1Checkpoints: what they check and why there

Learn each checkpoint as three things: when, what it checks, and why there.

If the problem is DNA damage, the cell is held at the checkpoint while repair enzymes work. If the damage cannot be repaired, the checkpoint signals trigger apoptosis, removing the cell before it can pass the damage on.

A checkpoint is a stop before a step that cannot be undone: G1 before copying the DNA, G2 before dividing it, M before pulling the copies apart.

Each checkpoint sits just before a commitment: replication, entry into mitosis, the separation of chromatids. That is the answer to "why at that point?"

2Cyclins and CDKs: the engine that drives the cycle

• A cyclin-dependent kinase (CDK) is a kinase that is present in the cell at a fairly constant level throughout the cycle, but is inactive on its own.
• A cyclin is a protein whose concentration rises and falls through the cycle. It is made steadily during one stage and then rapidly broken down.
• When a cyclin binds its CDK, the cyclin–CDK complex is active. It phosphorylates target proteins that carry the cell into the next stage, for example into S phase or into mitosis.
• When the cyclin is broken down, the CDK becomes inactive again, and the cell cannot simply rush through the next checkpoint until the next cyclin builds up.

Figure 1 shows two cyclins through a 24-hour cycle. One builds up during G1; when it passes a threshold, enough cyclin–CDK complex is active to take the cell into S phase, and the cyclin is then broken down. A second builds up during S and G2; when it crosses the threshold the cell enters mitosis, and its destruction during mitosis lets the cell leave mitosis and divide. CDK levels, not drawn, would be flat; kinase activity follows the cyclin.

Which cyclin pairs with which CDK, and the names of the external growth signals that start the cycle, are not examined. What you need is the mechanism: a constant kinase, switched on by a rising cyclin, switched off when the cyclin is destroyed. External signals that tell a cell to divide work through signal transduction pathways (topic 4.3) that end by increasing cyclin production.

3Reading cell-cycle data: where are cells held?

Experiments often sort cells by DNA content: G1 cells have one amount, G2 or mitotic cells twice that, S cells something between.

An invented data set. A culture of human cells was exposed to a dose of radiation that breaks DNA, and the percentage of cells in each phase was measured 12 hours later. The same was done with a mutant line whose G1 checkpoint does not work.

Claim: DNA damage holds normal cells at the G1 checkpoint, and the mutant line fails to stop.

Evidence: after radiation, the percentage of normal cells in G1 rises from 58% to 79%, and the percentage in S falls from 26% to 6%. In the mutant, the distribution after radiation (54%, 29%, 17%) is close to that of normal untreated cells.

Reasoning: cells stopped before S pile up in G1 and few enter S, as in the normal cells. The mutant's cells keep entering S with damaged DNA, so its G1 checkpoint is not working, and they will copy and pass on the damage.

The percent change for normal cells in S after radiation is $\frac{6 - 26}{26} \times 100 = -77\%$.

4When control fails: cancer and apoptosis

Cancer. Cancer is a disease of uncontrolled cell division. It arises when mutations disrupt the cell-cycle controls. Two kinds of fault work together:

• a protein that drives the cycle (a signal pathway component, a cyclin, a CDK) becomes stuck on, so the cell divides without the signal to do so;
• a protein that stops the cycle at a checkpoint is lost, so damaged or unneeded cells are no longer held or sent to apoptosis.

Usually several such mutations build up in one cell line. The cells divide again and again and form a tumour, collecting more mutations as they go because damaged DNA is no longer stopped.

Apoptosis. Apoptosis is programmed cell death (topic 4.3). In cell-cycle control it is the cell's last line of defence: a cell with damage it cannot repair is destroyed in an orderly way. If the signals for apoptosis are lost, damaged cells survive and keep dividing, which is another route to cancer.

Predicting disruptions. Use the same reasoning as for pathways: