hands, and
many congenital malformations, especially of the heart, among other
manifestation.
-Certain syndromes
are associated with abnormalities in the
number of sex chromosomes. Kleinfelter syndrome results when an
individual possesses three sex chromosoms (XXY). These persons
exhibit the male phenotype but they do not develop secondary
sexual characteristics and are usually sterile.
-Turner syndrome is another example of aneuploidy called
monosomy of the sex chromosomes. The karyotype exhibits only
one sex chromosome (XO). These individuals are females whose
ovaries never develop and who have undeveloped breasts, small
48
uterus and mental retardation.
HIGHLIGHTS:
CELL NUCLEUS:
-bounded by a double nuclear membrane (nuclear
envelope),
-contains the cellular DNA as chromatin,
-contains the nucleolus responsible for making
ribosomes,
-moves substances in and out through nuclear pores.
49
Practical Histology. General principles can be applied usefully
when looking at cells either in cytological preparations to
assess their activity.
Nucleus:
-A metabolically inactive cell has a compact round nucleus
which typically stains intensely as little chromatin is being
transcribed. No nucleoli are visible as ribosome production is
minimal.
-A protein –synthesizing cell has a large pale-staining nucleus
with large or multiple nucleoli, reflecting active transcription
of chromatin. Similar nuclear changes are evident in cells in
an active phase of multiplication.
-A dead cell has a shrunken nucleus, which appears as an
amorphous compact mass of intensely staining material. This
later fragments into separate particles, and is completely
lysed, leaving the cell devoid of any discernable nucleus.
50
Fig.1.
Mitosis,
whitefish
blastula, x270.
Different stages of mitosis are
shown. Prophase displays the
short thread-like chromosomes
in the center of the cell. The
nuclear membrane is no longer
present. During metaphase the
chromosomes line up at the
equatorial plane of the cell. The
chromosomes begin to migrate
toward the opposite poles of
the cell in early anaphase and
proceed farther and farther
apart as anaphase progresses.
Note
the
dense
regions
(centrioles) toward which the
chromosomes migrate.
51
Fig.1. Mitosis, whitefish blastula,
x540.
During the early telophase stage of
mitotic division, the chromosomes
have reached the opposite poles of
the cell. The cell membrane
constricts to separate the cell into
the two new daughter cells, forming
a cleavage furrow (arrowheads). The
spindle apparatus is visible as
parallel, horizontal lines (arrow) that
eventually form the mid-body. As
telophase progresses, the two new
daughter cells will uncoil their
chromosomes and the nuclear
membrane
and
nucleioli
will
become re-established.
52
Cell cycle.
A diagram
demonstrating the
cell cycle in actively
dividing
cells.
Nondividing cells ,
such as neurons,
leave the cycle to
enter the Go phase
(resting
stage).
Other cells, such as
lymphocytes, may
return to the cell
cycle.
53
Mitosis. x 9423.
Observe the interphase nucleus possessing a typical nuclear
envelope, perinuclear chromatin,nucleolus, nuclear pores. A cell
undergoing the mitotic phase loses its nuclear membrane and
nucleolus while its chromosomes are quite visible. These
chromosomes are no longer lined up at the equatorial plate but are
migrating to opposite poles, indicating that the cell in the early-to midanaphase stage of mitosis. Organelles are present in the cytoplasm.
54
M
CELL
CYCLE
G2
GO
cell cycle
(dividing cell)
S
G1
exit from
cell cycle
(nondividing
cells)
Cells can enter a
phase
of
proliferation
in
which they divide.
Cells which leave
the cycle are said to
be in the G0 phase.
Cell division for growth and renewal is achieved by the process
of mitosis.
An essential feature of development is the ability of cells to
divide and reproduce. In addition, death of mature cells in the adult
needs to be compensated for by production of new cells.
Cells reproduce by duplicating their contents & dividing into 2
daughter cells. The phases involved in cell replication can be regarded
as a cell cycle. The phases of cell division are visible histologically and
involve duplication of cellular cytoplasmic contents, duplication of
DNA, separation of cellular DNA into two separate areas of the cell and
55
finally cell division (cytokinesis).
THE cell cycle is divided into two major events:
Mitosis, the short period of time during which the cell divides its
nucleus and cytoplasm, giving rise to two daughter cells, and
interphase, a longer period of time during which the cell increases its
size and content and replicates its genetic material.
