Biology › Gene regulation, genomics and biotechnology › Mutation and cancer: what goes wrong, and where it goes wrong
Mutation and cancer: what goes wrong, and where it goes wrong
Most mutations do nothing. A few change one amino acid and are survivable. A very few land in the machinery that decides how often a cell divides, and those are the ones that turn into tumours — sometimes without changing a single base, because a promoter can be silenced instead of broken.
Before this The genetic code: triplet, degenerate, non-overlapping · Transcription factors and the promoter region · DNA methylation and histone acetylation · The cell cycle and its checkpoints · Stem cells and levels of potency
Before you start
A base substitution that does not change the amino acid has no effect — a silent mutation is a mutation that never really happened. The first clause is right and the conclusion drawn from it is not. A synonymous substitution leaves the primary structure of the protein unchanged, and that is what an exam means by silent. But 'silent' is a statement about one triplet in a coding sequence, and it says nothing about the rest of the genome. Change a base in a promoter and you change how much protein is made. Change a base at the boundary of an intron and the pre-mRNA is spliced wrongly and the protein is ruined. Some forms of β-thalassaemia are caused exactly that way, with the coding sequence of the haemoglobin gene entirely intact. A mutation that changes no amino acid can still change everything about how much of that protein a cell has.
What you should be able to do
- Name the types of gene mutation and give the consequence of each at the level of the polypeptide.
- Explain why many base substitutions change nothing about the protein, and why a deletion of one base usually changes everything.
- Distinguish a mutation in a gene from one in a control sequence, and give an example of each.
- Explain the difference between a proto-oncogene and a tumour suppressor gene, including how many alleles must be affected.
- Explain how methylation of a promoter can silence a tumour suppressor gene without altering a single base.
- Describe the therapeutic uses of stem cells and set out the ethical arguments on both sides of the embryonic case.
The kinds of mutation, and what each does to a protein
A mutation is a change in the base sequence of DNA. You have already met the two commonest kinds as copying errors during replication; here they are set out with what each does downstream, because that is what questions ask for.
A substitution replaces one base with another and affects one triplet only. Three outcomes are possible. If the new triplet codes for the same amino acid — which happens often, because the code is degenerate and triplets sharing an amino acid usually differ in the third base — the mutation is silent and the polypeptide is unchanged. If it codes for a different amino acid the mutation is missense, and the effect ranges from nothing at all to catastrophe, depending entirely on where in the protein the changed residue sits. If it produces a stop codon the mutation is nonsense, translation ends early, and the truncated polypeptide is almost always useless.
An insertion or a deletion of one or two bases is different in kind, because the code is non-overlapping and read in threes from a fixed start. Every triplet downstream is read from a new position — a frame shift — so every amino acid after the mutation is potentially wrong, and a stop codon usually turns up early by chance. One base lost near the start of a gene destroys the protein. Three bases lost remove one amino acid and shift nothing, which is why the commonest cystic fibrosis allele, a deletion of three bases removing a single phenylalanine, produces a protein that is made and folded but delivered to the wrong place rather than one that is gibberish.
| Mutation | What changes in the DNA | Effect on the protein |
|---|---|---|
| Substitution, silent | One base swapped; new triplet codes for the same amino acid | None to the amino acid sequence |
| Substitution, missense | One base swapped; new triplet codes for a different amino acid | One amino acid different; effect depends on where it is |
| Substitution, nonsense | One base swapped; new triplet is a stop codon | Polypeptide cut short, almost always non-functional |
| Deletion or insertion | One or two bases lost or gained | Frame shift: every triplet after it is misread |
| Duplication | A section is repeated | Frame shift unless the repeat is a multiple of three |
| Inversion | A section is reversed within the chromosome | The reversed section codes for a different sequence entirely |
| Translocation | A section moves to a different chromosome | Genes at both break points may be disrupted or newly switched on |
A human inherits roughly sixty to seventy new point mutations that neither parent had, most of them from the father, since sperm come from many more rounds of division than eggs do; almost all land in non-coding DNA and do nothing. The rate rises with exposure to mutagens: ultraviolet light, which links adjacent thymine bases; ionising radiation, which breaks strands; and chemicals such as the benzopyrene in tobacco smoke, which distorts the helix so that polymerase misreads it.
