Cell Cycle/Cancer (D.M) Flashcards
What are the two main types of cell death?
Apoptosis –> opposite to cell proliferation
- Programmed cell death –> apoptosis –> A lot more regulated –> contents that need to be broken down are placed in lipid-bound vesicles which are then digested by phagocytes.
- Necrosis –> cells basically explode –> contents of the cell are released –> not useful as its a waster plus it may be harmful –> often results in inflammation.
Describe what an apoptotic cell would look like during different stages of apoptosis.
A –> cell provided with apoptotic stimulus
B –> shrinks
C –> shrinks further
D –> vesicle formation
Is apoptosis a normal process?
Apoptosis –> normal process
During development, it plays an essential role –> for example tissue between fingers removed via apoptosis + blood vessels are solid –> centre removed via apoptosis + neurons removed from the brain (connected in the wrong way)
Also important during adulthood –> cell damaged too much? –> cell dies via apoptosis/T cell elimination
What are the different stages in the cell cycle?
- Interphase –> majority of the time in this stage (G1 phase, S phase and G2 phase)
- Prophase –> chromosomes become visible
- Metaphase –> align on the central equator + spindle formation
- Anaphase –> separation –> move to opposite ends
- Telophase –> separation into two cells.
- Cytokinesis
2-6 –> Mitosis (division process)
What happens in G1, S and G2 phase?
Interphase
G1 –> Sensing phase –> good conditions for division?
S –> Replicates DNA
G2 –> Checking and cell gathers energy
G –> Growth or Gap
What happens in S phase?
DNA replication –> end up with two sister chromatids which will come together later in prophase to form a chromosome. Both sister chromatids are held together by cohesin complexes at the centromere which is a constriction point.
What happens in prophase?
Prophase
- Chromosome condensation –> important because the average length of DNA in chromosome about 5cm – nuclear diameter 5um (0.005mm).
How doe this happen?
Naked DNA –> wrap it around nucleosomes (beads on a string arrangement) –> wrap the nucleosomes around each other to form chromatin fibre –> loop the fibres in a protein scaffold –> associate scaffolds –> forms chromosomes.
- Mitotic spindle starts to form –> co-ordinated by centrosomes (type of organelle –> allow microtubules to grow from a specific position)
Structure of the spindle?
Microtubules originate from the centrosome
- Microtubules that move away from the centre –> called astral
- Microtubules that overlap with each other –> called overlap microtubules
- Microtubules that attached to chromosomes –> called kinetochore microtubules
What happens in metaphase?
In ProMetaphase
- Chromosome fully condensed
- Centrosomes at opposite ends of the cell
- Spindle formation is mainly complete
- Nuclear envelope breakdown (one of the main changes)
In Metaphase
- Chromosomes align along the equator –> achieved using spindle fibres that push and pull the chromosomes –> microtubules attach to kinetochore (the protein complex assembled at the centromere that binds to microtubules of the spindle) -> balanced pushing and pulling leads to central alignment.
What happens in anaphase?
Anaphase
- Sister chromatids separate –> allows chromatids to move opposite sides
- There are a bunch of cohesin proteins that hold the two sister chromatids together –> proteolytic enzymes needed –> three key players: APC (anaphase-promoting complex, Securin (inhibitor of separase), separase (protease –> specific for cohesins).
1. When chromosomes are aligned along equator –> APC is activated.
2. Cleaves securin –> releases active separase
3. Active separase can now cleave the cohesin –> sister chromatids are no longer attached.
4. No force holding chromatids together –> pulling action of microtubules pulls chromatids to opposite poles.
Explain what happens to the microtubules when the chromosomes separate.
Microtubules can be found in two forms:
- Growing –> has a GTP cap promotes elongation
- Shrinking –> GTP cap no longer present –> dynamic instability –> allows the microtubules to shrink rapidly.
Hence –> when chromosomes need to be separated –> Cap is removed –> shrinkage of microtubules –> seperation.