56
In most tissues only a small
proportion of cells will be in
the cell cycle, the majority
being differentiated cells in
a G0 phase. Stem cells may
be in a G0 phase and only
come to re-enter the cell
cycle if there is a demand,
for example following cell
death.
CELL CYCLE
The DNA of cells is only replicated during certain phases of a cell’s growth
pattern, which takes place in several stages. The cell cycle is divided into two
main periods: mitosis and interphase which includes G1, S & G2 phases. G1
cells have just entered a phase of cellular growth. S phase cells actively
synthesize DNA, G2 cells have a double compliment of cellular DNA and are
resting prior to cell division, and M phase cells are in mitosis which comprises
5
57
stages.
CELL CYCLE
58
nuclear membrane
centriole center
of spindle
centromere
two sister chromosomes
microtubules
held together at centromere of spindle
The duplicated chromatin becomes
condensed
into
parallel
sister
chromosomes imparting a coarse stippling
to the nuclear region, which is associated
with loss of the nucleolus.
The centriole replicates to form two
microtubule organizing centers at opposite
poles of the cell (the mitotic spindle).
spindle
pole
nuclear
membrane vesicle
STAGES OF MITOSIS.
PROPHASE.
59
kinetochore microtubule
polar microtubule
The nuclear membrane breaks down to form small
vesicles, allowing the microtubules of the spindle to
interact with chromosomes. Each chromosome pair has an
attachment site (kinetochore) which binds to spindle
microtubules from each pole of the spindle (kinetochore
tubules). The chromosome pairs move to the center of the
spindle.
MITOSIS. Prometaphase.
60
cell equator
Movement of chromosomes along the microtubules lead
to alignment of the chromosomes at the equator of the cell
between the poles of the spindle.
MITOSIS.
Metaphase.
61
nuclear
envelope
vesicles
migrate
towards
poles
chromatids
pulled toward
pole of
spindle
.
elongation of
polar microtubule
shortened
kinetochore
microtubule
The kinetochore attachments to the paired
chromosomes separate and the chromosomes move to
opposite poles of the spindle. In late anaphase the spindle
microtubules elongate causing elongation of the cell and
further separation of the spindle poles.
MITOSIS. Anaphase.
62
chromosome decondense and
lose microtubular attachment
nuclear envelope reforms
The separated chromatides become separated from the
kinetochore microtubules and the nuclear membrane
reforms around each group of chromosomes. The cell
elongates further by elongation of the spindle
microtubules. This phase signals the end of mitosis.
MITOSIS. Telophase.
63
centriole
actin-myosin belt
nuclear membrane
Cleavage in two separate cells is produced by aggregation of an
actin-myosin belt immediately beneath the equator of the telophase
cell. The connecting region eventually separates with fusion of cell
membranes to form two daughter cells. At this stage a nucleolus
appears in the dense chromatin mass in the newly formed nucleus.
The cell now enters the G1 phase of the cell cycle.
MITOSIS. Cytokinesis.
64
Meiosis
Meiosis results in
the
formation
of
four
daughter cells, each with
half
the
normal
chromosomal complement
(i.e.haploid).
65
Anti-cancer drugs.Many drugs used to treat cancer act specifically on
cells in the cell cycle, the aim being to remove abnormally growing
cells. Unfortunately these drugs act on normal body cells as well as
cancer cells and have adverse effects particularly on renewing cell
populations, which depend on a high proportion of cells being in cycle.
Thus, blood cell production, hair production and gut-lining cell
production are all impaired by the administration of such anti-cancer
drugs. Nuclear changes in cancer. A cell containing a very large
nucleus relative to the amount of cytoplasm is generally in a phase of
cell division. Cells with unappropriately large nuclei raise the
suspicion od cancerous change. For example cells on the surface of the
uterine cervix should have small nuclei unless there is abnormal cell
growth such as that associated with the development of a malignancy.
In any specialized cell type, all the nuclei in adjacent cells should be
roughly the same size and have the same staining characteristics. In
cancer, however, nuclei vary in size and shape (nuclear
polymorphism) and commonly show dense-staining chromatin in a
66
coarse clumped pattern (nuclear hyperchromatism).
