Cycle 8+9 Workshop Flashcards
Define:
Stem cells
Cells that are able to differentiate into multiple types of cells
List:
Types of stem cells
- Embryonic
- Somatic
- iPS
- Umbilical cord stem cells
How are stem cells different from normal cells?
They have to express telomerase as they persist for much longer than normal cells and have to maintain telomere length
Where do umbilical stem stells come from?
Come from embryogenesis and some stay with us in adult life
List and define:
Potency/origin of stem cells
- Totipotent: Can form an entire viable organism
- Pluripotent: Can form nearly all cells
- Multipotent: Can differentiate into a family of cells
- Unipotent: Can only form one cell type
Give an example of:
Totipotent cell
Only the zygote is totipotent
Give an example of:
Pluripotent cells
Nearly all cells
Give an example of:
Multipotent cells
Cells of a specific tissue type
Give an example of:
Unipotent cell
Epidermal cells
State the difference between:
Somatic cell and a Unipotent Stem Cell
Unipotent stem cells
* Give rise to cells that won’t reach cell senescence (self-renew)
Somatic cells
* Eventually reach their Hayflick limit, must enter irreversible cell cycle arrest
True or False:
All stem cells have the exact same genome
True
What are the two types of regulation of gene expression that determine what cell type a stem cell can give rise to?
Temporal
Spatial
What plays a big role in determining the type of cell the stem cell give rise to?
Tissue-specific transcription factors
Define:
Spatial regulation (in terms of stem cell regulation)
Where (in what cell/part of the body)
Define:
Temporal regulation (in terms of stem cell regulation)
When (at what time/under what conditions)
List:
The stages that we can control gene expression
- Transcription
- Post-transcription
- Translation
- Post-translation
Why is regulation at the transcriptional level very important?
Prevents the waste of resources and energy to transcribe a protein when it is halted down the process line
What is transcription controlled by?
Transcription factors at its regulatory sites (promoter, proximal and distal regulatory sites)
How do stem cells divide? Describe
- Symmetric self-renewal: From 1 to 2 stem cells (maintains count, extra cell can stay or go)
- Asymmetric: From 1 stem cell, to 1 stem cell and 1 progenitor (maintains count, other is committed to differentiation)
- Symmetric differentiation: From 1 stem cell, to 2 progenitors (loses count)
In symmetric differentiation, how is the stem cell count maintained?
The neighbour has to symmetric self-renew to maintain count
Define:
Progenitor cells
Stem cells that have committed to differentiation, cannot go back to the niche
What are the two types of progenitor cells? Explain
- Multipotent: Upon differentiation
- Committed: After further differentiation, they are more differentiated and committed than multipotent
What are the role of stem cells in the body?
Maintain tissues in the body
* Epidermal stem cells replenish epithelial skin cells
* Intestinal villi stem cells stay in quiescent stage until transcription factors trigger them to migrate
How is potency of a stem cell tested?
- The unknown stem cells are labeled with fluorescent marker
- Injected in inner cell mass of blastocyst to form a chimera
- Implanted into pseudopregnant mouse
- The offspring is scanned for fluorescent marker
How does the appearance of the marker determine the potency of the stem cell?
- Marker is in wide range of tissue: Pluripotent
- Marker is in 1 type of tissue: Unipotent
Define:
iPSCs
Induced Pluripotent Stem Cells (iPSCs)
* Fibroblasts (obtained easily and non-invasively) that are reprogrammed by adding the 4 Yamanaka factors
What are Yamanaka factors?
Transcription factors (TF) that tell the fibroblast to revert to a stem cell
Why are iPSCs very important?
- Very powerful as they can recreate organ systems in vitro without having to get embryonic stem cells
- More predictable behaviour and easier to culture
List:
The 4 main components of the Lac operon
- Promoter
- Operator within promoter
- 3 lac genes
- Regulatory lacI gene upstream of lac operon
What are the 3 lac genes?
- lacZ: Involved in galactose metabolism, encodes beta-galactosidase
- lacY: Involved in galactose metabolism, encodes permease
- lacA
True or False:
The three genes of the operon can be transcribed separately
False, as part of an operon, either all 3 genes are transcribed and expressed simultaneously or none are
What does lacI code for?
The lac repressor, which binds the operator of the lac operon to suppress expression
How is expression of the lac operon regulated?
via transcriptional regulation
Describes:
What happens when lactose is present to the lac operon?