A mutation in a gene, and a mutation in the switch
The genome is not only genes. It is genes plus the sequences that decide when and how much each gene is transcribed: promoters, enhancers, the boundaries between introns and exons, and the genes coding for the transcription factors themselves. A change in any of those alters the protein's quantity or timing without altering its sequence — and quantity and timing are often what matters.
Three examples make the point better than a definition.
Lactase persistence. Most adult humans stop making lactase after weaning, which is the ancestral condition. The variant that keeps it switched on in people of northern European descent is a single base change, and it is not in the lactase gene. It lies about fourteen thousand base pairs away, inside an intron of a neighbouring gene, in a sequence that acts as an enhancer for lactase. The lactase protein of a lactose-tolerant adult is identical to everyone else's. All that changed was a switch.
β-thalassaemia. Many of the alleles causing it are not in the coding sequence of the β-globin gene at all. Some sit in the promoter and reduce transcription; others sit at an intron boundary, so the spliceosome cuts in the wrong place and the mature mRNA carries intron sequence. Either way the cell makes too little normal β-globin.
Tumours. A promoter that has been heavily methylated produces no protein at all from a gene that is entirely intact. That one is the subject of the next section, and it is the reason this distinction is in the specification rather than in a footnote.
- Mutation
- A change in the base sequence of DNA.
- Silent mutation
- A base substitution that produces a triplet coding for the same amino acid, so the primary structure of the polypeptide is unchanged.
- Frame shift
- The displacement of the reading frame caused by insertion or deletion of a number of bases that is not a multiple of three.
- Control sequence
- A region of DNA that is not translated but determines whether, when and how strongly a gene is transcribed.
Oncogenes and tumour suppressor genes
A tumour is a mass of cells produced by uncontrolled division. A benign tumour grows slowly, stays within a capsule, keeps cells that look much like the tissue it came from, and does damage only by pressing on something. A malignant tumour grows faster, is not contained, has cells that have lost the appearance and often the function of their tissue, and sheds cells into the blood and lymph that establish secondary tumours elsewhere. That last property, metastasis, is what makes cancer lethal rather than merely unpleasant.
Two classes of gene are involved, and they fail in opposite directions.
A proto-oncogene is a perfectly normal gene coding for a protein that stimulates cell division when a growth signal arrives — a growth factor, a receptor for one, or a protein that relays the signal inside the cell. A mutation can convert it into an oncogene: a version that is permanently active, or one that is present in too many copies, or one that has been moved next to a promoter that never switches off. The cell then receives a constant instruction to divide whether or not any growth factor is present. One faulty allele is enough, because a permanently active protein does its damage regardless of the normal protein made by the other allele.
A tumour suppressor gene codes for a protein that does the opposite: it halts the cell cycle at a checkpoint, or triggers apoptosis in a cell whose DNA is damaged beyond repair. Losing it removes a brake, and here both alleles must go, because one working copy still makes the protein. That is why inherited predispositions behave as they do: someone born with one faulty BRCA1 allele does not have cancer, but every cell is one event from having no brake instead of the two events everybody else needs.
The route that matters for this unit is the one that involves no mutation. A tumour suppressor gene whose promoter becomes heavily methylated is transcribed less, or not at all, and the cell loses the brake exactly as if the gene had been deleted. Sequence the gene and it reads perfectly. This is common in real tumours — the mismatch repair gene MLH1 is silenced this way in a substantial fraction of colorectal cancers — and it explains why cancer is described as a disease of gene expression and not only a disease of mutation.
- Proto-oncogene
- A normal gene whose product stimulates cell division in response to a growth signal.
- Oncogene
- A mutated or over-expressed proto-oncogene whose product stimulates division continuously.
- Tumour suppressor gene
- A gene whose product slows the cell cycle or triggers apoptosis; loss of both alleles removes that control.
- Metastasis
- The spread of cells from a malignant tumour through the blood or lymph to form secondary tumours.
Oestrogen returns here, and it should be described with care. Breast tissue is a target for oestrogen, and in a cell where a proto-oncogene lies downstream of an oestrogen response element, more oestrogen means more transcription of a gene that stimulates division. After the menopause the ovaries stop producing oestrogen but adipose tissue continues to, and higher concentrations of oestrogen in post-menopausal women are associated with a higher risk of breast cancer. Notice the shape of that claim: an association, with a plausible mechanism behind it. It is not a claim that oestrogen causes breast cancer in the way that a mutation in a tumour suppressor does, and questions on this topic reward candidates who keep the distinction visible.
