Transcript 3. Regulation of cell type during mitosis

BIO 121 – Molecular Cell Biology
Lecture Section 5
A. Review of the mitotic cell cycle and cell
death
B. Regulation of cell number and quality
during mitosis
C. Regulation of cell type during mitosis
D. Wound Healing
Control of Cell Number
A. Review of the mitotic cell cycle and cell death
B. Regulation of cell number and quality during
mitosis
C. Regulation of cell type during mitosis
Numbers in a Cell Population
• Cell number is a combination of....
• Cell divisions – Cell deaths (necrotic + programmed)
• Necrosis is premature cell death
– disease, injury, starvation, toxicity, excitotoxicity
• Programmed cell death is death by design
– apoptosis, anoikis, cornification, autophagy
• Same for an organism, system, organ or tissue,
and for single cell populations in an ecosystem
Programmed cell death is an essential component
It guarantees appropriate ennervation patterns
Figure 18-13 Molecular Biology of the Cell (© Garland Science 2008)
Even gives us our fingers and toes!
Figure Q18-1 Molecular Biology of the Cell (© Garland Science 2008)
Overview of the Mitotic Cell Cycle and Cell Division.
Diploid cells duplicate the contents of their cytosol and
nucleus prior to splitting to form two genetically exact
daughter cells.
Figure 17-51a Molecular Biology of the Cell (© Garland Science 2008)
We’ve learned to both control it.........
A mutation in a signal molecule that limits muscle cell division has been bred in.
Figure 17-69 Molecular Biology of the Cell (© Garland Science 2008)
Fig. 11-19
And fear it.........
2 µm
A normal cell next to a tumor derived from uncontrolled cell divisions
Like everything else, the process of
cell division has evolved over time
Fig. 12-11-1
Origin of
replication
E. coli cell
Prokaryotic Division
is called binary fission.
Two copies of circular
DNA, beginning at the
origin of replication,
actively move apart from
each other.
Two copies
of origin
Cell wall
Plasma
membrane
Bacterial
chromosome
Fig. 12-11-2
Origin of
replication
E. coli cell
Two copies
of origin
Origin
Cell wall
Plasma
membrane
Bacterial
chromosome
Origin
Fig. 12-11-4
Origin of
replication
E. coli cell
Two copies
of origin
Origin
When the daughter
chromosomes reach
the opposite poles,
the cell separates into
two daughter cells
Cell wall
Plasma
membrane
Bacterial
chromosome
Origin
Fig. 12-12
Since prokaryotes
evolved before
eukaryotes, mitosis
probably evolved from
binary fission.
Certain protists exhibit
types of cell division
that seem intermediate
between binary fission
and mitosis.
Bacterial
chromosome
(a) Bacteria
Chromosomes
Microtubules
Intact nuclear
envelope
(b) Dinoflagellates
Kinetochore
microtubule
Intact nuclear
envelope
(c) Diatoms and yeasts
Kinetochore
microtubule
Fragments of
nuclear envelope
(d) Most eukaryotes
The complex mitotic cell cycle is
used by the organism to control:
cell number
cell quality
cell type
Target
Mechanisms
Cell
Type
Cell
Quality
Figure 15-8 Molecular Biology of the Cell (© Garland Science 2008)
Cell
Number
We’ve even
looked at one
pathway already
Figure 17-62 (part 1 of 3) Molecular Biology of the Cell (© Garland Science 2008)
Fig. 12-5
G1
S
(DNA synthesis)
G2
Interphase does the real work of mitosis
• Gap 1 (G1) is when the cell gets ready for DNA synthesis
– Need DNA synthase complexes, repair enzymes, histones, etc..
• S-phase is when the cell synthesizes and edits chromosomes
– Synthesis and error editing are the primary objectives.
• Gap 2 (G2) is when the cell gets ready for cell division
– Need organelles, cytoskeletal proteins, molecular motors,
metabolic enzymes, etc.
