Non-membranous cell organelles.
Cytoskeleton – structure, function
and tissue specificity.
Specialization of the cell surface.
Intercellular junctions.
Biological motors – molecular principles
Institute of Histology and Embryology
Author: Prof. MUDr. Jindřich Martínek, DrSc.
Subject: General Histology and general embryology
Code: 82241
th
th
Date: 2013, October, 10 and 12
NON-MEMBRANOUS CELL ORGANELLS
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NUCLEOLUS
RIBOSOME
CENTRIOLE
CYTOSKELETON
thin filaments - actin
intermediate filament
lamins (A, B, C)
cytokeratins (1 – 20)
desmin, vimentin
nestin
microtubules
Becker: The world of the cell, 1986
NON-MEMBRANOUS CELL ORGANELLES
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NUCLEOLUS – localization in the karyoplasm – nucleolar organizing centers (NORs) two components (amorphous and granulous) – site of transcription of r-RNAs (ribosomal)
– compact, network and ring-form type
CENTRIOLE – specific organization of microtubeles (9 triplets) – MTOC – microtubular
organizing center function, simple and multiple centriolar replication. Role in formation of
mitotic and meiotic spindle and basal bodies of cilia and flagella
CYTOSKELETON – complex of thin (5-7 nm) and intermediate (10-15 nm) filaments
and microtubules (24 nm) – actin (thin) filaments with myosin and tubulins (of
microtubules) with dynein and kinesin represent molecular motors for intracellular
traffic – intermediate filaments are nonpolar – nestin, cytokeratins, vimentin, desmin –
tissue specific – cells of epithelial origin (cytokeratin), mesenchymal origin (desmin),
muscle tissue (vimentin)
RIBOSOMES – (15-30 nm) composed of two subunits (small and large) – play a role in
translation – protein synthesis – as free polysomes and membrane bounded (GER)
NUCLELUS – non-membranous cell organelle derived from activated parts of some
chromosomes – nucleolar organizing regions (NORs) or nucleolar organizing centers (NOCs)
Types: compact – initial transcription of rRNA at the single NORs; network – massive transcription in gradually confluent NORs
ring-form – decreasing transcription at the single NOR
FC = fibrilar center – NOC
PF = pars fibrosa – accumulated transcript
PG = pars granulosa – ribosomal assembly
PNC = perinucleolar chromatin
PNC
PF
PG
FC
PNC
VISUALISATION OF NUCLEOLAR ORGANISERS USING
IMMUNOHISTOCHEMICAL PROBES
1
2
3
4
2
3
1
TYPES OF NUCLEOLI:
a) compact – currently activated nucleolar
organiser
b) with nucleolonema – massive
formation of ribonucleoproteins around NORs
- typical network appearance
c) ring form – decreased of RNA transcription in
the inactivated nucleolar organiser
1 = perinucleolar chromatin
3 = nucleolonemata
5 = lamelae annulatae
2 = nucleolar organiser
4 = nuclear channel system
6 = lysosomes
5
6
RIBOSOMES (15 – 30 nm)
free polysomes
GER
PROTEIN SYNTHESIS
NUCLEUS
NUCLEAR
PORES
GER
GER
RIBOSOME – TRANSLATION MECHANISM – POLYSOMES
Start codon (AUG)
Currently translating codon (arbitrary)
Scheme of the protein synthesis on free
polysomes (Junqueira´s Basic Histology, Mesher, 2010)
ribosome
GER
GER
mRNA
free protein
in the cytoplasm
Electron micrograph: Histology, Ross, Pawlina 2010
Arrows: free polysomes
Rough (Granular) Endoplasmic Reticulum rER (GER)
GER
IMMUNOHISTOCHEMICAL DETECTION OF SOME CYTOSKELATAL COMPONENTS
Microtubules (red), actin filaments (green)– fluorescence microscope
CHARACTERISTICS OF CYTOSKELETAL COMPONENETS
Ross, Pawlina: Histology, 2006
Actin filaments
Diameter:
6 – 8 nm
Composition:
Polymer of G-actin
Intermediate filaments
Structure: Double-stranded F-actin helix
Thin flexible filament
Readily dissociate
and reassamble
Enzyme
activity: ATP hydrolitic activity
ATP-dependent polymeration
Location and function
in the cell:
Terminal web
Zonula adherens
Core of microvilli
Contractile ring in the
dividing cell
Contractile elements
of muscles
Microtubules
10 – 12 nm
Various proteins
Ropelike fiber
Strong, stable structure
24 nm
Dimers of α- and ß-tubulin
Hollow non-branched cylinder
Exhibit dynamic instability
None
Extend across cytoplasm
connecting desmosomes
and hemidesmosomes
Nuclear lamina of nucleus
Support of cell processes
Provide mechanical strenght
and resistence to shearing
forces
GTP hydrolytic activity
GTP-dependent polymeration
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
ACTIN
thin filaments (5 – 7 nm)
G and F actin – ATP dependent
(poly- and depolamerization)
MOLECULAR MOTOR COMPONENT
together with myosin
Smooth muscle cells
Hearth muscle cells - cardiomyocytes
FUNCTION OF MICROFILAMENTS
AND MICROTUBULES IN REGULATIONS
AND INTRACELLULAR TRAFIC
THE MOLECULAR MOTOR PROTEINS WORKING WITH MICROFILAMENTS (ACTIN)
Unipolarly organized myosin
(myosin monomers – myosin I) –
one way movement of cargo
Bipolar organization of myosin
(myosin monomers – myosin II) –
ontractile activity with opposite
direction movement of actin
filaments – muscle tissue
Unipolarly working myosin (I) –
attached to the cytoplasmic
membrane – formation of pseudopodia movement of cell
THE MOLECULAR MOTOR PROTEINS WORKING WITH TUBULINS OF MICROTUBULES
Kinesins move along the MT tothe plus end and can transport cargo (organelles) from the cytocentrum
toward the cell periphery
Dyneins move along the MT to the minus end, transport cargo (endocytotic vesicles) from the cell
Scheme: Histology, Ross, Pawlina, 2010
periphery toward the MTOC.
