GENE EXPRESSION

20.1 Mutation – any change to the quantity of structure of DNA in an organism that arises during the replication of DNA. Gene mutation – any change to or more nucleotides bases or any rearrangement of bases.

Types of gene mutations:- Substitution of bases – when a nucleotide in a section of DNA molecule is replaced by

another nucleotide that has a different base. There are three possible consequences from this: Formation of one of the three stop codons that mark the end of a polypeptide chain.

The production of the polypeptide will be stopped prematurely. The final protein almost certainly would be different and so the protein would not able to perform its function.

The codon would code for a different amino acid, so the structure of a polypeptide would different in a single amino acid. This protein may have a different shape and so not function properly.

The codon is different but codes for the same amino acid, so the protein will function properly. This is because the code is degenerate. No effect.

- Deletion of bases – los of a nucleotide base from a DNA sequence. This creates a frame shift because the reading frame of triplets has been shifted to the left by one letter. Most triplets will be different, so the amino acid sequence (primary structure) and therefore the shape of polypeptide (tertiary or quaternary structure) would be altered. So it would be a non-functional protein that could considerably alter the phenotype. A deleted base at the start will have a greater impact than one near the end.

- Addition of bases – an extra base is inserted in the sequence, this also causes a frame shift and the whole sequence of triplets become altered after it. The frame shift is to the right. If three bases are added or any multiple of three bases, there will not be a frame shift. The polypeptide will be different, but not to the same extent.

- Duplication of bases – one or more bases are repeated, frame shift to the right. - Inversion of bases – a group of bases become separated from DNA sequence and re-join at

the same position but in the inverse order. - Translocation of bases – a group of bases become separated from the DNA sequence on one

chromosome and become inserted into the DNA sequence on a different chromosome. Translocation has significant effects of gene expression leading to an abnormal phenotype. This includes development of certain cancer forms and reduced fertility.

Gene mutations can arise spontaneously during DNA replication. Spontaneous mutations are permanent changes in DNA that occur without any outside influence. Mutations, though random, occur with a predictable frequency. Natural mutation varies from species to species, but is typically around one to two mutations per 100,000 genes per generation. Mutagenic agents or mutagens are outside factors that can increase this basic mutation rate. These include:

- High energy ionising radiation , e.g. alpha and beta particles, short wavelength radiation such as X-rays and ultra violet light. It can produce highly reactive agents, free radicals, in cells, which alter the shape of bases in DNA so that DNA polymerase can no longer act on them. Ultraviolet radiation affects thymine, causing it to form bonds with the nucleotides on either side, disrupting DNA replication.

- Chemicals such as nitrogen dioxide may directly alter DNA structure or interfere with transcription. Benzopyrene, a constituent of tobacco smoke, is a powerful mutagen that inactivates a tumour-suppressor gene TP53 leading to a cancer. Some chemical can remove groups from nucleotide bases, such as nitrous acid that removes –NH2 group from cytosine

in DNA, changing it into uracil. Or chemical can add groups to nucleotides Benzopyrene adds a large group to guanine that makes it unable to pair with cytosine. When DNA polymerase reaches the affected guanine it inserts any of other bases.

20.2 Cell differentiation – the process by which each cell develops into a specialised structure suited to the role they carry out.

Single-celled organisms perform all essential life functions but cannot be totally efficient in all of them as some function would require a different cellular structure.

In multicellular organisms, different cells are adapted for different role they perform for the organism to work efficiently overall. In early development its cells are not specialised – totipotent cells. They have all the genes, as all cells start from zygote and can become any specialised cell. But when the cells is mature and specialised, not all of its genes are switched on and therefore is not coding for every possible characteristic.

- Some genes are permanently expressed/ switch on in all cells, such as genes that code for enzymes are other proteins involved in respiration, translation, membrane synthesis, ribosomes and tRNA synthesis.

- Other genes are permanently not expressed/ switched off . - Some genes are switched on and off as and when they are needed.

