Non-membranous organelles
Cell inclusions
Cell cycle
Institute of Histology and Embryology
Doc. MUDr. Zuzana Jirsová, CSc.
Histology and Embryology – B81131
Lecture - 9. 10. 2013
NON-MEMBRANOUS ORGANELLES
(Nucleolus)
Ribosomes
Centrioles
Cytoskeleton
actin filaments/microfilaments
intermediate filaments
microtubules
RIBOSOMES
Function: translation of mRNA, protein synthesis
Structure
Electron dense granular particles (20-30 nm in diameter)
composed of the ribonucleoproteins (rRNA and proteins)
Ribosome consists of the small and large subunits
Distribution: free or attached to membranes of ER (rough ER)
Ribosomes synthesizing proteins occur in clusters – polyribosomes
(polysomes), individual ribosomes are held together by a strand
of mRNA
Synthesis on free polyribosomes: proteins for use within the cell
(hemoglobin, actin, myosin, proteins of the intermediate filaments,
tubulin, most mitochondrial enzymes)
Synthesis on the ribosomes attached to membrane of ER
(proteins are segregated into the cisternae of ER): proteins that will
be secreted, lysosomal enzymes, and integral membrane proteins
Scheme of the protein synthesis on free
polyribosomes (Junqueira´s Basic Histology, Mesher, 2010)
ribosome
rER
rER
mRNA
rER
free protein
in cytoplasm
Electron micrograph: Histology, Ross, Pawlina 2010
Arrow: free polyribosomes
Rough endoplasmic reticulum: rER
CENTRIOLES
Mitotic spindle (cell in metaphase of mitosis)
centrioles
MT
ch
are cylindrical structures, each centriole is
composed of nine sets of MT triplets arranged
in the fashion of a pinwheel. In the triplets,
microtubule A is complete (consists of 13
protofibrils), while B and C share 2–3 subunits.
A pair of centrioles is found in non-dividing cell.
Long axes of the centrioles are at right angles
to each other. Before cell division each
centriole duplicates itself.
triplet
ch
centrioles
MT
Junqueira, Carneiro and Kelly, Basic Histology, 1995
MT – microtubules, ch - chromosomes at equatorial plate
MICROTUBULES
Non-branching rigid hollow tubes
Diameter 25 nm, wall 5 nm
Tubulin – heterodimer consisting of the α- and ß- tubulins
Polymerization in circle (13 circularly arrayed tubulin molecules),
longitudinal contacts link tubulin molecules into protofilaments
Dynamic instability - polymerization = elongation, and depolymerization = shortening, depend on the presence of GTP, Mg2+
MTOC - Microtubule organizing center, MT grows from tubulin ring
-(minus) non-growing end, + (plus) growing end
MT associated proteins regulate polymerization and anchor of MT to
organelles
MT are found in: centrioles, mitotic spindle, cilia, flagellum,
elongating cell processes, cytoplasm
MT are involved in: cell elongation and movement, intracellular vesicular
transport, movement of chromosomes, maintenance of cell shape
Motor proteins: ATP-driven MT are associated with proteins which are attached
to the moving structure and pull it along a MT. Dynein moves along MTs toward
their minus end, and kinesin toward plus (growing) end.
Microtubule formation (Ross, Romrell, Kaye, Histology, 1995)
Distribution of the microtubules (MT) in cell
MT
MTOC
nucleus
Microtubule-organizing center (MTOC)
centrosom
Consists of
(a) a pair of
centrioles,
(b) amorphous
protein material,
and (c) gamatubulin rings
(nucleation sites
for microtubule
formation)
NBBC connector is not
in humans
Scheme: Molecular Biology of the Cell, Alberts et al., 2002
Scheme: Histology, Ross, Pawlina, 2011
The molecular motor proteins associated with microtubules (MT)
Kinesins move along the MT toward plus end, transport organelles from the cell
center toward the cell periphery
Dyneins move along the MT toward minus end, transport organelles from the cell
periphery toward the MTOC.