Reading methylation data from a tumour
Samples of healthy colon tissue and of tumour tissue from the same patient are analysed. The promoter of a tumour suppressor gene is found to be methylated at 8% of its cytosine sites in the healthy tissue and at 74% in the tumour. Sequencing shows the coding sequence of the gene is identical in both samples. Explain what has happened and why the cells divide uncontrollably.
The gene has not been mutated: the sequences are identical, so any answer built on a changed base is ruled out by the data given. What has changed is the methylation of the promoter, which has risen roughly ninefold.
A heavily methylated promoter cannot be bound by the transcription factors and RNA polymerase that would normally start transcription, and the methyl groups also attract proteins that recruit histone deacetylases, so the chromatin around the gene condenses further. Transcription falls to nothing, no mRNA is made, and the tumour suppressor protein is not synthesised.
Without that protein, the checkpoint it enforces in the cell cycle is not applied. Cells with damaged DNA are neither held for repair nor sent into apoptosis, so they continue through the cycle, divide, and pass their damage on. Division becomes uncontrolled and a tumour forms.
One further inference the data supports: because no base has changed, the silencing is in principle reversible, which is why drugs that inhibit methylation or deacetylation are used in some cancers. A deleted gene offers no such possibility.
Stem cells, and the argument that surrounds them
Cancer is a failure of control over division. Stem cell medicine is an attempt to use controlled division deliberately, and the two sit in the same unit for that reason. You met the levels of potency earlier in the course; what matters here is where the cells come from and what they are being used for.
| Source | Potency | Where it stands |
|---|---|---|
| Embryo, four to five days old | Pluripotent: any cell type of the body | Widest use in principle; the source of the ethical objection |
| Umbilical cord blood | Multipotent, mainly blood-forming | Routinely banked; used in the same way as bone marrow |
| Adult tissue, such as bone marrow | Multipotent, limited to that tissue's range | Established in treatment; supply and range both limited |
| Induced pluripotent, from the patient's own cells | Pluripotent, after reprogramming with transcription factors | No embryo used, no rejection; still largely at trial stage |
One use is genuinely routine: bone marrow transplantation replaces a patient's blood-forming stem cells after their own have been destroyed by leukaemia or its treatment, and has been standard care for decades. A second is licensed — sheets grown from a patient's own limbal stem cells restore the corneal surface after chemical burns. Beyond those, trials are running for macular degeneration, Parkinson's disease, type 1 diabetes and spinal cord injury, some with encouraging early results, and none of them is yet ordinary treatment. A question asking you to evaluate a stem cell claim is usually testing whether you know the difference between a trial and a treatment.
Now the ethics, which examiners want set out rather than settled. The disagreement is genuine, it is between thoughtful people, and it turns on a question biology cannot answer: what moral status a five-day-old human embryo has.
Those who oppose the use of embryonic stem cells generally hold that a human embryo has the full moral status of a person from fertilisation onwards, since that is when a genetically distinct human organism begins and no later point marks a non-arbitrary threshold. On that view, destroying an embryo to obtain its cells is the deliberate destruction of a human life, and the good that might be done with the cells does not license it — the Roman Catholic Church argues this position formally, and it is held by many people on secular grounds as well.
Those who support it generally hold that moral status develops gradually, and that an embryo of about a hundred cells with no nervous system, no capacity for experience and no individuality yet fixed does not have the interests that make killing a person wrong. On that view the suffering of identifiable patients with untreatable conditions carries more weight than the destruction of a blastocyst, particularly when the embryos used are surplus from IVF and would otherwise be discarded.
A middle position accepts the use of surplus IVF embryos while rejecting the creation of embryos specifically for research, on the ground that there is a moral difference between using something that will be destroyed anyway and bringing it into existence in order to destroy it. British law reflects a version of this compromise: research on human embryos requires a licence from the Human Fertilisation and Embryology Authority, and no embryo may be kept beyond fourteen days.