– Kinetochores are refined and finalized.
Fig. 12-UN1
G1
S
Cytokinesis
Mitosis
G2
MITOTIC (M) PHASE
Prophase
Telophase and
Cytokinesis
Prometaphase
Anaphase
Metaphase
M-Phase
• Prophase: chromosomes condense with kinetichores,
centrisomes move to poles, nuclear membrane disintegrates
• Metaphase: spindle fibers attach and push to midline
• Anaphase: kinetichores pull sister chromatids to poles
• Telophase: the reversal of prophase activities
• Cytokinesis: actin-based separation of cytosol into daughters
Sister chromatids are attached to each other at centromeres and
each is individually attached to a kinetochore from the opposite pole
metaphase
Figure 17-43a Molecular Biology of the Cell (© Garland Science 2008)
Sister chromatids separate from each other, producing 92 individual
chromosomes in humans, and are pulled by kinetichores to the poles.
anaphase
Figure 17-43b Molecular Biology of the Cell (© Garland Science 2008)
dynein motors
Figure 17-37 Molecular Biology of the Cell (© Garland Science 2008)
Figure 17-40 Molecular Biology of the Cell (© Garland Science 2008)
Cytokinesis results from the assembly of an actin-myosin ring that gets smaller
and smaller as mysoin pulls actin along actin. Ring assembly is microtubuledependent and myosin light chain kinase must be phosphorylated to begin.
Figure 17-49a Molecular Biology of the Cell (© Garland Science 2008)
The cleavage furrow
Cell Division in Plants
In some plants growth continues over the life of the organism
Regulation of the Cell Cycle
• There are four major ‘checkpoints’ that monitor sensor
systems and trigger molecular switches to get to next stage
–
–
–
–
The G1/S checkpoint – to enter the cycle or not
The S-checkpoint – to synthesize DNA or not
The G2/M checkpoint – to divide the cell or not
The M-checkpoint – to shift from metaphase to anaphase
• The sensor systems are focused outside the cell as well as
inside the cell to make sure conditions are appropriate
• When ‘off’, the molecular switches halt progression, when
‘on’ they biochemically start the next stage
Fig. 12-14
G1 checkpoint
Control
system
S
G1
M
M checkpoint
G2 checkpoint
S checkpoint
G2
The book claims 3
and describes 4,
so we’ll go with 4
• For many cells, the G1 checkpoint seems to be the most
important one
• If a cell receives a go-ahead signal at the G1 checkpoint,
it will usually complete the S, G2, and M phases and
divide
• If the cell does not receive the go-ahead signal, it will
exit the cycle, switching into a nondividing state called
the G0 phase
Copyright © 2008 Pearson Education, Inc., publishing as Pearson Benjamin Cummings
Fig. 12-15
G0
G1 checkpoint
G1
(a) Cell receives a go-ahead
signal
G1
(b) Cell does not receive a
go-ahead signal
Stop and Go Signals at the G1 Checkpoint
• In multicellular organisms, the classic required external signals
are growth factors, cytokines and hormones
• Most cells also exhibit anchorage dependence, in which they
must be attached in order to divide
• Most cells also exhibit density-dependent inhibition, in which
crowded cells stop dividing
• Many single cells also monitor external nutrient availability and
will not divide if it is inadequate
Copyright © 2008 Pearson Education, Inc., publishing as Pearson Benjamin Cummings
The Cell Cycle Switches at the First Three Checkpoints are
Molecules Called Cyclins and Cyclin-Dependent Kinases
Cyclins are expressed and degraded, when present the cell moves forward
1. G1 and G1/S cyclins regulate the G1 checkpoint
2. S cyclins regulate S checkpoint
3. M cyclins regulate G2/M checkpoint
Cyclin-dependent kinases are always present and are
activated by the binding of their appropriate cyclin
Figure 17-15 Molecular Biology of the Cell (© Garland Science 2008)
1. Cdk is always there, 2. if conditions are OK cyclin is expressed,
3. the combination is an active kinase, and 4. activity is lost with cyclin
Cdk
Cyclin accumulation
Cyclin is
degraded
G2
checkpoint
MPF
Cyclin
What does cyclin-cdk phosphorylate?