REPLICATION
The two strands of a DNA molecule first separate, & a new strand is
then synthesized alongside each of the separate strands. The result is that each
newly formed double-stranded molecule is identical to the original molecule
67
whose strands became separated.
Part of a doublestranded DNA molecule,
showing the deoxyribonucleotides of one strand
joined to those of the second
strand. Adenine is joined to
thymine and cytosine is
joined to guanine.
68
The Process of Transcription.
69
CELL DIVISION, Lymph node,
Rat, Mallory's stain, 1416 x.
This plate illustrates the
nuclear events in mitosis.
Interphase
A): Non-dividing or
resting stage. The chromatin
appears as an irregular
reticular meshwork. The
nuclear membrane, or
envelope, and the nucleolus
are distinctly seen.
Chromosomes are not visible.
Early prophase (B): NM
& nucleolus disappear. Granularity of the N. is markedly increased, and
filamentous structures are seen. These granules and filaments represent
the chromosomes, which become shorter and thicker in this stage.
Late prophase (C): The thread- or rod-like character of the
chromosomes is more apparent. Each chromosome consists of two
coiled chromatids, which are not visible in this preparation. The
disappearance of the nuclear membrane allows mixing of nuclear and
70
cytoplasmic material.
CELL DIVISION
Lymph node, Rat,
Mallory's stain, 1416 x.
Metaphase (D): Chromosomes
appear condensed and line up in
the equatorial plane (metaphase
plate) of the cell. Each chromosome is still composed of two
paired chromatids.
Anaphase (E): The daughter
chromosomes (chromatids)
separate and are drawn to
opposite poles of the cell. They
remain separate and tightly
coiled, and appear at this
magnification to be fused.
Cytoplasmic division begins.
Telophase (F): The two distinct groups of daughter
chromosomes (chromatids) appear fused and tightly packed.
Cytoplasmic division is completed. Nuclear membranes re-form and
71
nucleoli reappear.
Division of HeLa cells in culture, SEM.
The rounded cell
elongates
progressively &
a cleavage furrow
appears
The cell are
pushed
farther
apart, but a narrow
intercellular
bridge
persists
before
separation
72
occurs
PHASES OF SPERMATOGENESIS
Spermatogonia are influenced by testosterone at puberty to enter the cell
cycle. AD spermatogonia are reserve cells, they either proliferate
proliferate or
differentiate into AP.
Phase 1. Spermatocytogenesis (Multiplication Phase)
Nucleus
Spermatogonia
round nucleus, deeply
staining speckled chroma
tin, central pale nuclear
vacuole, one or two small
Nucleoli
On the contrary to
female gametogenegametogenesis, male germ cell
mitosis
TYPE AD
population is conticontinuously
renewing
round nucleus, pale staining
system, that depdepnuclear chromatin, one or
ends on spermatotwo nucleoli
spermatomitosis
TYPE AP
TYPE AD
cyte
replacement
from
the
stem cells,
coarsely clumped
DNA replication
4n
chromatin, pale central
and spermatogenespermatogenenucleolus
sis is sustained even
TYPE B
in old men.
AP spermatogonium has an equal chance of remaining as pale type (self(self73
renewal by division) or to differentiate into B spermatogonia.
Phase II of the SPERMATOGENESIS –
Growth (meiotic prophase, 22 days).
Nucleus
Primary spermatocytes
meiosis
similar to type B
PRELEPTOTENE
fine filamentous
chromatin with fine
Beading
LEPTOTENE
fine filamentous
chromatin with
coarse beading
ZYGOTENE
EARLY
PACHYTENE
large nuclei, thick
prominent short
rods of chromatin
LATE
PACHYTENE
B-spermatogonia lack
capacity
for
selfselfrenewal. They divide
and give rise to
primary spermatocyspermatocytes.
Shortly after their forformation primary sperspermatocytes
duplicate
their chromosomes to
obtain 4n DNA content
and diploid chromochromosome number (2n)
DIPLOTENE
74
Phase III – Maturation
(Meiotic divisions I and II)
1st meiotic division
secondary
spermatocyte
(2n)
2nd meiotic division
small central
nucleus, finely granular
chromatin
with occasional larger
chromatin
masses
spermatids
(n)
Phase IV Transformation
(spermiogenesis)
spermatozoa
(n)
During
1st
meiotic
division
the
DNA
content is halved (to
2n DNA) and the
chromosome number
reduces to haploid.
