Mechanisms of Disease I : Cell growth + differentiation Flashcards
Define cell growth
An increase in cell size and an increase in number of cells.
Define cell differentiation
When cells become more complex and specialised , usually also when cells stop growing
What is cell potency. ?
a cells ability to differentiate into other cell types. the more cell types a cell can differentiate into the more potent.
what are the three main groups diseases related to cell growth and differentiation can fall in to?
Developmental conditions - can be related to cell growth or differentiation
Neoplasia (and Metaplasia)
Other e.g. cardiac hypertrophy
Give an example of a developmental condition
Neural tube defects such as spina bifida (complete closure of the spine)
What is neoplasia ?
it is the uncontrolled growth of cells. the replacement of one differentiated cell type with another mature differentiated cell type that isnt normally present
e.g. Cancer, tumours
What is metaplasia ?
Transformation of one cell type into another e.g. cancer
What are the two main forms of cell growth ?
Hypertrophy
Hyperplasia
Define hyperplasia
More cells caused by cell division and increased cell proliferation. Hyperplasia is the most common form of cell growth.
Define hypertrophy
Cells growing bigger. This is caused by cells making more proteins, macromolecules and membranes. Elevated protein synthesis is a big driver of increased cell size.
What are differentiated cells called ?
post-mitotic. Cell morphology and function will have changed. They will show specific cell type-specific genes
Similarities between cell growth + differentiation
both governed by integration of multiple signals :
intra and extracellular signals
growth and inhibitory factors
cell adhesion etc
What are co-incidence detectors ?
Promotors. signals converge on promotors of key genes.
Define extracellular signals
Ligand binding to a receptor that induces intracellular cascade
What are the three broad classes of extracellular signals
Paracrine
Autocrine
Endocrine
Describe paracrine signals
produced locally to stimulate proliferation of a different cell type that has the appropriate cells surface receptor (cell to cell)
Describe autocrine signals
produced by a cell that also expresses the appropriate cell surface receptor (hormone signally but binds on same cells)
Describe endocrine signals
released systematically for distance effects (hormone signalling)
What is the role of extracellular signals in cell growth + differentiation ?
Proteins that stimulate proliferation and promote survival Mitogens e.g. growth factors and interleukins
Induce differentiation and inhibit proliferation e..g TGF
induce apoptosis e.g. TNF-alpha and other members of TNF family
How do extracellular signals induce gene expression ?
Growth factor binds to growth factor receptor
this will activate signal transduction via a kinase cascade
this results in transcription factor activation in the nucleus
Transcription factors drive transcription of the downstream genes
mRNA created which is exported into the cytoplasm
Translation + protein synthesis take place
Proteins are formed which remain in cytoplasm exerting
their functions
4 phases of the cell cycle
M, G1, S, G2
G0
Describe M phase
Mitotic phase, where the cell divides giving us two daughter cells
Describe interphase
G1,S,G2. phases of the cell cycle that is not mitosis
Describe G1 phase
One of the gap phases.
The cell increases in size and the cellular contents are duplicated.
Describe G2 phase
The other gap phase.
Cell grows in size. organelles and proteins develop in prep for cell division
Describe S phase
Synthesis phase. where the genome / chromosomes are replicated
Describe G0 phase
These are the Quiescent cells (cells that have left the cell cycle)
These cells may be permanently differentiated so change shape and have a diff function. They ultimately will undergo apoptosis / cell shedding (death)
OR
they can re join the cell cycle at G1
OR
they can permanently remain in G0
When are cells diploid or tetraploid ?
How can we use chromosome numbers ?
After mitosis cells are diploid (2N)
after S phase cells are tetraploid (4N) since everything is replicated.
we can check the number of chromosomes and use this to measure cell proliferation
Describe FACS
FACS = fluorescence and flow cytometry.
It’s a fluorescent activating cell sorting method. Method of flow cytometry.
If DNA stain is applied, FACS can measure (strength of stain which corresponds to) DNA content of every cell in a population.
Difference in a FACS graph between high and low proliferation
In highly proliferative cells where the rate of division is HIGH, the percentage of cells in G1 is lower whilst those in S have increased. G2/M remain similar. a higher S phase % indicated a larger number of mitotic cycles.
Fluorescence microscopy
blue = DNA red = gamma tubulin (involved in anchoring microtubule complexes) green = CHEK2 (mitotic protein / tumour suppressor gene) yellow = centrioles (means that gamma tubulin and CHEK2 are at same place (colocalised) )
define ploidy
no. of chromosome sets in a cell
what are stages in mitosis
PMAT. prophase metaphase anaphase telophase
Prophase
Nucleus becomes less definite
micro-tubular spindle apparatus assembles
centrioles migrate to poles
PROMETAPHASE
nuclear membrane breaks down
kinetochores attach to spindle in nuclear region
Metaphase
chromosomes align in equatorial plane
Anaphase
Chromatids separate and migrate to opposite poles
telophase
daughter nuclei form
Cytokinesis
division of cytoplasm
chromosomes decondense
why do we have cell cycle checkpoints ?
these controls (involving specific protein kinases and phosphatases) ensure the strict alteration of mitosis and DNA replication
Where do we have control checkpoints and their purpose
Restriction point near G1
Checks DNA is not damaged, cell size, metabolite and nutrient stores.