The lactose is converted to another form called allolactose, which binds to the lac repressor and inactivates it, meaning it can no longer bind to the promoter and start transcription
What are spliceosomes?
Complexes made of snRNPs (protein + RNA) and the pre-mRNA
Where is pre-mRNA cut?
5’ and 3’ splice site recognition sequences
What is the 3’ splice site?
NCAG
True or False:
Exons are removed, introns are joined together
False, the intron is removed, exons are joined together
What is abberant mRNA splicing?
When one splice site is mutated beyond recognition, and the splice is not correct (may be non-functional as it may contain intronic region)
What causes beta-thalassemia?
- Single base substitution
- Creates new 3’ splice site in the middle of intron
- Causes some of the intron to be left in the mature mRNA
- Creates early stop codon, causing much smaller hemoglobin protein to be synthesized
What is the basic idea behind beta-thalassemia treatment?
- Using CRISPR Cas9 gene editing
- Drug is a specific form of fetal hemoglobin
- Targeting BCL11A, not transcribed meaning that it doesn’t suppress gamma globin gene transcription, cells can produce fetal hemoglobin
What is Alzheimer’s disease?
- Progressive and fatal neurodegenerative disease, most common cause of dementia
- Starts in hippocampus and spreads outwards
- Loss of neurons, memories and cognitive function
Define:
Dendrites
Receive signals from surrounding neurons
Define:
Soma
Cell body with organelles
Define:
Axon
Long extension of the cell, carries neurotransmitters, nutrients and signals from presynaptic neuron to axon terminal; has myelin sheath
Define:
Synaptic cleft
Gap where presynaptic neuron axon terminals meet postsynaptic neuron dendrites
Define:
Microtubules
Essential for nutrient transport and structural support. Stabilized by tau proteins
What is the function of cholinergic neurons?
Release acetylcholine transmitters (acetyl-CoA + choline)
What synthesizes acetylcholine? Where does it do it?
Choline acetyl transferase in the neuron
What is acetycholinesterase (AChE)?
An enzyme on the postsynaptic neuron that degrades acetylcholine back into its components
Describe:
The function of acetylcholine in a healthy individual
- Released from the axon terminals of one cholinergic neuron
- Bind to a receptor on the dendrite of the postsynaptic cholinergic neuron
- Binding sends out a signal
Describe:
Function of acetycholine in patients with AD
- Low levels of choline acetyl transferase (CAT)
- Pathway is downregulated
Why is acetycholine binding to the postsynaptic cholinergic receptor important?
Important in initiating a signal cascade
What drug is used to treat cholinergic neuron degeneration in AD? Why?
- Acetycholinesterase inhibitors (e.x. rivastigmine)
- Inhibits the enzyme that breaks down acetycholine, meaning there is more acetycholine around to transmit signal
Define:
Amyloid precursor protein (APP)
A transmembrane protein that is found in neurons
* Essential in the function, maintenance and repair of neurons
* Not present indefinitely (has lifespan, eventually broken down)
List:
The two pathways that APP can be degraded
- Alpha-secretase + gamma-secretase
- Beta-secretase + gamma-secretase
Describe:
Alpha + Gamma Secretase degredation pathway for APP
- Cut into small, soluble peptides released into extracellular environment
- SOLUBLE (cell knows how to deal with this)
- Dominant degradative pathway
Describe:
Beta + Gamma Secretase pathway for APP
- Beta secretase cuts at different locations, released into extracellular environment
- Produces larger, INSOLUBLE amyloid beta peptide (36-42 amino acids long)
- Involvement of microglia, astrocytes, and apolipoprotein E are needed to get rid of the amyloid beta peptide
Describe:
APP degredation in patients with AD
- More beta secretase than alpha secretase
- Higher production of insoluble amyloid beta
- Lower clearance of amyloid beta
- Leads to plaques
How do amyloid beta plaques cause brain deterioration?
- Excessive release of toxic cytokines leads to inflammatory response, thus neuron loss
- Overactivation of microglial cells and astrocytes leads to glutamate build up and neuron damage
- Crowded synaptic clefts blocks receptors, leads to decreased communication between neurons
What are NMDA antagonists?