To make proteins the cell does not require would be wasteful in terms of energy and resources. A variety of stimuli/controlling factors ensure that genes that not needed are not expressed. These include:

- Preventing transcription- Preventing translation

Specialisation is irreversible in most animal cells. Once the cells have matured and specialised they can no l0nger develop into other cells. However, non-specialise cells, stems cells can.

Stem cells – undifferentiated dividing cells that occur in adult animal tissue and need to be constantly replaced. So they have ability to divide to form an identical copy pf themselves in self-renewal process. They originate from:

- Embryotic stem cells – come from embryos in early stages of development- Umbilical cord blood stem cells – derived from umbilical cord blood, similar to adult stem

cells- Placental stem cells – found in placenta- Adult stem cells – found in body tissues of foetus through to adult; specific to particular

tissue or organ and produce cells to maintain and repair tissues.

Types of stem cells:- Totipotent stem cells – found in early embryo and can be differentiate into any type of cell.

A zygote is a totipotent cell. - Pluripotent stem cells – found in embryos and can differentiate into almost any type of cell.

They are slightly more specialised cells that the totipotent cells develop into as they divide and mature.

- Multipotent stem cells – these can differentiate into a limited number of specialised cell, includes adult stem cells and umbilical cord blood stem cells. For example, stem cells in the bone marrow can produce any type of blood cell.

- Unipotent stem cells – can only differentiate into a single type of cell. They are derived from multipotent stem cells and are made in adult tissue.

Induced pluripotent stem cells (iPS cells) – type of pluripotent cell that has been produced from unipotent stem cells. This is done by inducing genes and transcriptional factors to turn on genes that were previously off. One of the features of iPS is that they are capable of self-renewal, therefore can potentially divide indefinitely to provide a limitless supply. This can be used to replace embryotic stem cells in medical research and treatment and this would overcome the many ethical issues of using embryos in stem cell research.

Uses of pluripotent cells:Type of cell Disease that could be treatedHeat muscle cells Heart damage, e.g. result of a heart attackSceletal muscle cells Muscular dystrophyΒ cells of pancreas Type 1 diabetesNerve cells Parkinson’s disease, multiple sclerosis, strokes,

Alzheimer’s disease, paralysis due to spinal injury

Blood cells Leukaemia, inherited blood diseasesSkin cells Burns and woundsBone cells Osteoporosis Cartilage cells OsteoarthritisRetina cells of the eye Macular degeneration

20.3 Regulation of transcription and translation

For transcription to begin the gene is switched on by specific molecules that move from the cytoplasm into the nucleus – transcriptional factors. Each transcriptional factor has a site that binds to a specific base sequence of the DNA in the nucleus. When it does, that region of DNA begins the process of transcription. mRNA is produce and information is translated into a polypeptide. When a gene is not expressed the site where the transcription factor binds to is inactive. So no polypeptide can be synthesised from that gene.

Hormones like oestrogen can switch on a gene and so start transcription by combining with a receptor site on the transcriptional factor. Detailed process:

- Oestrogen is lipid-soluble molecule and so can diffuse through the cell-surface membrane- It binds to the site whose shape is complementary to oestrogen on a receptor molecule of

transcriptional factor- The oestrogen changes the shape of DNA binding site on the transcriptional factor, which

can now bind to DNA (it is activated), - The transcriptional factor enters the nucleus through a nuclear pore and bind to specific

base sequences on DNA- The combination of transcriptional factor with DNA stimulates transcription of that gene.

20.4 Epigenetics – a scientific field that looks at how environmental influences, such as diet stress, toxins, can subtly alter the genetic inheritance of an organism’s offspring.

Epigenome – chemical tags that cover the DNA and histones. The epigenome determines the shape of the DNA-histone complex. It can keep genes inactive in a tightly packed arrangement and so ensures that they cannot be read i.e. keep them switched off. This is called epigenetic silencing. It can also switch them on buy unwrapping active genes, making them exposed and easily transcribed.

The DNA code is fixed but the epigenome is flexible. The tags can be switched off/inhibit and on/activate with response to environmental factors, such as diet and stress. Epigenome of a cell – accumulation of signal it has received during its lifetime and it therefore acts like a cellular memory.

Environmental signal stimulates proteins to carry its message inside the cell from where it is passed by a series of other protein into the nucleus. Here the message passes to a specific protein which can be attached to a specific sequence of bases of the DNA. This has two possible effects – acetylation of histones which leads to activation or inhibition of a gene; or methylation of DNA by attracting enzymes that can add or remove methyl groups.

Acetylation – process whereby an acetyl group is transferred to a molecule. In this case this is acetylcoenzyme A. Deacetylation – process whereby an acetyl group is removed from a molecule.

Methylation – addition of a methyl group, CH3 to a molecule. In this case the methyl group is added to the cytosine bases of DNA.

Switched on/activated – the association of histones with DNA is weak, making it less condensed/loosely packed. DNA is accessible by transcription factors that instated the production of mRNA.

Switched off/inhibited – the association between histones and DNA is strong, making it more condensed/tightly packed. DNA is no accessible by transcription factors and so cannot initiate production of mRNA.

Condensation of the DNA-histone complex is caused by deacetylation of the histones or by methylation of DNA.

- Deacetylation increases positive charges on histones and so increases their attraction to the phosphate groups of DNA. The association becomes stronger and so DNA becomes more condensed.

- Methylation inhibits transcription by preventing the biding of transcriptional factors to the DNA and attracting proteins that condensed the DNA-histone complex (by inducing deacetylation of the histones) making the DNA inaccessible to transcription factors.

There is lots of evidence to support that epigenetic inheritance takes place. For example, experiments on rats showed that those female rats who received good care when young respond better to stress in later life and nurture their offspring better. In humans, when a mother has a condition of gestational diabetes, the foetus is exposed to high concentrations of glucose. This cause epigenetic changes in the daughter’s DNA, increasing the likelihood she will develop the disease herself.

It is thought that in sperm and eggs during the earliest stages of development, a specialised cellular mechanism searches the genome and erases its epigenetic tags in order to return the ells to a genetic ‘clean state.’ But some tags escape this process and are passed to offspring.

Some changes are responsible for certain diseases. Alternations can causes abnormal activation or silencing of genes. This included cancer.

Epigenetic changes do not alter the sequence of DNA. But they can, however, increase the incidence of mutations. Some active genes normally help repair DNA and so prevent cancers. In people with various types of inherited cancer, it is found that increased methylation of these genes has led to these protective genes being switched off. As a result, damaged sequences are not repaired and can lead to cancer.

To treat cancer, you can use epigenetic treatments to counteract these changes. These treatments use drugs to inhibit certain enzymes involved in either histone acetylation of DNA methylation. Epigenetic therapy must be specifically targeted on cancer cells. If the drugs were to affect normal cells they could activate gene transcription and make them cancerous. Another use of epigenetics in disease treatment has been the development of diagnostic tests that help to detect the early stages of diseases such as cancer, brain disorders and arthritis.

In eukaryotes and some prokaryotes the translation of mRNA produced by a gene can be inhibited by breaking mRNA down before its coded information can be translated into a polypeptide. One type of small RNA molecule that may be involved is small interfering RNA (siRNA).

- An enzyme cuts large double-stranded molecules of RNA into smaller sections called small interfering RNA (siRNA).

- One of the two siRNA strands combines with an enzyme. - The siRNA molecule guides the enzyme to a messenger RNA molecule by pairing up its bases

with the complementary ones on a section of mRNA molecule. - Once in position, the enzyme cuts mRNA into smaller sections. - The mRNA is no longer capable of being translated into a polypeptide. - The gene is not expressed i.e. blocked.

20.5 Cancer – a group of diseases caused by damage to the genes that regulate mitosis and the cell cycle. This leads to unrestrained growth of cells. A tumour – a group of abnormal cells – develops and constantly expands in size. Cancer is a common and destructive disease. Cancer is to some extent avoidable and if diagnosed early, successfully treatable.

Malignant are cancerous tumours.Benign are non-cancerous tumours.