• G1-cdk
– Rb protein: loses inhibition of E2F
– E2F then activates transcription of S-cyclin
• M-cdk
– histones: chromosome condensation
– lamins: nuclear membrane fragmentation
– myosin: purse-string cytokinesis
So, back to our growth
factor example.....
What does immediate
early gene expression
cause?
Figure 17-62 (part 1 of 3) Molecular Biology of the Cell (© Garland Science 2008)
Immediate early gene, Myc, expression
activates the expression of G1 cyclin
Figure 17-62 (part 2 of 3) Molecular Biology of the Cell (© Garland Science 2008)
M Checkpoint Regulation
• If any kinetochores are not attached to spindle
microtubules during metaphase they send a
molecular signal that delays cycle progression
• The signal activates the anaphase-promoting
complex, or cyclosome, known as APC/C
• APC/C is a protease that destroys:
– cyclins: stopping cell cycle
– securin: which drives sister chromatid separation
Apoptosis During Mitosis Controls Cell Quality
• Every cell cycle in multicellular organisms are
molecularly hardwired with apoptosis as a
potential outcome if things aren’t just right
• The primary cause is imperfect DNA copying
• This is one of our primary defenses against
erroneous or uncontrolled proliferation
Characteristics:
1. Cessation of DNA repair mechanisms
2. Cell shrinkage
3. Nuclear membrane blebbing
4. DNA fragmentation
5. Death
Can result from bad copies
during DNA replication
Mdm2 will cause
p%#
p53 degradation
Figure 17-63 (part 1 of 2) Molecular Biology of the Cell (© Garland Science 2008)
p21 has been shown to
block cyclin/cdk activity at
G1-S, S and G2-M
Figure 17-63 (part 2 of 2) Molecular Biology of the Cell (© Garland Science 2008)
The Roles of p21 and PCNA
• The apoptotic clock:
– PCNA = the proliferating cell nuclear antigen
– If [p21] rises to 5X [PCNA] apoptosis will happen
– p21 will reach binding affinity for PCNA
– The dimer will shut off DNA repair mechanisms
When apoptosis is inactive, Bcl2 is bound to APAF-1 and the
dimer acts to block assemblage of mitochondrial membrane
channel proteins, BH123 (Bax and Bac) ,
Figure 18-11a Molecular Biology of the Cell (© Garland Science 2008)
The loss of DNA repair mechanisms:
1. Activates BH3-only proteins
2. Signal Bax translocation to the mitochondria
BH3-only inactivates Bcl2/APAF-1 inhibition and along
with Bax dimerization with Bac forms active channels
Bax/Bac dimers
apoptosis
inducing
factor
Figure 18-11b Molecular Biology of the Cell (© Garland Science 2008)
Release of cytochrome C from the mitochondria outcompetes
Bcl2 for APAf-1 and activates assembly of the apoptosome
Figure 18-8 Molecular Biology of the Cell (© Garland Science 2008)
(8, 9, 10)
(3, 6, 7)
Figure 18-5b Molecular Biology of the Cell (© Garland Science 2008)
Characteristics:
1. Cessation of DNA repair mechanisms
2. Cell shrinkage
3. Nuclear membrane blebbing
4. DNA fragmentation
5. Death
Extrinsic apoptotic cascade also uses the common caspase cascade
Figure 18-6 Molecular Biology of the Cell (© Garland Science 2008)
3. Regulation of cell type during mitosis
• Review
– Symmetric vs. Asymmetric Cell Division
– Stem Cell and Embryonic Cell Divisions
• Regulation
– Control in the Cytosol
– Control in the Nucleus
Common Cell Divisions Produce Daughters
Like the Parent to Replenish the Population
Common during growth and the repair of damaged tissues
The Stem Cell Concept
• Asymmetric division of stem cells produces one new
stem cell and one differentiated daughter
– Many stem cells in the embryo and adult
• In some organs: frequent replenishing divisions
– gut, epidermis, bone marrow