Induced pluripotent stem cells change the argument without ending it. They are made by treating an adult cell — usually a skin or blood cell — with a small set of transcription factors that switch previously silent genes back on, returning the cell to a pluripotent state. No embryo is involved and, because the cells come from the patient, tissue grown from them is not rejected. Those who object to embryo research welcome them for exactly that reason. But they are not free of difficulty: reprogrammed cells can carry mutations acquired during the adult cell's life, their tendency to form tumours has to be controlled, and the fact that they can in principle be turned into gametes raises questions of its own.
TRY IT — Evaluating a claim about stem cell treatment
A private clinic advertises an injection of the patient's own bone marrow stem cells as a treatment for multiple sclerosis, a disease in which the myelin sheaths around neurones are destroyed. The advertisement says that because the cells are the patient's own there is no ethical objection and no risk of rejection. Evaluate the claim biologically, and say what you would want to see before accepting it.
Check your answer
The rejection point is sound. Cells taken from the patient carry the patient's own antigens, so the immune system does not identify them as non-self and there is no rejection. The ethical point about embryos is also sound as far as it goes: no embryo is destroyed, so the objection raised against embryonic stem cells does not apply.
The biological problem is potency. Bone marrow stem cells are multipotent, and the range they can produce is the blood cells — red cells, the various white cells and platelets. They are not pluripotent, so there is no reason to expect them to produce the oligodendrocytes that make myelin in the central nervous system. The advertisement asserts a benefit without a mechanism.
The word 'ethical' is also doing hidden work. Absence of an embryo is not absence of ethical difficulty: selling an unproven treatment to people with an incurable disease raises questions about consent, about exploitation of hope and about the money involved.
Before accepting the claim you would want published results from randomised controlled trials with a placebo group, since multiple sclerosis relapses and remits on its own and testimonials cannot distinguish a treatment effect from a remission; a proposed mechanism by which the injected cells reach the central nervous system and produce myelin; and regulatory approval rather than an advertisement.
In the exam
- Name the mutation and then say what it does to the protein. 'A mutation occurs' scores nothing; 'a substitution produces a stop codon, so the polypeptide is shortened' scores twice.
- Silent mutations exist because the code is degenerate; frame shifts happen because it is non-overlapping. Name the property, not just the outcome.
- A deletion of three bases does not cause a frame shift. Check the number before you write 'frame shift' — questions set this trap deliberately.
- One faulty allele can be enough for an oncogene; both alleles must be lost for a tumour suppressor gene. Getting these the wrong way round undoes an otherwise complete answer.
- Hypermethylation of a promoter silences a gene without changing its sequence. If a question gives you methylation data and identical sequences, it is testing precisely that.
- For oestrogen and breast cancer, keep the language of association and mechanism separate. Data showing a correlation supports a hypothesis; it does not establish a cause.
- In an ethics question, state who holds each view and the reason they hold it. Marks are for the arguments, not for your conclusion, and a one-sided answer caps itself.
Check yourself
A gene codes for a protein 480 amino acids long. In one patient a single base is deleted at base 30. In a second, a single base is substituted at base 1437, near the end of the coding sequence. In a third, three bases are deleted at position 900. Predict which of the three is likely to be most severely affected, and explain your reasoning fully.
Answer
The patient with the single base deletion at base 30 is likely to be worst affected by a wide margin.
Base 30 is inside the tenth triplet, so nine amino acids are translated normally and everything after that is read in a shifted frame. Because the code is non-overlapping and read in threes from a fixed start, removing one base pulls every subsequent base one place forward, so all of the remaining amino acids are potentially wrong. A shifted frame also throws up stop codons at random, so the polypeptide is usually cut short as well as scrambled. A protein of nine correct amino acids followed by nonsense will not fold into anything, and the cell effectively has no product from that allele.
The substitution at base 1437 falls in the 479th triplet, near the very end of a 480-residue protein, and changes one amino acid. That may do nothing at all: the code is degenerate, so the new triplet may specify the same amino acid, and even if it does not, a single change at the end of a chain rarely disturbs an active site or a binding site. The most likely outcome is a protein that works normally.
The three-base deletion at position 900 removes exactly one amino acid and shifts nothing, because three is a multiple of three. Every triplet after it is read as before. The result is a protein 479 amino acids long with the rest of its sequence intact. It may still be faulty — the commonest cystic fibrosis allele is precisely this kind of mutation and produces a protein the cell fails to deliver to the membrane — but it is a protein, and it is far closer to normal than a frame-shifted one.