Inhibit NMDA receptors on the postsynaptic neuron so that excess glutamate has no effect on neurons
* Relieves symptoms, but does not cure disease
Define:
Microtubules
Key structures of cells (allow molecules/nutrients from stoma to reach axon)
Define:
TAU
Type of protein that forms subunits in microtubules to hold them together
Describe:
TAU and microtubules in AD patients
- TAU protein is hyperphosphorylated
- Causes TAU protein to dissociate from the microtubule subunits
- Microtubules then start to fall apart
True or False:
If microtubule is non-functional, the cell is mutated but doesn’t die
False, the cell dies
What happens to hyperphosphorylated TAU proteins?
Start to stick to one another, forming intracellular clumps, contributes to cell death
What are the clumps formed by hyperphosphorylated TAU proteins known as?
Fibrillary tangles
What genetic risks increase the chances of AD?
- Down Syndrome
- Presenilin 1 and 2
- ApoE4
Describe:
Down Syndrome and relation to AD
- Extra chromosome 21 (encodes APP)
- Higher expression of APP and a much higher risk of early onset AD
Describe:
Presenilin 1 and 2, its relation to AD
- Autosomal dominant inheritance
- Mutant gamma-secretase creates larger amyloid beta peptides (42 aa) leading to greater aggregate production
Describe:
Inheritance of ApoE4, its relation to AD
- A less effective variant of ApoE4 that is not effective at removing amyloid beta aggregates
- High risk of early onset AD
What is single cell genome sequencing?
The genome of each individual cell within a tumour splice can be sequenced
Define:
Intratumoral heterogeneity
- Genetic differences between cells of one tumour in one affected patient
- A single tumor has multiple patterns of gene expression of different cells
Why is intertumoral heterogeneity important?
- Contributes to the complexity of cancer and treatment
- Precision medicine, takes into account individual cariability in genes, environment, and lifestyle for each person
Define:
Intertumoral Heterogeneity
- Between many individuals (even with the same cancer type), you observe differences in gene expression, mutation sequencing
Why is intertumoral heterogeneity important?
- Personalized medicine looks at intra-tumoral problems, wondering which mutations exactly to target
True or False:
As cells accumulate mutations, the probability of having an oncogenic cell stays the same
False, the probability of having an oncogneic cell increases
Oncogenic cells will:
- Proliferate
- Avoid apoptosis
- Grow
- Move and invade
- Create new blood vessels
- Evade body’s immune response
What is the term for creating new blood vessels? What is the purpose of this?
Angiogenesis
* To ensure a constant supply of nutrients
What are rounds of mutations known as?
Clonal expansions
Does the order of mutation introduction have an effect?
Yes
How does clonal evolution and genetic heterogeneity pose an issue to cancer treatment?
Which cell mutations should we target?
Define:
Passenger mutations
Mutations that do not contribute to cancer development, ‘along for the ride’
Describe:
Passenger mutations
- Results from normal mutation processes (when DNA divides, endogenous factors)
- Not really selected for, inert mutations
Define:
Driver mutations
- Mutations that contribute to cancer development, and encourages excessive cell growth
Describe:
Driver mutations
- First does not cause severe consequence
- Classified as a mutator phenotype when compuled with a second
- Third one is early invasive cancer
- Usually takes 2-5 to get cancer
- Required for causation, progression, and maintenance of cancer
- Growth advantage (selected for)
What are the two categories of driver mutations (causes of cancer)?
- Proto-ongenes
- Tumour-suppressor genes
Why do driver mutations tend to be in proto-oncogenes and tumour-suppressor genes?
They are more likely to lead to cancer progression
What are proto-oncogenes?
Genes involved in promoting cell survival, growth, and movement
* Mutations in genes are likely to lead to oncogenic phenotypes (e.x. growth factors)
In proto-oncogenes, what does a mutation imply?
Gain of function: Pathway is overexpressed, cancer cells can induce receptor to be constitutively active
Give an example of a growth factor receptor
Receptor Tyrosine Kinase (RTK), which binds to growth factor, activating a transcription factor that will bind to promoter/regulatory regions; activating expression genes necessary for cell growth
What are tumour suppresor genes? What happens when they mutate?
Genes that are involved ins topping cell cyle, inducing apoptosis, recognizing DNA damage
* Mutations causes loss of regulation of cell cycle
What is the most common mutation involving tumor suppressing gene?
P53 (over 60% of cancers involve this mutation)
* Mutations in the DNA binding domain region of P53 hinders ability to be a transcription factor
What are cancer stem cells?
- Slowly dividing cancer cells
- Resistant to chemotherapy
- Results in tumour lapse