G2 / M phase checkpoint
Ensure S phase is complete
Check DNA completely replictated
Check DNA not damaged
when chromosomes aligned on spindle
when mitotic spindle separates correct no. of each chromosome goes to each cell
Cyclin dependent kinases (CDK) and Cyclins
10 genes encode CDK
> 20 genes encode cyclins
expression of cyclin is induced by growth factors
How do CDKs work ?
Cyclins expressed as a result of growth factors inducing it
when there is sufficient amount of cyclin it forms a complex with CDK
this forms an active kinase complex
This phosphorylates specific substrates
how is CDK regulated ?
cycles of synthesis (gene expression) and destruction (by proteasome)
post translational modification by phosphorylation , may result in activation, modification or destruction
dephosphorylation
binding of cyclin-dependent kinase inhibitors (CDKIs
how do CDKIs work ?
they are able to bind to complexes and inhibit them
How does the Retinoblastoma protein work ?
Retinoblastoma protein (RB) is a key substrate of G1 and G1/S cyclin-dependent kinases
Unphosphorylated RB binds to E2F (transcription factor) preventing its stimulation of S phase protein expression (prevents E2F from binding to promotor of genes)
when RB is in the presence of cyclin D-CDK4 and Cyclin E-CDK2, its phosphorylated thus meaning it dissociates from E2F
E2F is no longer suppressed. it is able to bind to promotor of Cyclin E allowing replication to start.
what do mitogens do ?
act in G1 phase which will signal to the nucleus and through transcription will cause expression of “early genes” which include transcription factor and will cause expression of delayed genes
delayed genes contain cyclin D which can form active complexes with CDK4/6
cyclin + CDK will cause hypo phosphorylate to the RB. some expression of E2F which will increase cyclin E expression. cyclin E-CDK causes second level phosphorylation of RB which loses all repressive ability
E2F remains high
this will then activate E2F responsive genes required for S phase
Cyclin E-CDK2 activated
what are the sequence of events triggered by mitogens
Growth factor signalling activates early gene expression (transcription factors – FOS, JUN, MYC)•
Early gene products stimulate delayed gene expression (includes Cyclin D, CDK2/4 and E2F transcription factors)•
E2F sequestered by binding to unphosphorylated retinoblastoma protein (RB)•
G1 cyclin-CDK complexes hypophosphorylate RB and then G1/S cyclin-CDK complexes hyperphosphorylate RB releasing E2F•
E2F stimulates expression of more Cyclin E and S-phase proteins (e.g. DNA polymerase, thymidine kinase, Proliferating Cell Nuclear Antigen etc.)•
S-phase cyclin-CDK and G2/M cyclin-CDK complexes build up in inactive forms. These switches are activated by post-translational modification or removal of inhibitors, driving the cell through S-phase and mitosis.
What happens if there is DNA damage
This triggers cell cycle arrest or apoptosis
- stop the cycle via cyclin dependent kinase inhibitors e.g. CHEK2
- attempt DNA repair via nucleotide or base excision enzymes, mismatch repair etc.
- programmed cell death IF repair is impossible. (BCL2 family which blocks apoptosis of some cells e.g. lymphocytes. Caspases which are protease enzymes essential in apoptosis)
the roles of TP53
TP53 = tumour protein 53.
- if DNA is fine then TP53 destroyed by proteasome
-if DNA damaged then mutagen bind. mutagen is detected an causes the activation of protein kinases resulting in TP53 phosphorylation. TP53 can no longer be destroyed by a proteasome.
TP53 can cause expression of CDKI resulting in cell cycle arrest
Activates DNA repair = repair
if it cannot be fixed causes cell death
TP53 and cancer
TP53 loss of function mutations are amongst the most frequent in cancer
- this prevents cell cycle arrest, preventing apoptosis and preventing DNA repair
- this results in more mutations, more heterogeneity (heterogenous tumour mass stops it from adapting), more adapting = cancer progression.
How is chemotherapy involved in cell cycle ?
Traditional chemotherapeutic drugs act on the cell cycle
- stop proliferation and induce apoptosis
S phase drugs cause DNA damage
e.g. 5-fluorouracil (prevents synthesis of thymidine)
Cisplatin binds to DNA causing damage and blocking repair
Discuss M phase drugs
they target the mitotic spindle.
Vinca alkaloids stabilise free tubulin, prevent microtubule polymerisation, arrest cells in mitosis
Paclitaxel (taxol) stabilises microtubules, preventing de-polymerisation, arrests cell in mitosis
not just cancer : colchine is used for immune-suppresion
