Cellular Control

A mutation is a random change to the genetic material. Some mutations involve changes to the structure or number of chromosomes. A gene mutation is a change to the DNA. 

Mutations may occur spontaneously during DNA replication before cell division. Certain chemicals, such as tar in tobacco smoke, and ionising radiation such as UV light, X-rays and gamma rays, may be mutagenic. 

The structure of the DNA molecule makes it stable and fairly resistant to corruption of the genetic information stored within it. Errors may occur, however, during the replication of a DNA molecule. Mutations associated with mitotic division are somatic mutations and are not passed to offspring. They may be associated with the development of cancerous tumours. 

Mutations associated with meiosis and gamete formation may be inherited by offspring. 

Gene mutations may affect protein production and function. There are two main classes of DNA mutation:

• Point mutation: one base pair replaces (is substituted for) another. 
• Insertion of deletion (indel) mutation: one or more nucleotides are inserted or deleted from a length of DNA. These may cause a frameshift. 

The genetic code consists of nucleotide base triplets within the DNA. During transcription of a gene, this code is copied to a length of mRNA as codons, complementary to the base triplets on the template strand of the length of DNA.

The sequence of codons on the mRNA is therefore a copy of the sequence of base triplets on the gene (coding strand of the DNA). There are three types of point mutation:

• Silent 
• Missense 
• Nonsense 

Apart from methionine, all the amino acids involved in protein synthesis, have more than one base triplets code. This reduces the effect of point mutations, as they do not always cause a change to the sequence of amino acids in a protein. This is often called the 'redundancy' or 'degeneracy' of genetic code.

A point mutations involving a change to the base triplet, where that triplet still codes for the amino acid, is a silent mutation. The primary structure of the protein, and therefore the secondary and tertiary structure, is not altered. 

A change to the base triplet sequence that leads to a change in the amino acid sequence in a protein is a missense mutation. Within a gene, such a point mutation may have a significant effect on the protein produced. The alteration to the primary structure leads to a change to the tertiary structure of the protein, altering its shape and preventing it from carrying out its usual function. 

Sickle cell anaemia results from a missense mutation on the sixth base triple of the gene for the B-polypeptide chains of haemoglobin: the amino acid valine, instead of glutamic acid, is inserted at this position. This results in deoxygenated haemoglobin crystallising within erythrocytes, causing them to become sickle shaped, blocking capillaries and depriving tissues of oxygen. 

A point mutation may alter a base triplet, so that it becomes a termination (stop) triplet. This particularly distributive point mutation results in a truncated protein that will not function. This abnormal protein will most likely be degraded within the cell. The genetic disease Duchenne muscular dystrophy is the result of a nonsense mutation.

If nucleotide base pairs, not in multiples of three, are inserted in the gene or deleted from the gene, because the code is non-overlapping and read in groups of three bases, all the subsequent base triplets are altered. This is a frameshift.

When the mRNA from such a mutated gene is translated, the amino acid sequence after the frameshift is severely disrupted. The primary device of the protein, and subsequently the tertiary structure, is much altered. Consequently, the protein cannot carry out its normal function. If the protein is very abnormal, it will be rapidly degraded within the the cell. 

Some forms of thalassaemia, a haemoglobin disorder, result from frame shifts due to deletions of nucleotide bases. 

Insertions or deletions of a triplet of base pairs result in the addition or loss of an amino acid, and not in a frameshift. 

Some genes contain a repeating triplet such as -CAG CAG CAG-. In an expanding triplet nucleotide repeat, the number of CAG triplets increases at meiosis and again from from generation to generation. 

Huntington disease results from an expanding triplet repeat. If the number of repeating CAG sequences goes above a certain critical number, then the person with that genotype will develop the symptoms of Huntington disease later in life. 

Many mutations are beneficial and have helped to drive evolution through natural selection. Different alleles of a particular gene are produced via mutation. 

The mutation that gave rise to blue eyes arose in the human population 6000-8000 years ago. Such a mutation may be harmful in areas where sunlight intensity is high, as the lack of iris pigmentation could lead to lens cataracts. However, in more temperate zones, it could enable people to see better in less bright light. 

Early humans in Africa would have had black skin, the high concentrations of melanin protecting them from sunburn and skin cancer. When humans migrated to temperate regions, a paler skin would be an advantage, enabling vitamin D to be made with a lower intensity of sunlight. In such areas, people with fairer skin would have an advantage and be selected, as vitamin D not only protects us from rickets, it protects us from heart disease and cancer. 

Some mutations appear to be neutral, being neither beneficial or harmful, such as those that, in humans, cause the inability to smell certain flowers, including freesias and honeysuckle and differently shaped ear lobes.

Enzymes that catalyse the metabolic reactions involved in basic cellular functions are synthesised at a fairly constant rate. Enzymes that may only be needed under specific conditions are synthesised at varying rates, according to the needs of the cell.

The bacterium E.coli normally metabolises glucose as respiratory substrate. However, if glucose is absent and the disaccharide lactose is present, lactose induces the production of two enzymes:

• Lactose permease, which allows lactose to enter the bacterial cell
• B-galactosidase, which hydrolyses lactose to glucose and galactose.

The lac operon consists of a length of DNA, about 6000 base pairs long, containing an operator region (lacO) next to the structural genes lacZ and lacY that code for the enzymes B-galactose and lactose permease, respectively.

Next to the operator region, lacO, is the promotor region, P, to which the enzyme RNA polymerase binds to begin transcription of the structural genes lacZ and lacY.

The operator region and promoter region are the control sites.

A small distance away from the operon is the regulatory gene, I, that codes for a repressor protein (LacI). When this regulatory gene is expressed, the repressor protein produced binds to the operator, preventing RNA polymerase from binding to the promoter region. The repressor protein therefore prevents the genes lacZ and lacY from being transcribed (expressed), so the enzymes for lactose metabolism are not made. The genes are ‘off’.

When lactose is added to the culture medium, once all the glucose has been used, molecules of lactose bind to the LacI repressor protein molecules; this alters the shape of the LacI repressor protein, preventing it from binding to the operator. The RNA polymerase enzyme can then bind to the promotor region and begin transcribing the structural genes into mRNA that will then be translated into the two enzymes. Thus lactose induces the enzymes needed to break it down.

Every cell in a eukaryotic organism has the same genome but, because different cells use it differently, they function differently. This is the basis of cell differentiation. In neurones, the genes being expressed differ to some extent from those being expressed in a liver or kidney cell, although all cells express the basic ‘housekeeping’ genes.

Transcription factors are proteins, or short non-coding pieces of RNA, which act within the cell’s nucleus to control which genes in a cell are turned on or off. Transcription factors slide along a part of the DNA molecule, seeking and binding to their specific promotor regions. They may then aid or inhibit the attachment of RNA polymerase to the DNA, and active or suppress transcription of the gene. They are essential for the regulation of gene expression in eukaryotes, making sure that different genes in different types of cells are activated or suppressed. Some transcription factors are involved in regulating the cell cycle. Tumour suppressor genes and proto-oncogenes help regulate cell division via transcription factors. Mutations to these genes can lead to uncontrolled cell division or cancer.

About 8% of genes in the human genome encode transcription factors. Many genes have their promoter regions some distance away, along the unwound length of DNA but, because of how the DNA can bend, the promotor region may not be too far away spatially.

Within a gene there are non-coding regions of DNA called introns, which are not expressed. They separate the coding or expressed regions, which are called exons.

All the DNA of a gene, both introns and exons, is transcribed. The resulting mRNA is called primary mRNA. Primary mRNA is then edited and the RNA introns – lengths corresponding to the DNA introns – are removed. The remaining mRNA exons, corresponding to the DNA exons, are joined together. Endonuclease enzyme may be involved in the editing and splicing processes.

Some introns may themselves encode proteins, and some may become short non-coding lengths of RNA involved in gene regulation. Some genes can be spliced in different ways. A length of DNA with its introns and exons can, according to how it is spliced, encode more than one protein.

Post-translational regulation of gene expression involves the activation of proteins. Many enzymes are activated by being phosphorylated. Cyclic AMP (cAMP), is an important second messenger involved in this activation.

1. A signalling molecule, such as the protein hormone glucagon, binds to a receptor on the plasma membrane of the target cell. 

2. This activates a transmembrane protein which then activates a G protein.

3. The activated G protein activates adenyl cyclase enzymes.

4. Activated adenyl cyclase enzymes catalyse the formation of many molecules of cAMP from ATP.

5. cAMP activates PKA (protein kinase A).

6. Activated PKA catalyses the phosphorylation of various proteins, hydrolysing ATP in the process. This phosphorylation activates many enzymes in the cytoplasm, for example those that convert glycogen to glucose.

7. PKA may phosphorylate another protein (CREB, cAMP response element binding).

8. This then enters the nucleus and acts as a transcription factor, to regulate transcription.

The large and ancient family of the genes, the homeotic genes, are involved in controlling the anatomical development, or morphogenesis, of an organism, so that all structures develop in the correct location, according to the body plan. Several of these genes contain Homeobox sequences, and they are sometimes called Homeobox genes.

Each Homeobox sequence is a stretch of 180 DNA base pairs (excluding introns) encoding a 6—amino acid sequence, called a homeodomain sequence, within a protein. The homeodomain sequence can fold into a particular shape and bind to DNA, regulating the transcription of adjacent genes. These proteins are transcription factors and act within the cell nucleus. The shape that these homeodomain-containing proteins fold into it called H-T-H. It consists of 2 alpha-helices (H) connected by one turn (T). Part of the homeodomain amino acid sequence recognises the TAAT sequence of the enhancer region (a region that initiates or enhances transcription) of a gene to be transcribed.

Homeobox genes are highly conserved across different phyla including the arthropods (such as fruit flies) and vertebrates (such as mice and humans), so these sequences have remained little changed since the Cambrian explosion of animal body plans some 500 million years ago. 

Highly conserved regions typically indicates that natural selection has continually eliminated forms with mutations in that sequence which suggests that any altercations to the sequence causes death in an early stage of life or the inability to reproduce.

The Hox genes regulate the development of embryos along the anterior-posterior (head-tail) axis. They control which body parts grow where. If Hox genes are mutated, abnormalities can occur. Hox genes are arranged in clusters and each cluster may contain up to 10 genes. In tetrapods (four-limbed vertebrates) including mammals and therefore humans, there are four clusters. At some stage during evolution, the Hox clusters have been duplicated.

In early embryonic development, Hox genes are active and are expressed in order along the anterior-posterior axis of the developing embryo. The sequential and temporal (in time) order of the gene expressions corresponds to the sequential and temporal development of the various body parts, a phenomenon known as collinearity.

Hox genes encode homeodomain proteins that act in the nucleus as transcription factors and can switch on cascades of activation of other genes that promote mitotic cell division, apoptosis, cell migration and also help to regulate the cell cycle.

Hox genes are similar across different classes of animals; a fly can function properly with a chicken Hox gene inserted in place of its own. This is because in plants, animals and fungi have similar Homeobox gene sequences which are highly conserved.

Hox genes are regulated by other genes called gap genes and pair-rule genes. In turn, these genes are regulated by maternally supplied mRNA from the egg cytoplasm.

From zygote to embryo to fully formed adult, there are many mitotic cell divisions. Mitosis is part of the cell cycle that is regulated with the help of Homeobox and Hox genes. It ensures that each new daughter cell contains the full genome and is a clone of the parent cell.

During cell differentiation some of the genes in a particular type of cell are ‘switched off’ and not expressed.

In 1962, Leonard Hayflick showed that normal body cells divide a limited number of times (around 50 times – known as the Hayflick constant) before dying.

Apoptosis is programmed cell death.

1. Enzymes break down the cell cytoskeleton

2. The cytoplasm becomes dense with tightly packed organelles

3. The cell surface membrane changes and small protrusions called blebs form

4. Chromatin condenses, the nuclear envelope breaks and DNA breaks into fragments

5. The cell breaks into vesicles that are ingested by phagocytic cells, so that cell debris does not damage any other cells or tissues. The whole process happens quickly.

Many cell signals help to control apoptosis. Some of these signalling molecules may be released by cells when genes that are involved in regulating the cell cycle and apoptosis respond to internal cell stimuli and external stimuli such as stress. These signalling molecules include cytokines from cells of the immune system, hormones, growth factors and nitric oxide. Nitric oxide can induce apoptosis by making the inner mitochondrial membrane more permeable to hydrogen ions and dissipating the proton gradient. Proteins are released into the cytoplasm where they bind to apoptosis inhibitor proteins, allowing apoptosis to occur.

Apoptosis is an integral part of plant and animal tissue development. Extensive proliferation of cell types is prevented by pruning through apoptosis, without release of any hydrolytic enzymes that could damage surrounding tissues.

During limb development, apoptosis causes the digits to separate from each other. Apoptosis removed ineffective or harmful T-lymphocytes during the development of the immune system. In children aged between 8 and 14, 20-30 billion cells per day apoptose; in adults about 50-70 million cells per day apoptose.

The rate of cells dying should equal the rate of cells produced by mitosis:

• Not enough apoptosis leads to the formation of tumours

• Too much apoptosis leads to cell loss and degeneration.

Cell signalling plays a crucial role in maintaining the right balance.