Three different types of motion that occur
- Kinetochore microtubules –> dynamic instability –> pull sister chromatids
- Overlap microtubules –> Push against each other –> push away centrosomes –> pushes centrosomes to opposite ends of the cell
- Astral microtubules –> Pulling from the edge of the cell –> further separating microtubules.
Explain what happens in telophase.
Telophase
- Chromosomes at opposite poles must decondense
- Nuclear envelope forms
- Microtubules/Spindle breakdown
What happens in cytokinesis?
Formation of two cells
- Twisting cells –> forms constriction point –> cells pop apart –> actin ring contracts separating the membranes
How are organelles distributed?
- When organelles are really abundant –> stochastic/random process
- Only one of the organelle –> active distribution of organelle –> undergo their own division
What experiments were conducted on cdc mutants?
Yeast were given mutations which resulted in…
- Yeast cells that grew too large before dividing –> resulted in elongated yeast.
- Yeast cells that didn’t grow much before dividing –> resulted in small circular yeast.
They then figured out the what mutations the yeast cells had –> complementation analysis –> From this analysis they figured out that there were 60 different genes that regulated the cell proliferation cycle (varying amounts of importance –> fundamental and accessory)
What was one of the main genes involved in cell cycle regulation?
Gene called cdc2 –> codes for a protein kinase (involved in signalling –> phosphorylation) –> one of the key regulators of proliferation process.
cdc2 can phosphorylate serine and threonine side chains on other proteins –> essential for cell cycle –> without you get elongated cells.
cdc2 concentration remains constant, however, its activity level fluctuates.
What are cyclins and how does their concentration differ along cell cycle?
Cyclin is a family of proteins that control the progression of cells through the cell cycle by activating cyclin-dependent kinase (CDK) enzymes.
How does CDK and cyclin activity relate?
CDK and cyclins activity similar activity levels –> maximum activity in G2 phase.
This is because the cyclin is a regulatory subunit for CDK (same as cdc2 but for humans) –> no cyclin –> kinase is not activated –> no activity.
How many different cyclins are there? How do they function?
There is a cyclin equivalent for each stage of the cell cycle –> they tell that specific phase to progress.
Basically –> specific cyclin is made at a particular time –> activates the CDK –> initiates a particular step in the cell cycle –> cyclin is destroyed at a defined time –> stops action CDK (step cannot be repeated) –> allows for ordered progression of cycle.
Note
G2 is the regulatory step for yeast
However, in humans, G1 is the regulatory step.
What is the restriction point?
Cell passes restriction point –> it must go through the entire cell cycle –> occurs in late G1 phase.
What is a key protein in restriction point control?
Retinoblastoma protein (Rb) –> 1 05kDa
Two forms of the Retinoblastoma protein –> Hypo- (active) and hyper-phosphorylated (inactive) form –> Low and high phosphate content respectively –> the phosphorylated forms have different conformations
G1 –> Hypophosphorylated form
S/G2/M –> Hypophosphorylated and Hyperphosphorylated form
Phosphorylation is cyclical
Note –> RB can be phosphorylated by G1 cyclins/CDKs
Explain the different phosphorylated forms of Rb effect the cell cycle.
Model of Cell cycle
Rb –> stops hyperproliferation (division)
G1 –> hypophosphorylated –> active form –> inhibits continuation in cell cycle
S/G2/M –> hyperphosphorylated form –> Rb is not stopping hyperproliferation –> allows for the latter stages in cell cycle.
How does Rb specifically influences the cell cycle?
When Rb is in the hypophosphorylated form…
Rb interacts with E2F protein –> transcription factors that bind to DNA in a site-specific manner –> by binding to E2F –> it can not act as a transcription factor.
What gene expression are impacted by E2F?
- S-phase cyclin
- DNA polymerase
- DHFR (metabolic gene –> responsible of nucleotide production for DNA replication)
- etc….
What happens step by step to Rb when a cell decides to go through the division process?
- G1 cyclins are activated
- Cyclins bind to the cdks
- Phosphorylates the Rb protein –> Hypo to Hyper phosphorylated form
- It undergoes a conformational change —> no longer bonded to E2F.