– example: billions of blood cells are destroyed by the spleen
every hour
• In others, they only divide in response to stress or
the need to repair the organ
– heart, prostate
Stem Cell Mitosis
HSC
Each division produces daughter
cell(s) unlike the parent cell:
- The first two are asymmetric
and produce one stem cell
- The last one produces two
like daughters unlike parent
Stem Cell Types: Embryonic Stem Cells
The Inner Cell Mass
Produces all cells
of the embryo
Stem Cell Types: Adult Stem Cells
• Committed stem cells with limited potential
– hematopietic stem cells
- hair stem cells
– mesenchymal stem cells
- melanocyte stem cells
– epidermal stem cells
- muscle stem cells
– neural stem cells
- tooth stem cells
– gut stem cells
- germline stem cells
– mammary stem cells
The hair shaft is composed of keratinocytes, lubricated
by sebaceous secretions, with melanin for color.
- “Bulge” is the stem cell niche for hair basal cells,
sebocytes and melanocytes.
- The first two arise from a common stem cell but
melanocytes arise from a committed stem cell.
Hematopoietic stem cells in the bone marrow
are the kings of differentiation choices
Progenitor cells will produce two daughters like each
other but not like the parent, with the choice being
determined by the current, local signaling combination
Control of Assymetrical Division in the Cytosol
• G2 is the key to differentiation events
• The daughter cells can be built differently
–
–
–
–
DNA is the same but....
RNA’s can be different
Proteins can be different
Cytoskeleton and organelles can even be different
• The parent cell must focus its placement of these
components on either side of the furrow
Isolation of transcription factors across midline
Fig. 18-15a
Unfertilized egg cell
The unfertilized
egg is the queen Sperm
of cytoplasmic
isolation of cell
Fertilization
fate determinants
Nucleus
Two different
cytoplasmic
determinants
Zygote
Mitotic
cell division
Two-celled
embryos
have two
different
cell types
All stem cells
do it effectively
We use the embryonic
process of cleavage to
disperse the careful
distribution of cytoplasmic
determinants laid down in
the egg.
The G-phases of somatic mitosis allow for cytoplasmic growth
so that the daughter cells are equal in size to the parent cell.
In cleavage we want to use the egg cytoplasmic material so we
just skip the G-phases all together.
It makes the cell cycle go very fast!
- Frogs can make 37,000 cells in 43 hours.
- Fruit flies can make 50,000 in 12 hours (10 min!)
They don’t even bother to make plasmamembranes until later!
Control of asymmetric division in the nucleus
Two DNA methyltransferases are
important in modifying DNA
Development of pluripotency in the inner cell
mass depends on methylation pattern
Both parent gametes have methylation patterns that must be
removed for all genes to be available to the developing organism.
One of the big hurdles to somatic cell nuclear transfer
cloning was overcoming the adult methylation pattern
It took hundreds of failed
attempts before the
successful cloning of Dolly
from adult mammary
epithelium.
Each step of differentiation of a given cell type
depends on changing methylation patterns
These genes maximize effectiveness when coding for
transcription factors, SNuRPs, signaling receptors, etc.
Cutaneous Wound Healing
The skin is a complex organ...
Many cells and activities involved
Many cells and activities involved in Healing
Clotting
Scarring
Re-establishing
Function
• Four overlapping stages to wound healing
– Hemostasis
– Inflammation
– Proliferation
– Maturation
Blood flows into the exposed ECM of
the injured tissue.