INTERMEDIATE FILAMENTS – non-polar
lamins A, B, C – nuclear lamina
cytokeratins – cells of germ layer origin
desmin – cells of mesenchymal origin
vimentin – cells of mesenchymal origin neurofilaments –
nerve cells
GFAP – glial fibrillary associated protein (acidic)
nestin
– during cell development
CYTOKERATIN 18 – fission products – APOPTOTIC MARKER
MICROTUBULES (25 nm)
tubulin α, β, γ
(GTP – dependent polymerization)
MOLECULAR MOTOR COMPONENT –
together with dynein (+ -) and kinesin (- +)
CENTRIOLAR PAIR - CENTROSOME
CILIUM – KINOCILIUM
Ross, Pawlina: Histology, 2006
CENTRIOLES AND CENTRIOLAR REPLICATION
Centriolar pair = diplosome – centrosome
Centriole:
Procentriole:
 = 200 nm
 = 200 nm
L = 400 nm
L = 200 – 400 nm
Centriolar DNA – MTOC – centriolar precursors
SIMPLE REPLICATION – begins from
existing centriolar pair
MULTIPLE REPLICATION – starts as synthesis of precursor material (tubulins)
and continues as induced assembly of nine procentriolar triplets of microtubules –
typical for cells with kinociliary apparatus at the apical surface
ZONULA OCCLUDENS – tight junctions – missing intercelluar space
Mucus layer
ZONULA OCCLUDENS – freeze fracturing technique
Ridges
Grooves
Ciliary
necklaces
Ross, Pawlina: Histology, 2006
Golgi complex
Zonula occludens
Zonula adherens
Desmosome – macula adherens
Cytokeratin –
intermediate filaments
ZONULA ADHERENS – terminal web –
transverally arranged actin filaments inserting via α-actinin
and vinculin and catenin into E-cadherin moleculs
GAP JUNCTIONS – NEXUSSES –
communicative cell to
cell interconnections
connexon – canal formation
connexins – membrane proteins
MICROVILLI
 = 0.1 m
lenght = 1 – 5 m – core – actin filaments
Regularly arranged and numerous microvilli
represent specific surface specialization of an
absorptive epithelium – brush border –
at the apical pole of cells.
Stereocilium – long and branched microvillus
Ross, Pawlina: Histology, 2006
Microvilli at the cell suface and intercellular interconnection
ZO = zonula occludens; ZA = zonula adherens; D = desmosome
zo
ZA
D
SOLITARY (single or individual) CILIUM
 = 0.25 m
L = 3 – 5 (7) m (50 for tail of spermatozoon) Consist of basal body (9 triplets – MTOC) and axonemal complex (9 doublets
and 1 central pair of microtubules).
Basal body develops from one of
centrilar pair and therefore it is located often in the CYTOCENTRUM.
Pinocytotic vesicle
Ciliary sheat
Basal body
Axonemal complex
Ross, Pawlina: Histology, 2006
KINOCILIARY APPARATUS
Oviduct – simple columnar
ciliated epithelium
Cilia and microvilli
at the cross section
CILIA, MICROVILLI, JUNCTIONAL COMPLEXES
AND TERMINAL WEB
INTERDIGITATIONS
IMMUNOFLUORESCENT DETECTION
OF PANCADHERIN IN SEMINIFEROUS
TUBULES OF TESTIS (Zonulae adherentes)
BETWEEN SERTOLI CELLS PROCESSES
BASOLATERAL LABYRINT
Typical for absorptive type of
epithelial cells as in the
proximal tubules (kidney)
or in lining of so called
striated ducts in some
salivary glands.
Specific for an intesive
Ion transport (energy
needed) by e.g.
Na+, K+ ATP-ase as
an integral protein of the plasma membrane
Stevens, Lowe: Histology, 1993
Ross, Pawlina: Histology, 2006
Stevens, Lowe: Histology, 1993