Cell Cycle Flashcards
In cell cycle eukaryotes…
Replace lost body cells
Replace old and worn out cells
Undergo clonal expansion
Embryonic development
Interphase
G1, S, G2 sub-phases
(G- gap (growth) between main phases)
(S- synthesis of DNA)
M phase
Mitotic phase
- Mitosis: division of nucleus
- Cytokinesis: division of cytoplasm and organelle
Cells out of cell cycle are what?
G0 (G zero)
What cells lose their ability to enter cell cycle?
Post-mitotic cells: terminally differentiated cells that have lost their
ability to replicate.
- They are said to be permanently arrested.
- Usually highly specialised cells.
- Contact inhibition can also cause cells to exit the cell cycle
What cells can enter the cell cycle?
Cells with high mitotic activity
• High turnover cells are constantly in the cell cycle.
Cells that divide upon appropriate stimulation
• Most of the cells in our body only divide upon stimulation from growth
factors, hormones or other signals.
G1 phase
- Stimulus or signal (eg growth factor) required for entry from G0.
- G1 is the longest phase of cell cycle & cell volume doubles
• Period where the cell grows and prepares for S phase.
(e.g metabolic changes preparation for S phase ensures adequate reserves for organelles to replicate, Synthesis of enzymes and other cellular components needed for DNA synthesis)
• Entry to S phase will be delayed if the cell is not fully prepared
- In unfavourable conditions such as growth deprivation or in the presences of growth inhibitory signals the cell can withdraw from G1 and go back to G0 until conditions improve
- Many biological processes in this phase are driven by regulatory proteins specific to G1
S phase
- Replication is semi-conservative
- Original DNA strands used as templates for the synthesis of new strands.
- Daughter cells inherit one parental DNA strand and one new strand.
- Occurs in 3 steps
- Initiation > Elongation > Termination
DNA replication: INITIATION
DNA UNWINDS
• DNA Helicases are necessary for this. Process requires ATP to
the break hydrogen bonds between base pairs
UNWINDING = SUPERCOILING IN OTHER REGIONS OF DNA
• DNA topoisomerases relieve this tension by unwinding
‘supercoiled’ regions in the DNA, they also prevent knotting
downstream of the DNA so that Helicase can continue strand
separation
• Topoisomerases work by cutting one or both strands of the
DNA duplex at key points, untwisting the strands to relieve the
tension, then resealing the breaks.
UNWOUND DNA KEPT OPEN FOR REPLICATION
• Single strand binding proteins (SSB) bind temporarily to
separated DNA strands as soon as they have been unwound by
DNA Helicase to keep the strands apart during replication
Origins of Replication
- The size & complexity of the genome necessitate multiple replication starting points called Origins of Replication (ORC) for rapid DNA synthesis
- ORC occur along the length of each chromosome, In humans approx 30,000 to 50,000 ORC are visible along the length of chromosomes during S phase
Helicases unwind DNA in what way?
Helicases unwind DNA away from the origin of replication in a bidirectional manner. This creates replication bubbles that grow bigger in both directions as DNA unwinding progresses
Replication fork
The Y-shaped region at the furthest end of each replication bubble is referred to as the replication fork.
DNA replication: ELONGATION
- In humans, DNA Polymerase (delta) is the key player in this step. However it cannot synthesise DNA de novo so a primer is required
- Primases (DNA Polymerase ) short RNA primers, which serve as starting points for the DNA elongation process. Primers can be 10-20 primers long and provide the initial hydroxyl group for elongation
- DNA polymerase then sequentially adds DNA nucleotide in the 5’ to 3’ direction of the DNA strand.
- The DNA double helix is anti-parallel. One strand runs in the 5’ to 3 direction. The other strand runs in the 3’to 5’
- DNA replication is therefore bi-directional
Leading strand
5-3 direction, replication is continuous in elongation
Lagging strand
Anti-parallel strand
in the opposite direction to the leading strand but in a fragmented manner (in keeping with the 5’ to 3’ extension rule)
The fragmented DNA strands are known as Okazaki fragments
DNA replication: TERMINATION
- An endonuclease removes the RNA primer
- DNA polymerase fills the gaps with DNA nucleotides according to the rule of base pairing
- DNA ligase joins the Okazaki fragments to produce a continuous chain
- DNA polymerase can proofread its own work. If mistakes happen it removes them immediately.
- Other DNA repair mechanisms can be activated to correct errors as necessary
TELOMERES
• Removal of last RNA primer at the very end of lagging strand creates a gap which cannot be filled since DNA polymerase works from a primer template.
• Telomerase enzymes correct this shortfall by adding non coding 6-8 bp DNA repeats
to the end fragments.
- These repeat sequences of non coding DNA at chromosomal ends are called telomeres. They ‘cap’ (stabilise) chromosome ends and prevent loss of genetic information. They also prevent the ends from becoming tangled.
- Telomeres progressively shorten (senescence) in certain types of cells. When telomere length shortens to a critical point the cell dies.
DNA replication vs PCR
PCR:
- No Helicase, Topoisomerase or Primase necessary
- DNA primers (not RNA)
- No leading or lagging strands
S phase: DNA REPLICATION SUMMARY
DNA REPAIR MECHANISMS FOR
- Single strand DNA damage
- Double strand DNA damage
Single:
Base excision repair
Nucleotide excision repair
Mismatch repair
Double:
Non-homologous end joining
Homologous recombination repair
DNA repair failure in human cancers
BRCA1 and BRA2 defects due to double strand breaks- cannot be repaired so join together by non-homologous recombination so lose a piece of DNA in breast cancer
What happens at end of S phase?
Chromosomal DNA content doubles (2N to 4N)
• Two identical strands of each chromosome.
• Sister chromatids joined at the centromere.
moves on to G2 phase
M phase
MITOSIS (PMAT):
• Division of nucleus.
• Involves 4 main phases, based on the physical state of the
chromosomes and spindle.
• Prophase, Metaphase, Anaphase & Telophase
CYTOKINESIS
• Final step in the cell cycle.
• Division of cytoplasm and organelles
MITOSIS STEPS
- Prophase: chromosomes condense, centrosomes move to opposite poles, mitotic spindle forms
- Metaphase: centrosome are at opposite poles, chromosomes are at their most condensed and line up at the equator of the mitotic spindle. Mitosis will not continue if chromosomes are not properly aligned
• Anaphase: sister chromatids separate
synchronously, each new daughter chromosome moving to the opposite spindle pole.
• Telophase: chromosome arrives at the spindle poles, chromosomes decondense, nuclear envelope reforms
Meiosis vs Mitosis
What are the 3 major cell cycle checkpoints?
- G1 or G1/S checkpoint: Controls progression from G1 to the S phase.
- G2 or G2/M checkpoint: Controls progression from the G2 phase to the Μ phase.
- Metaphase (M) checkpoint: Controls progression from Metaphase to Anaphase.
What is phosphorylation?
- Addition of a Phosphate molecule to a protein.
- Most common post-translational protein modifications in eukaryotic cells.
Effects: The strong negative charge on a phosphate group can:
• Alter the physical properties modification of the protein shape.
• Alter the biochemical e.g how it interacts with water (a hydrophobic protein can will become hydrophilic when phosphorylated.
• Result in acute and reversible regulation of protein function e.g modulating protein folding, substrate affinity, stability and activity.
Advantages as a control mechanism
• It is rapid, taking as little as a few seconds.
• It does not require new proteins to be made or degraded.
• It is easily reversible.
• Particularly important regulation mechanism in the control of cell cycle progression
What are cyclins?
- Family of proteins that control the progression of cells through the cell cycle undergo a constant cycle of synthesis and degradation during cell division.
- Their concentrations is tightly regulated to ensure the cell cycle progresses in a proper sequence.
- Levels and type expressed vary depending on the cell cycle phase.
- At their peak levels when the protein they need to regulate is need.
What are cyclin-dependant protein kinases?
- Enzyme that adds negatively charged phosphate groups to other molecules by Phosphorylation
- Levels fairly stable throughout the cell cycle.
- Inactive until a Cyclin binds to it.
- Degradation of the bound Cyclin terminates the Cdk activity.
What is the cyclin-CDK complex?
- The active complex sole function is Phosphorylation of proteins in a cell cycle phase specific manner.
- The phosphorylated protein will then trigger specific events within the cell cycle (e.g. DNA polymerase gene expression (G1) through interaction with Retinoblastoma (Rb) protein.
- After the event, the bound cyclin is degraded and the CDK becomes inactive again
Phosphorylation of what allows transcription of more Cyclin E?
Rb-E2F complex
Retinoblastoma protein (RB): • A key cyclin-CDK target protein in G1 and G1 to S progression. • Transcription Regulator (Gatekeeper). • A major tumour suppressor protein. • can prevent entry into S phase and induce cell-cycle arrest
Elongation factor-2 (E2F)
• Transcription factors that regulate the expression of genes important in cell proliferation,
particularly those involved in progression through G1 and into the S-phase of the cell
What families of CKIs are key players in stopping cyclin-CDK activity during cell cycle?
p53 in cell cycle
- Activated in response to DNA Damage e.g Radiation by Phosphorylation.
- Phosphorylated p53 is active form and has a longer half life.
- Induces for CDKI expression (e.g p21) which causes the cell to arrest in G1 until the damaged DNA is repaired.
- If the DNA cannot be repaired it induces APOPTOSIS (programmed cell death).
- Important tumour suppressor protein.
What are mitogens and how do they regulate the cell cycle?
- Cytokines (small proteins 5-80 kDa) that stimulate cell proliferation.
- Bind to their cell-surface receptors and activate the signalling pathways that lead to cell cycle initiation and ultimately cell proliferation.
• Differentiated cells enter from G0
to G1 after the action of growth factors.
• Some mitogen genes and mitogen receptor genes are proto-oncogenes (genes which can mutate to become oncogenes/cancer promoting genes)