- E2F is free –> Activates transcription of genes required for S-phase
- The cell is now committed to S-phase
Once this occurs –> there is no going back –> irreversible switch.
How do cells leave the G0 phase?
- Growth hormone binds to cell
- Activates transcription of G1 cyclins –> bind to cdk
- Phosphorylates Rb
- Activates Cell cycle
What are Capases? How are they activated?
Humans code for a whole bunch of caspases –> they are cysteine aspartate proteases –> they cleave target proteins C terminal after an aspartate residue
- Cysteine (in active site) and targets aspartate
Activation
Inactive procaspase (largely inactive) –> to convert to active form you need two cleavages –> remove prodomain and between the red and pink area –> this allows the protein to fold and form active caspase.
What are adaptor proteins?
Allow bridging between two protein to allow processes to occur –> for apoptosis –> bridge caspase and the pro-apoptotic input signal.
What are regulator proteins?
Regulators
2 Forms
- Pro-apoptotic regulators
- Anti-apoptotic regulators
All these regulators have TM domain in their structure
What are the two types of apoptosis?
- Intrinsic pathway –> cell detects that it has been damaged (DNA damage –> leading to cancer cell formation) –> cell will recognise whether it can repair damage or not –> if not –> it will commit to apoptosis.
- Extrinsic pathway –> signal comes from the outside –> for the greater good of the organism.
Explain the process of the triggering apoptosis (intrinsic pathway).
Mitochondria act as a source to trigger apoptosis –> molecule responsible for this is cytochrome C –> found on intermembrane space but very rarely in the cytosol.
However, if it is found cytosol –> acts as a trigger for apoptosis –> due to leakage from the mitochondria.
Process
- The signal that causes the mitochondrial membrane to disrupt
- Cytochrome C is released into the cytosol
- Binds to adaptor proteins –> Apaf-1 –> results in conformation change of Apaf-1
- Apaf-1 can bind to more Apaf-1 molecule and procaspases.
- Procaspases have low levels of activity –>, however, when brought together in large quantities –> expect higher level of activity.
- When one caspase is in its activated to its active form –> chain reaction with the rest –> caspase activation cascade.
Regulator proteins –> regulate the stability of the outer mitochondrial membrane –> Pro membranes of this family can form pores in the membrane whereas the Anti membranes prevent this –> both members do this by integrating to the membrane using their T.M domains –> Interplay between regulators determines whether pores form or not.
Cancer cells –> overexpresses Bcl-2 –> prevents proe formation.
In the extrinsic pathway what protein triggers apoptosis?
Protein is called Fas –> Transmembrane protein –> has a cytoplasmic ‘Death domain’.
Fas ligand binds to the transmembrane protein –> tells the cell that it has to die.
Killer lymphocyte express Fas ligand –> Fas ligand interacts with its receptor –> results in the assembly/grouping of many Activated Fas receptors –> cause a conformational change of cytoplasmic domain –> allows it to interact with adaptor proteins –> recruit caspases –> when brought together in large quantities –> caspase cascade –> leads to cell death.
Why do a lot of vesicles appear on the surface of the cell during apoptosis?
Vesicles budding of the surface of the cell –> filled with degraded proteins, DNA etc….
These vesicles are then released in the circulatory system –> engulfed by macrophages –> fuse with lysosomes –> further degradation to a.a, nucleotides, etc.
- Phosphatidylserine is useful –> expressed on vesicle –> tell macrophages to digest.
Definition of cancer? Breakdown the definition –> explain it.
A group of diseases generally characterised by genomic instability and uncontrolled cell division and leading to the invasion of surrounding tissue and eventual dispersal to distant sites.
The definition is not absolute (applies mostly to solid tumours) –> does not apply to all cases
Breakdown
- ‘A group of diseases’ –> many different conditions –> can literally affect all cells in the body + even though two people have the same condition –> completely genetically unique.
- ‘genomic instability’ –> genomes of the cancer cells change over time –> cells are under evolutionary pressure to survive –> constant genetic change that accumulates overtime –> ‘snowball effect’.
- ‘uncontrolled cell proliferation’ –> forms tumour –> useful for detection.
- ‘Lending to the invasion of surrounding tissue’ –> surface of skin forms lump/internal results of invasion of other tissue (disrupts organ function) –> disrupts tissue architecture.
- ‘dispersal to distant sites’ –> usual cause of death –> called: metastasis –>
What are some common types of cancer called?
What are the three main causes of cancer?
- Inheritance –> from parents
- Genetic change –> most cancers caused by genetic change –> Changes to DNA (mutations)
- Viral infection –> uncommon
Describe how genetic change leads to tumour formation.
Genetic change –> main cause of cancer –> for example 90% of the time cancer is caused by random genetic change.
Caused by defective replication or mutagens (irradiation, chemicals, etc…)
Possible mutagens:
Ionising radiation - x-rays, ultraviolet radiation and sunbathing
Chemicals - environmental aflatoxin, man-made and self-made –> Metabolic origins of mutagens - oxygen species
Results in different types of changes to DNA:
- Point mutation –> thousands of possibilities
- Deletion of segments of chromosomes –> after multiple divisions –> lose these segments (may be important for cancer suppression)
- Translocation
- Inversion
The relationship between Age and incidence of cancer?
Longer you live —> more likely to get tumours.
How can we use Darwinian selection to understand what type of tumour cells proliferates the most?
Initially, tumour cells only have a couple of mutations –> overtime due to the genomic instability –> they accumulate more and more mutations –> different cancer cells with different mutations –> over time the cell type that is the ‘fittest’ will predominate in tumours.
Note –> there is always constant genomic change –> however, tends to be one founder cell with a core set of mutations.
Makes it difficult to treat –> different cells with different mutations –> constant mutations/’moving target’.
What are the differences in the genome that make a virus able to turn a normal cell into a cancer cell?
Case study –> Rat sacroma virus
Normal basic Retroviral RNA –> LTR-GAG-POL-ENV-LTR
Cancerous retroviral RNA –> LTR-GAG-POL-ENV-RAS-LTR
Call it v-RAS (v=Viral)
Humans also have RAS proteins present in all cells –> Called c-RAS –> very similar to c-RAS –> main difference is a single substitution –> glycine substituted for valine
Are there many different types of retrovirus with their own oncogene associated with it?
YES!
All the oncogenes have a normal cellular equivalent –> viral versions –> result in these proteins not being switched off –> activates downstream signalling –> leads to cell proliferation.
In the human papilloma virus, which genes/proteins are required for a transformed phenotypes (no cancerous to cancerous)
Deletion analysis indicates that only the E6 and E7 proteins are required for transformation of the phenotype.
How do E6 and E7 cause tumour formation?
E6 and E7 are tumour supressors
E7 binds very well to the Rb protein –> binds better to Rb than E2F –> when E7 is present in cells –> E2F is free –> acts on DNA (transcription factor) –> drives S phase transcription.
E6 binds to p53 protein (found on chromosome 17) –> influences apoptosis and cell cycle arrest.
- Prevents apoptosis by –> E6 stops p53 from inhibiting Bax –> Bax is free to inhibit Bcl-2 proteins –> Bcl-2 normally increases the formation of pores in the mitochondrial membrane –> leads to the leakage of cytochrome C –> process is inhibited as Bcl-2 is inhibited by bax.
Prevents cell cycle arrest (E6 triggers continuous division)–> E6 binds and inhibits P21Cip1 from inhibiting CDK –> now CDK is free to phosphorylate Rb –> releases E2F –> cell division –> normally P21Cip1 inhibits CDKs from phosphorylating.
So E6 and E7 force the cell to proliferate in an uncontrolled manner and also stops the cell from killing itself.
Note –> in order to get cancer –> we need to lose both alleles of the normal functioning forms –> because we are not activating an oncogene but we are inactivating proteins that suppress cancer (only one allele affected –> still have the other producing normal proteins).
Difference between oncogene mutation and interactivity mutation.
In the cell cycle what key players are gonna be a source of potential oncogenes and tumour suppressor genes.