During the 2nd meiotic
division
the
DNA
content
of
each
daughter cell reduces
to haploid (1n DNA)
whereas the chromochromosome number remains
unaltered (haploid).
75
SCHEME OF SPERMATOGENESIS
76
The process of spermatogenesis from
spermatogonia to mature spermatozoa
requires about 64 days in man.
77
Mitotic Division of the Cell. Prophase
In prophase the cell assumes
a more spherical shape & appears
more refractile, while in the nucleus
chromosomes become more visible
and appear as threadlike structures.
Each chromosome consists of two
coiled chromatids that are closely
associated along their length. Each
chromosome contains a double strand
of DNA. As prophase progresses, the
chromatids continue to coil, thicken &
shorten, reaching 1/25th of their length
by the end of prophase.
Then chromosome approach
the nuclear envelope. Nucleoli become
smaller & finally disappear. The
nuclear envelope breaks down. When
this occurs the center of the cell
becomes
more
fluid,
&
the
chromosomes move freely, making
their way to the equator of the cell.
Simultaneous with the nuclear events the centrioles replicate, and the
resulting pairs migrate to the opposite poles of the cell.
78
The disappearance of the nuclear envelope mark the end of
prophase and the beginning of metaphase, which is characterized by
formation of the mitotic spindle and the alignment of chromosomes along
the equator to form the equatorial plate. The spindle is formed of
microtubules. They extend from the poles of the spindle to attach to the
centromere (kinetochore) of each chromosome and form the chromosomal
fibers.
METAPHASE
The
centromere is a specialized
region of unduplicated
DNA & protein that
holds
together
the
chromatid
of
each
chromosome
and
forms an attachemnt
site for the chromosomal fibers. The final
act of metaphase is
simultaneous
duplication of the DNA
at the centromeres of
all chromosomes, after
which the centromeres
79
split.
MITOSIS. ANAPHASE.
The
initial
separation
of
the
chromatids marks the
beginning of anaphase.
All
the
chromatids
simultaneously move to
the opposite poles, the
centromeres
travel
in
advance of the telomeres
(arms) that train behind.
Movements
of
chromatids which are now
daughter chromosomes is
an
active
dynamic
process.
80
As
daughter
chromosomes
reach
their respective poles,
discontinuous portions
of ER form around each
their group and begin to
re-form
nuclear
envelope.
This
event
initiates
telophase.
When nuclear envelope
is completely reformed,
the
chromosomes
uncoil,
become
indistinct and the 2
nuclei reassume the
interphase
configuration.
The
normal complement of
MITOSIS. TELOPHASE.
nucleoli reappears at
this time.
Their development is associated with NORs present on certain
chromosomes. This is the end of karyokinesis. The mitotic spindle begins to
81
disappear.
Blood Smear, Giemsa, x 800.
The fibers of the spindle between the two forming nuclei appera
stretched, the are termed interzonal fibers. Midway between the two nuclei, in
the region formerly occupied by the equatorial plate, a constriction of the
plasmalemma forms a furrow that extends around the equator of the cell. The
constriction extends deeper into the cytoplasm, separating the daughter
chromosomes until they are joined only by a thin cytoplasmic bridge called
midbody containing interzonal fibers. Eventually the daughter cells pull away
from each other by ameboid movement thus completing the separation of the
82
cells & ending cytokinesis.
The Result of the Cytokinesis:
1) Cell organelles are evenly distributed between the
daughter cells.
2) Immediately after division the daughter cells enter a
phase of active RNA and protein synthesis resulting in
an increase of the volume of both nucleus and
cytoplasm.
3) The ER and Golgi are restored to their original
concentrations.
4) Mitochondria reproduce by fission, centrioles replicate
in the daughter cells just before next division.
83
?
?
84
?
?
85
?
?
86
87
88
89
90
The Fate of Each
Homologous Pair of
Chromosomes in
Mitosis & Meiosis.
1.
Meiosis includes 2
sequentional cell
divisions, the 1 st
resulting in reduction of
chromosome
complement to haploid
and the 2nd resulting in
the production of 4
haploid daughter cell.
2.
2.Chiasmata formation
occurs in meiosis only
so that every gamete is
genetically different.
91
92
93
94