RBC and Platelets Trapped in Fibrin Clot
Clotting factor
VII from the
blood contacts
tissue factor on
cells in the
damaged
tissues to
activate clotting
Platelet activation in the. clot makes them sticky
and releases their signal storage vesicles
©2000 by Lippincott Williams & Wilkins
Camacho A , Dimsdale J E Psychosom Med 2000;62:326-336
Positive feedback
activates even more
Platelet activation
releases growth
factors by regulated
secretion
Inflammation is a process mediated primarily
by WBC as part of our innate immunity
- Resident mast cells and macrophages
- Recruited monocytes and neutrophils
Resident mast cells also degranulate
rubor = redness
calor = heat
tumor = swelling
dolor = pain
Activated mast cell activities
Figure 1 Development and differentiation of macrophages.
Rickard A J , Young M J J Mol Endocrinol 2009;42:449-459
© 2011 Society for Endocrinology
Activated macrophage activities
The special case of extravasation
• Circulating WBC must get out of the vessel
• Combines activation of the WBC with
‘Cell Rolling’, ‘Adhesion’ and ‘Diapedesis’
1.
The presence of environmental cues associated with injury
and infection change endothelial surface selectins
2.
These catch closely matched WBC surface oligosaccharides
and make them roll to a stop on endothelial surface
3.
The white blood cell then activates an integrin that binds
tightly to ICAM on endothelial cells
4.
Diapedesis uses basic migratory mechanisms along with WBC
shape change to squeeze between endothelium
Intercellular Diapedesis
Transcellular
Diapedesis
Activated neutrophils are phagocytic
Pseudopod-Driven Phagocytosis in Neutrophils
Figure 13-46 Molecular Biology of the Cell (© Garland Science 2008)
Actin polymerization
is required to extend
pseudopods
Actin depolymerization is
required to seal off the
phagosome
Figure 13-47a Molecular Biology of the Cell (© Garland Science 2008)
Proliferation re-establishes tissue function
• Reconnection of
the dermal
connective tissue
• Integrity of the
epidermal layers
• Re-establishment
of blood flow
Reconnection of
the dermal CT
Cell Migration or “Crawling”
• The Basic Mechanism
–
–
–
–
Triggered by signals from outside the cell
Actin-myosin based movement
Requires attachments to outside to pull against
Gotta’ drag all of the cell contents along for the ride
Chemotaxis
Circumferential receptors
Rho-family GTPases (monomeric)
Rho-dependent kinases
1. Actin monomer nucleotide exchange
2. Actin fiber polymerization and disassembly
3. Myosin motor ATPase activity
Figure 17-62 (part 1 of 3) Molecular Biology of the Cell (© Garland Science 2008)
Formation of the scar matrix
1.
2.
3.
4.
glycosaminoglycans
proteoglycans
fibrous proteins
elastic proteins
Re-establishment of the
epidermal epithelium
involves both mitosis and
epithelial migration
Also must reform the basal lamina
Re-epithelialization below the scab
scar
Fi
Model depicting α3β1-integrin-mediated functions of epidermis that contribute to wound
healing.
Mitchell K et al. J Cell Sci 2009;122:1778-1787
©2009 by The Company of Biologists Ltd
Figure 23-34 Molecular Biology of the Cell (© Garland Science 2008)
Maturation Phase
Wound contraction by myofibroblasts
Stitches Perform Wound Contracture
Collagen Remodeling
A scar never reaches the strength of
undamaged tissue
Healing Abnormalities
• Failure to heal: Excessive Inflammation
• Excessive scarring: Wound Fibrosis
– Hypertrophic Scarring
– Keloid Scarring
Biofilms May Block Healing
Hypertrophic scars result from failed
fibroblast contracture
Don’t extend beyond the original wound edge
Keloid scars result from excessive
TGF-b receptors on fibroblasts
Extend to fibroblasts outside the wound
People have
exploited these
conditions to create
the ‘keloid tattoo’