Scheme: Histology, Ross, Pawlina, 2010
ACTIN FILAMENTS, MICROFILAMENTS
G (globular) actin, polymerization in double-stranded helix of the
F (fibrillar) actin
Actin filaments in non-muscle cells (diameter 6-8 nm)
readily dissociate (depolymerization) and
reassemble (Ca2+, ATP - dependent polymerization)
Terminal web (association with cell m., actin-binding proteins)
Structural framework within cell
Core of microvilli and sterocilia (actin-linking proteins)
Anchoring cell junction – zonula adherens
Moving and shifting cytoplasmic components
(motor proteins of AF are myosins, type I and II)
Locomotion of cell (pseudopodial processes)
Contractile ring (constriction of cells during cytokinesis)
In muscle tissue
Actin filaments (6-8 nm) are structurally stable and integrated with
myosin filaments (15-16 nm)
Arrangement of the actin filaments in the microvilli (M) and into the terminal web (Tw)
Scheme: Stevens and Lowe, Histology, 1993
M
Capping proteins
Tw
myosin I
nucleus
Scheme: Molecular Biology of the Cell, Alberts et al., 2002
Actin filaments (MF = microfilament) and MT
intermediate filaments
Electronmicrograph: Junqueira´s Basic Histology, Mescher, 2010
Distribution of the intermediate filaments in cell
Scheme: Molecular Biology of the Cell, Alberts et al., 2002
Tw
D
D
D
H
IMF – blue, D = desmosome, H = hemidesmosome
Electronmicrograph: Junqueira´s Basic Histology, Mescher, 2010
Intermediate filaments (IMF) are ropelike filaments (diameter: 10-12 nm) consisting of the
stable structural proteins. IMF form three-dimensional meshwork, are concentrated
in the terminal web (Tw), connect desmosomes and hemidesmosomes.
Their proteins display tissue specifity: cytokeratins (epithelium), vimentin (cell of
mesenchymal origin), desmin (muscles), glial fibrillar acidic protein (glial cells –
astrocytes), neurofilament proteins (neurons), lamins (A,B,C) – nuclear lamina of the
nucleus. Immunocytechemical detection of the IMF proteins is used in the tumor
diagnosis
Characteristics of the three types of cytoskeletal elements
Actin filament
Diameter:
6 – 8 nm
Composition: Polymer of G-actin
Structure: Double-stranded F-actin helix
Thin flexible filament
Readily dissociate
and reassamble
Enzyme
activity: ATP hydrolitic activity
Location and function
in the cell: Terminal web
Zonula adherens
Core of microvilli
Contractile ring in the
dividing cell
Contractile elements
of muscles
Scheme: Histology, Ross, Pawlina, 2006
Intermediate filament
10 – 12 nm
Various proteins
Ropelike fiber
Strong, stable structure
None
Extend across cytoplasm
connecting desmosomes
and hemidesmosomes
Nuclear lamina of nucleus
Support of cell processes
Provide mechanical strenght
and resistence to shearing
forces
Microtubule
24-25 nm
Dimers of α- and ß-tubulin
Hollow non-branched cylinder
Exhibit dynamic instability
GTP hydrolytic activity
Core of cilia and flagellum
Centriole
Mitotic spindle
Provide network “railroad
tracks“ for movement of
organelles within cell
Movement cilia and
chromosomes (during
cell division)
CELL INCLUSIONS
Accumulation of metabolites or deposits of various nature
Glycogen (hepatocytes, cardiomyocytes)
PAS reaction with control test, Best´s carmine
Lipids (adipocytes, cells producing steroid hormones, sebaceus gland)
Oil red, Sudan dyes
Proteins - Reinke´s crystals in Leydig´s cells
Pigments – colored substances
a) Exogenous: dust, carbon particles (alveolar macrophages)
lipochromes (carotenoids, yellow color of adipose tissue)
b) Endogenous pigments
hematogenic: hemosiderin (iron-containing, brownish pigment)
ferritin (size 8-9 nm, iron protein complex)
melanin - melanosomes (organelles), dark brown pigment
melanocytes, pigment epithelium of retina
lipofuscin – yellowish brown pigment, in static cell population
(cardiomyocyte, neuron), derived from secondary
lysosomes, deposits of indigestible substances
GLYCOGEN in hepatocytes
Dust cells (macrophages filled with carbon
liver, PAS reaction + alum hematoxylin
particles) in the lung
lymphocytes
dust cells
Photomicrographs: Collection of ÚHIEM
artery
Lipid droplets in hepatocytes (oil red, alum hematoxylin)
Lipofuchsin in neurons of the spinal ganglion (HE)
Melanin in pigment epithelium of retina (HE)
Hemosiderin in macrophages (arrow)
melanocyte
Photomicrographs: Collection of ÚHIEM
CELL CYCLE
Morphological and biochemical events occurring in life of a cell
between its formation and end of its division into two daughter cells
Four phases:
G1- cell growth, RNA and protein synthesis, volume of cell is restored
some cells leave cycle to begin differentiation (G0)
S – DNA synthesis = DNA replication, chromosomes consist of two
chromatids, duplication of centrioles
G2- RNA and protein synthesis (tubulin), formation of energy for M
M - mitosis is process that produces two identical daughter cells
with the same chromosome number and DNA content
equal partition of genetic material
Regulation of cell division
Regulation is critical for normal development of multicellular organism
(loss of control ultimately leads to cancer)
Cyclins and cyclin dependent kinases
Proto-oncogenes (normal proliferation and differentiation)
Growth factors (stimulation), chalons (inhibition)
Cell cycle and checkpoints
Checkpoints – internal quality control mechanisms, monitor
and modulate the progression of cell through the cell cycle
prevents premature
entry into anaphase
prevents the cytokinesis until all the
chromosomes have been correctly separated
Non-proliferating
population of cells
terminal
differentiation
prevents progression of
cell into M phase
Irreparable damage
of DNA tumor-supressing
protein p53 –
programmed cell
death (apoptosis)
monitors quality
of replicating DNA
Cell self-evaluates its own replicative potential
before deciding to enter the S phase
Scheme: Ross, Pawlina, Histology, 2003
PHASES OF MITOSIS
Karyokinesis
Prophase - condensation of chromosomes (sister chromatids held
together by centromere)
disappearance of nucleoli, disintegration of nuclear
envelope (prometaphase)
Metaphase - centrioles at poles of cell, formation of mitotic spindle
(astral, polar and kinetochore mictotubules)
chromosomes at equatorial plane (metaphase plate)
Anaphase - separation of sister chromatids (cohesins break
down). Chromatids are pulled to opposite poles of cell
Telophase – reconstruction of nuclear envelope
decondensation of chromosomes, nucleoli reappear
Cytokinesis
Elongation of cell, furrowing of the cell membrane midway between
the poles of mitotic spindle (cleavage furrow). Separation is achieved
by a contractile ring (actin filaments, myosin II molecules)
kinetochere
microtubule
polar microtubule
astral
microtubule
cleavage furrow
contractile ring
MEIOSIS is the cell division that takes place in the process of gametogenesis. Meiosis (M)
includes two successive divisions, the first (M I) and the second (M II) meiotic divisions,
to reduce the number of chromosomes to haploid number.
FIRST MEIOTIC DIVISION
Prophase
Homologous chromosomes (HChs)
approach each other (A). Pairing of
HChs and formation of synaptonemal
complexes (B).
Crossing-over - interchange of
chromatid segments between paired
HChs. Segments of chromatids break
and are exchanged as HChs separate;
points of interchange are temporally
united and form chiasma (C).
Anaphase (E): random distribution of
(paternal and maternal) chromosomes to
the daughter cells gives rise to infinite
genetic diversity.
composed of
two chromatids
1n 2c
1n 1c
Scheme: Sadler, Langman´s Medical Embryology, 2004
Haploid number
of chromosomes
1n
SECOND MEIOTIC DIVISION is not
preceded by an S phase. During
anaphase centromeres of each
chromosome separates and chromatids
migrate to opposite poles of the spindle.
At completion of division, chromosomes
in each of the four daughter cells are
different from each other
CELL DEATH
In humans, as in all other multicellular organisms, the balance between cell
production and cell death must be maintained.
Cell death may result from accidental cell injury (necrosis) or mechanisms that
cause cell to self-destruct (apoptosis, programmed cell death).
Necrosis is a pathological process, is result of an acute cell injury. The
demage of cell membrane leads to the swelling of the cell and membrane
breakdown followed by the release of cytoplasmic content. This process is
often associated with the inflammatory response.
Apoptosis occurs under normal physiological conditions, and is characterized by the DNA fragmentation, decrease in cell volume, loss of the mitochondrial function, membrane blebbing, and formation of the apoptotic bodies.
The apoptotic bodies are removed without a trace by phagocytotic cells
that no inflammatory response is elicited.
Apoptosis can be activated by the external stimuli – tumor necrosis factor,
transforming growth factor ß, free radicals, oxidants, toxins, UV and ionizing
radiation. Internal activators are oncogenes, tumor supressors (p53).