The ranking is therefore: frame shift near the start, worst; three-base deletion in the middle, intermediate; substitution near the end, probably harmless. What decides severity is how much of the protein is affected and where, not how many bases changed.
Questions
Question 14 marks
Explain why the deletion of a single base near the start of a gene usually destroys the protein, while the deletion of three bases at the same point usually does not.
Mark scheme
- B1 the code is non-overlapping and read in triplets from a fixed starting point
- B1 losing one base moves every base after it one place forward, so the reading frame is shifted and every triplet downstream is read from a new position
- B1 all the amino acids after the deletion are therefore potentially wrong, and a stop codon usually appears early by chance, so the polypeptide is shortened as well as scrambled and will not fold into anything
- B1 three is a multiple of three, so the frame is not shifted: one amino acid is removed and every triplet after it is read exactly as before
Question 24 marks
Compare a proto-oncogene with a tumour suppressor gene, referring to what the product of each normally does and to how many alleles must be affected before division is lost from control.
Mark scheme
- B1 a proto-oncogene codes for a protein that stimulates cell division when a growth signal arrives, whereas a tumour suppressor gene codes for a protein that halts the cell cycle at a checkpoint or triggers apoptosis
- B1 a proto-oncogene does harm when a mutation makes it permanently active or present in too many copies, whereas a tumour suppressor gene does harm when its product is lost
- B1 one faulty allele is enough for an oncogene, because a permanently active protein acts whatever the other allele makes, whereas both alleles of a tumour suppressor gene must go, because one working copy still makes the protein
- B1 in both cases the outcome is the same: cells pass checkpoints they should not and divide repeatedly, so a mass of cells accumulates
Question 34 marks
A research group wishes to obtain stem cells from embryos left over from IVF treatment. Evaluate the case for allowing this work.
Mark scheme
- B1 for: embryonic stem cells are pluripotent and can give rise to any cell type of the body, so they offer a range of treatment that multipotent adult stem cells cannot
- B1 for: the embryos are surplus from IVF and would otherwise be discarded, and many people hold that there is a moral difference between using such an embryo and creating one in order to destroy it
- B1 against: those who hold that a human embryo has the full moral status of a person from fertilisation onwards regard its destruction as the deliberate ending of a human life, which the good done with the cells does not license
- B1 judgement: British law takes a middle position, licensing such research through the Human Fertilisation and Embryology Authority up to fourteen days; induced pluripotent cells narrow the disagreement, since they are pluripotent and need no embryo, though they can carry mutations acquired in the adult cell and a tendency to form tumours
Question 43 marks
A drug that inhibits the enzymes adding methyl groups to DNA is being trialled in patients whose tumours carry an intact but heavily methylated tumour suppressor gene. Suggest why the drug might restore control of division in those cells, and suggest one reason it may also harm healthy cells.
Mark scheme
- B1 with fewer methyl groups on the promoter, transcription factors and RNA polymerase can bind to it again, so the gene is transcribed and mRNA is made
- B1 the tumour suppressor protein is then synthesised and the checkpoint it enforces is applied again, so cells with damaged DNA are held for repair or sent into apoptosis instead of dividing; no base had changed, so nothing needed repairing
- B1 the drug reduces methylation across the whole genome and in every cell it reaches, so genes that were properly silenced in healthy cells are switched on as well
Question 52 marks
Name the kind of substitution that produces a stop codon, and name the property of the genetic code that allows a different substitution to change no amino acid at all.
Mark scheme
- B1 a nonsense substitution, which ends translation early
- B1 the code is degenerate, so more than one triplet codes for the same amino acid
Worth remembering
- Substitutions affect one triplet; insertions and deletions that are not multiples of three shift the whole reading frame.
- Silent, missense and nonsense are the three outcomes of a substitution, and each has a distinct effect on the polypeptide.
- A mutation in a control sequence changes how much protein is made without changing what the protein is.
- Oncogenes are proto-oncogenes stuck on; tumour suppressor genes are brakes that have been lost, and both alleles must go.
- Methylating the promoter of a tumour suppressor gene silences it without altering a single base.
- Bone marrow and limbal stem cell treatments are established; most other stem cell therapies are still in trials.