Benign tumours Malignant tumoursCan grow to a large size Can grow to a large sizeGrow very slowly Grow rapidlyCell nucleus has a relatively normal appearance Cell nucleus is often large and appears darker

due to an abundance of DNACells are often well differentiated (specialised) Cells become de-differentiated (unspecialised)Cells produce adhesion molecules that make them stick together and so they remain within the tissue from which they arise = primary tumours

Cells do not produce adhesion molecules and so they tend to spread to other regions of the body, the process called metastasis, forming secondary tumours.

Tumours are surrounded by a capsule of dese tissue and so remain as a compact structure

Tumours are not surrounded by a capsule and so can grow finger-like projections in the surrounding tissue

Much less likely to be life-threatening but can disrupt functioning of a vital organ

More likely to be life-threatening, as abnormal tumour tissue replaces normal tissue

Tend to have localised effects on the body Often have systemic (whole body) effects such as weight loss and fatigue

Can usually be removed by surgery alone Removal usually involves radiotherapy and/or chemotherapy as well as surgery

Rarely reoccur after treatment More frequent reoccur after treatment

Most oncogenes are mutations of proto-oncogenes. Proto-oncogenes stimulate a cell to divide when growth factors attach to a protein receptor on its cell-surface membrane. This then activates genes that cause DNA to replicate and the cell to divide. If a proto-oncogene mutates into an oncogene it can become permanently activated (switched on) for two reasons:

- The receptor protein on the cell-surface membrane is permanently activated- The oncogene may code for a growth factor that is then produced excessive amounts

The result is that cells divide too rapidly and out of control and a tumour or cancer develops. Most cancer-causing mutations involving oncogenes are acquired, but a few cancers are caused by inherited mutations.

Tumour suppressor genes slow down cell division, repair mistakes in DNA and ‘tell’ cell when to die – apoptisis, programmed cell death. They maintain normal rates of cell division and prevent formation of tumour. If a tumour suppressor gene is mutated it is inactivated/switched off. As a result the cell can grow out of control. The mutated cells are usually structurally and functionally different from normal cells. While most of these die, those that survive can make clones of themselves and form tumours. There are a number of tumour suppressor genes including TP53, BRCA1, BRCA2. TP53 is involved in the process of apoptosis. This process is activated when a cell is unable to repair DNA.

Some cancers are caused by inherited mutations of tumour suppressors genes but most are acquired, not inherited.

Oncogenes cause cancer as a result of activation of proto-oncogene; tumour suppressor genes cause cancer when they are inactivated.

Abnormal DNA methylation is common in the development of a variety of tumours. The most common abnormality is hypermethylation. The process of leading to it:

- Hypermethylation occurs in a specific region, promoter region, of tumour suppressor genes. - This leads to the tumour suppressor gene being inactivated. - Transcription of the promoter regions of tumour suppressor genes is inhibited- The tumour suppressor gene is therefore silenced. - Cell division is increasing and causes a formation of a tumour.

Abnormal methylation of this type is thought to occur in a BRCA1 and leads to the development of breast cancer.

Another form of abnormal methylation is hypomethylation. This has been found to occur in oncogenes where it leads to their activation.

Oestrogen play a central role in regulating the menstrual cycle, however an increase in oestrogen concentrations can increase risk in developing breast cancer. It is more likely to occur after the menopause as the fat cells of the breasts tend to produce more oestrogens after the menopause. This appears to trigger breast cancer in postmenopausal women. When the tumour has develop, this leads to increased development of the tumour. It also appears that white blood cells that are drawn to the tumour increase oestrogen production. This leads to even greater development of the tumour. Oestrogen also causes proto-oncogenes of cells in breast tissue to develop into oncogenes, which leads to development of tumour (breast cancer).

Two-hit hypothesis – it takes a single mutated allele to activated proto-oncogenes but it takes a mutation of both alleles to inactivate tumour suppressor genes.

Bioinformatics – science of collecting and analysing complex biological data such as genetic codes. It uses computers to read, store and organise biological data at much faster rate than previously. Uses algorithms.

Whole-genome shotgun (WGS) sequencing – technique to determine the genome. It involves cutting DNA into small, easily sequence sections and then using computer algorithms to align overlapping segments to assemble the entire genome.

Benefits: