Cell Cycle Regulation and Checkpoints

Cell Cycle Regulation

• The cell cycle is regulated by cyclins and cyclin-dependent kinases (Cdks).
• Changes in cell cycle regulation can lead to uncontrolled growth and cell division in cancer.
• It's important to understand the actions of oncogenes and tumour suppressor genes in cancer.
• P53 plays a significant role in the regulation of the cell cycle.

Cell Cycle & Mitosis Revision

• A review of the cell cycle and mitosis is recommended before this session.

Cell Cycle Strict Regulation

• Cells divide when they receive appropriate signals.
• Signals determine if more cells of a specific type are needed.
• Signals determine if cells are healthy and without mutations that could cause cancer.
• The cell cycle checks if each phase is complete before moving to the next to avoid errors.
• The cell cycle can be stopped at several checkpoints.

Checkpoints

• G₂ checkpoint: Checks if DNA and DNA damage are repaired.
• Enter Mitosis occurs after the G₂ checkpoint.
• Checkpoint in Mitosis: checks if chromosomes are attached to the mitotic spindle?
• Pull Duplicated Chromosomes Apart happens after the checkpoint in Mitosis.
• Enter S Phase occurs when the environment is favorable.
• G₁ Checkpoint assess environment favorability.

Eukaryotic Cell Signals

• Mitogens stimulate cell division by overcoming the cell cycle "brake”.
• Growth Factors stimulate growth by promoting synthesis and inhibiting the degradation of macromolecules.
• Survival Factors suppress programmed cell death.

Growth Factors Explained

• Growth factors lead to an increase in protein synthesis.
• Growth factors lead to a decrease in the degradation of macromolecules

Checkpoint - Spindle Assembly

• Mitosis must not complete until all chromosomes attach to the mitotic spindle.
• The mitotic checkpoint delays until all chromosomes are connected.
• Prolonged checkpoint activation results in cell death.
• This is a mechanism for many anti-cancer drugs.

Protein Kinases

• Most enzyme activity is regulated within a cell.
• The cell can switch enzymes on or off.
• A common regulation mechanism is phosphorylation/dephosphorylation.
• Protein kinases add phosphate groups and can induce changes in the 3D structure of a protein, altering enzyme activity.

Cyclins & Cyclin-Dependent Kinases

• Cyclin-dependent kinases (CDK) regulate the cell cycle.
• CDKs are inactive when not bound to a cyclin protein.
• When active, CDKs phosphorylate and activate proteins involved in regulating the cell cycle.

Cyclin Activity

• CDKs are present in a cell throughout its cycle.
• CDKs cannot regulate the cycle on their own.
• M-Cyclin concentration increases during interphase and falls during mitosis.

M-Cdk Roles

• M-cyclin and mitotic Cdk combine to form the M-Cdk complex.
• M-Cdk phosphorylates and activates proteins that condense chromosomes, break down the nuclear envelope, and form the mitotic spindle.

Other Cyclins

• In addition to M-cyclin/Cdk, other cyclins/CDKs regulate the transition to S-phase.
• S-Cdk activates proteins to recruit DNA polymerase and trigger DNA replication.

S-cyclin & M-cyclin Relationship

• The levels of S-cyclin and M-cyclin fluctuate during different phases of the cell cycle, indicating their roles in these phases.

Major Cyclins & Cdks

• G₁-Cdk: Cyclin D partners with CDK4 and CDK6.
• G₁/S-Cdk: Cyclin E partners with CDK2.
• S-Cdk: Cyclin E (early) and Cyclin A partners with CDK2.
• M-Cdk: Cyclin A (early) and Cyclin B partners with CDK1.

Cell Cycle Regulation Complexity

• Cell Cycle Regulation is Complex and cyclical
• Cyclin levels can change dramatically during the cell cycle - and have complex relationships

Cell Cycle Machinery

• A cell can withdraw from the cell cycle, entering a quiescent state known as G₀.
• Cell cycle machinery is disassembled in G₀.
• Cyclins and CDKs disappear in G₀.
• Cells in G₀ neither divide nor prepare for division and can remain in this state for life.
• Re-entering G₁ from G₀ can take days.

Cell Cycle Stops

• A cell stops cycling if encountering problems.
• Problems include damaged DNA, an unfavorable extracellular environment, incompletely replicated DNA, or improperly attached chromosomes.

CDK & Cancer

• Mutations in CDKs and cyclins are common in cancer cells.
• These mutations allow a cell to progress through the cell cycle without the normal checks.
• These mutations increase the chance of cell division with potentially cancerous mutations.
• Common mutations seen in skin cancers include CDK4 (G1 Cdk).

What Causes Mutations?

• Spontaneous genetic code changes happen when mistakes occur during depurination.
• Generation of free radicals occurs.
• Changes in DNA happen due to food components, potentially leading to colon cancer.
• Cigarette smoke causes DNA damage, increasing the risk of lung cancer and cancers of the throat, tongue, and palate.
• UV light damages DNA, increasing the risk of skin cancers.

Exposure to UV Light

• UV light can damage DNA.
• UV light promotes covalent linkages between adjacent pyrimidine bases such as creating thymine dimers
• UV light may cause mispairing during replication.
• If unrepaired, pyrimidine dimers inhibit transcription, cause mutations, cell death, and skin cancer.

Proto-Oncogenes & Oncogenes

• Proto-oncogenes are normal genes that promote cell division, including protein kinases and growth factors. A mutant form of a proto-oncogene is called an oncogene.
• Oncogenes can cause cell division from a single mutation.
• Oncogenes are the result of gain-of-function or dominant mutations.
• When expression is wrong, uncontrollable cell division and cancer can be the result.

Proto-Oncogene RAS

• Normal RAS flips between active, bound to GTP, and inactive, bound to GDP.
• When active, RAS stimulates regulators of cell proliferation.
• 30% of cancers have a mutation in RAS.
• Amino acid change impairs GTP hydrolysis.
• The continual stimulation cell division with RAS is always active.

Tumour Suppressor Genes

• Tumour suppressor genes prevent cell division, through cell cycle checkpoints and apoptotic inducers.
• A loss of genes that inhibit cell division causes cancer.
• Mutations of both copies of a tumour suppressor gene are needed to allow cell division; recessive mutation.
• This control results in loss of proliferation, and uncontrolled cell division by the loss of this control.

Tumour Suppressor Gene P53

• P53 was first discovered as a mutation in Li-Fraumeni syndrome.
• Over 50% of human cancers involve an abnormal p53.
• P53 is "Guardian of the Genome.”
• P53 expresses when cells are exposed to DNA-damaging agents.
• P53 stimulates the expression of p21 and cell cycle arrest.
• DNA repairs or cells undergo apoptosis.
• It protects cells from genomic instability caused by DNA damage.

P53 Mechanism

• Following cellular stress, p53 induces the expression of p21, a Cdk inhibitor (also known as CDKN1A).
• Cells arrest in G₁ to allow for DNA repair.
• P53 is degraded in proteasomes in the absence of DNA damage.
• P21 is not synthesized in the absence of p53 activity.
• Cells divide with damaged DNA when the cycle is not halted.

Oncology Genetics

• Most cancers are sporadic, involving acquired mutations and epigenetic changes during an individual's life.
• Mutations are somatic, affecting specific tissues and not passed down.
• Some inherit single gene mutations, increasing cancer susceptibility.
• Requires a 2nd hit → Knudson hypothesis is required.
• The original mutation can be passed from generation to generation resulting in inherited cancer.